INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(14598359, 'Rutkowski P, Kamińska J, Kowalska M, Ruka W, Steffen J', 'Cytokine and cytokine receptor serum levels in adult bone sarcoma patients: correlations with local tumor extent and prognosis.', 'J Surg Oncol', '2003', 'Cytokine and cytokine receptor serum levels in adult bone sarcoma patients: correlations with local tumor extent and prognosis. We analyzed the correlations between pretreatment serum levels of 11 cytokines and soluble cytokine receptors (interleukin 6 (IL-6); interleukin 8 (IL-8); interleukin 10 (IL-10); vascular endothelial growth factor (VEGF); basic fibroblast growth factor (bFGF); macrophage colony-stimulating factor (M-CSF); granulocyte colony-stimulating factor (G-CSF); interleukin 1 receptor antagonist (IL-1ra); sIL-2Ralpha; tumor necrosis factor receptor I (TNF RI), and TNF RII) with clinico-pathological features and survival of patients with bone sarcomas.Altogether, 72 patients with bone sarcomas without distant metastases before treatment (26 osteosarcomas-36%, 23 chondrosarcomas-32%, 13 Ewing''s sarcomas/PNET-18%, 10 giant-cell tumors-14%), 22 patients with benign non-inflammatory bone tumors and 50 age-matched healthy controls were included into this prospective study.Median serum levels of 9/11 cytokines, with the exception of sIL-2Ralpha and G-CSF, were significantly higher in sarcoma patients than in controls. Median serum levels of IL-6, IL-8, IL-1ra, TNF RI, and M-CSF were significantly higher in patients with bone sarcoma as compared to patients with benign bone tumors. In 45.9% of sarcoma patients, six or more cytokines and cytokine receptors, including those that are involved in bone destruction (e.g., IL-6 and IL-8) and bone formation (e.g., IL-1ra and TNFRI and TNFRII), were elevated in parallel. Serum levels of IL-6, IL-8, TNF RI, TNF RII, and VEGF correlated significantly with tumor size (<10 cm vs. >or=10 cm in diameter) and serum levels of IL-6, IL-8, TNF RI, and IL-1ra correlated significantly with local tumor extent (E2/4 vs. E5/6 according to the classification proposed by Spanier et al. 46). Moreover, serum levels of IL-1ra and IL-6 were significantly higher in patients with small tumors (<5 cm in diameter) infiltrating structures adjacent to the periosteum (E5/6) than in large tumors (>10 cm in diameter) but confined to the bone and periosteum (E < 4). The lowest median serum levels of 8/11 cytokines/cytokine receptors were found in patients with giant-cell tumors. In an univariate analysis, increased serum levels of IL-1ra, IL-6, IL-8, IL-10, sIL-2Ralpha, M-CSF, TNF RI, and TNF RII, the number of cytokines elevated, higher tumor grade, larger tumor size, greater local extent (E) and patients'' age >35 years correlated with poor overall survival (OS) (P < 0.05). Similarly, high serum levels of IL-1ra, IL-6, TNF RI and TNF RII, tumor grade, tumor size, and tumor local extent (E) (P < 0.05) affected disease free survival (DFS) in univariate analysis. Multivariate analysis using Cox''s proportional hazards model showed that high serum levels of IL-1ra (P = 0.039) and TNF RI (P = 0.048), the number of serum cytokines above normal cut-off values (0-1 vs. 2-5 vs. >or=6; P = 0.029), greater tumor local extent E (E2/4 vs. E5/6; P = 0.02) correlated significantly with shorter OS. Only E was found as an independent prognostic factor for DFS (P = 0.04).These findings indicate that cytokines and soluble cytokine receptors, both physiologically involved in bone destruction and bone formation, have an essential role in the progression of malignant bone tumors.2003 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/14598359', 'yes', 'yes', 'NA', 9),
(14598886, 'Iwaya K, Ogawa H, Kuroda M, Izumi M, Ishida T, Mukai K', 'Cytoplasmic and/or nuclear staining of beta-catenin is associated with lung metastasis.', 'Clin Exp Metastasis', '2003', 'Cytoplasmic and/or nuclear staining of beta-catenin is associated with lung metastasis. Beta-catenin is involved in cell motility in the extracellular matrix, and is expressed in normal and neoplastic mesenchymal cells. In order to clarify whether beta-catenin expression in the cytoplasm and/or nucleus is associated with a propensity for pulmonary metastasis in osteosarcoma, the LM8 murine osteosarcoma cell line with a high metastatic potential to the lung was compared with original Dunn cells in terms of the beta-catenin expression level. Both osteosarcoma cell lines lost membrane localization of beta-catenin. However, beta-catenin gene had no mutation in exon 3 by direct sequence analysis. A large number of LM8 cells showed diffuse cytoplasmic and/or nuclear staining of beta-catenin (30.8 per high power field (HPF)), while a much smaller number of Dunn cells showed expression of beta-catenin (7.7 per HPF). Cells with positive staining of beta-catenin were frequently seen at the invasive front and in intravenous tumor deposits within the metastatic lesions to the lung. Thus, LM8 cells express a larger amount of the beta-catenin protein than Dunn cells, as judged by immunoblot analysis. In five resected cases of pulmonary metastasis, translocation of beta-catenin to the cytoplasm and/or nucleus of osteosarcoma cells was detected, although seven primary osteosarcomas cells that did not metastasize for more than five years did not show beta-catenin expression. These data indicate that the cytoplasmic and/or nuclear staining of beta-catenin is a biological marker of metastatic potential of osteosarcoma to the lung.', 'http://www.ncbi.nlm.nih.gov/pubmed/14598886', 'yes', 'yes', 'NA', 1),
(14610316, 'Dong YB, Yang HL, Elliott MJ, McMasters KM', 'Increased mdm-2 expression in a p53-independent manner blocks UV-induced cell cycle arrest and apoptosis in human osteosarcoma cells.', 'Tumour Biol', 'NA', 'Increased mdm-2 expression in a p53-independent manner blocks UV-induced cell cycle arrest and apoptosis in human osteosarcoma cells. DNA damage results in an increase in P53 levels, which is required to initiate a P53-mediated cell cycle arrest and/or apoptosis. P53 and MDM-2 form a feedback control loop: while P53 can transactivate the MDM-2 gene, high levels of MDM-2 inhibit P53 transactivation as well as promote rapid degradation of P53. In the present study, we investigated the interaction between endogenous MDM-2 and P53 following UV-induced DNA damage in an MDM-2 overexpression cell line. A human osteosarcoma cell line (OsACL, which contains wild-type P53 and overexpresses MDM-2 protein) was used in this study. Here we show that following UV treatment, P53 levels increased in the OsACL cells despite the presence of high-level endogenous MDM-2; however, CAT assays using a P53 reporter system revealed that this P53 was transcriptionally inactive. Although P53 transactivation was inhibited, MDM-2 levels rose markedly following UV irradiation. Northern blot analysis revealed that the increase in MDM-2 protein levels was a result of increased levels of MDM-2 mRNA, possibly due to increased transcription. Cell cycle analysis revealed that OsACL cells were markedly resistant to UV-induced apoptosis. Transfection of OsACL cells with an anti-sense MDM-2 plasmid dowregulated MDM-2 expression and increased UV-induced apoptosis. In conclusion, MDM-2 overexpression can block UV-induced cell cycle arrest and apoptosis by inhibiting P53 transcriptional activity. Furthermore, increased expression of MDM-2 in OsACL cells following UV irradiation appears to be related to P53-independent mechanisms.Copyright 2003 S. Karger AG, Basel', 'http://www.ncbi.nlm.nih.gov/pubmed/14610316', 'no', 'no', 'NA', 2),
(14624456, 'Nielsen-Preiss SM, Silva SR, Gillette JM', 'Role of PTEN and Akt in the regulation of growth and apoptosis in human osteoblastic cells.', 'J Cell Biochem', '2003', 'Role of PTEN and Akt in the regulation of growth and apoptosis in human osteoblastic cells. Cancer cells are characterized by either an increased ability to proliferate or a diminished capacity to undergo programmed cell death. PTEN is instrumental in regulating the balance between growth and death in several cell types and has been described as a tumor suppressor. The chromosome arm on which PTEN is located is deleted in a subset of human osteosarcoma tumors. Therefore, we predicted that the loss of PTEN expression was contributing to increased Akt activation and the subsequent growth and survival of osteosarcoma tumor cells. Immunoblot analyses of several human osteosarcoma cell lines and normal osteoblasts revealed relatively abundant levels of PTEN. Furthermore, stimulation of cell growth or induction of apoptosis in osteosarcoma cells failed to affect PTEN expression or activity. Therefore, routine regulation of osteosarcoma cell growth and survival appears to be independent of changes in PTEN. Subsequently, the activation of a downstream target of PTEN activity, the survival factor Akt, was analyzed. Inappropriate activation of Akt could bypass the negative regulation by PTEN. Analyses of Akt expression in several osteosarcoma cell lines and normal osteoblasts revealed uniformly low basal levels of phosphorylated Akt. The levels of phosphorylated Akt did not increase following growth stimulation. In addition, osteosarcoma cell growth was unaffected by inhibitors of phosphatidylinositol-3 kinase, an upstream activator of the Akt signaling pathway. These data further suggest that the Akt pathway is not the predominant signaling cascade required for osteoblastic growth. However, inhibition of PTEN activity resulted in increased levels of Akt phosphorylation and enhanced cell proliferation. These data suggest that while abundant levels of PTEN normally maintain Akt in an inactive form in osteoblastic cells, the Akt signaling pathway is intact and functional.Copyright 2003 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/14624456', 'yes', 'no', 'NA', 2),
(14629833, 'Ferrari S, Dondi M, Fanti S, Zoboli S, Giacomini S, Mercuri M, Bacci G', 'Somatostatin receptor (SSTR) scintigraphy in patients with osteosarcoma.', 'Cancer Biother Radiopharm', '2003', 'Somatostatin receptor (SSTR) scintigraphy in patients with osteosarcoma. To investigate somatostatin receptors (SSTRs) in primary or metastatic high-grade osteosarcoma, 18 patients (12 without metastases, 4 with synchronous lung, and 2 with metachronous bone metastases) underwent SSTR scintigraphy. After intravenous (i.v.) injection of 200-250 MPq of 111In-pentreotide, images were recorded at 4 and 24 hours. All patients underwent 99mTc-methylene diphosphonate (MDP) bone scan (BS). Scans were interpreted both with qualitative evaluation and with semiquantitative image processing, by means of tumor-to-background ratio modification over time. Primary tumor: SSTR scintigraphy was positive in 12 (75%) patients: 10 (83%) of the 12 nonmetastatic patients, and 2 (50%) of those with synchronous metastases. Metastases: SSTR scintigraphy was positive only in one (17%) of the six patients with metastatic disease. No relation was found between SSTR scintigraphy results and gender, histologic subtype, site, and size of the tumor. After primary chemotherapy, 15 patients underwent surgery of primary tumor; a good histologic response was found in 7 (64%) of the 11 patients with positive SSTR scintigraphy versus 1 (25%) of the 4 patients with negative SSTR scintigraphy. High-grade osteosarcoma exhibits somatostatin receptors, usually detectable in the primary tumor, but not in the metastatic lesions. The different expression of somatostatin receptors observed in primary and metastatic lesions, and in patients with tumors having different chemosensitivity, could indicate a possible relation between somatostatin receptors and biological behavior of the tumor.', 'http://www.ncbi.nlm.nih.gov/pubmed/14629833', 'yes', 'yes', 'NA', 1),
(14631380, 'Fellenberg J, Krauthoff A, Pollandt K, Delling G, Parsch D', 'Evaluation of the predictive value of Her-2/neu gene expression on osteosarcoma therapy in laser-microdissected paraffin-embedded tissue.', 'Lab Invest', '2004', 'Evaluation of the predictive value of Her-2/neu gene expression on osteosarcoma therapy in laser-microdissected paraffin-embedded tissue. Histologic response to chemotherapy is currently the strongest prognostic factor in high-grade osteosarcoma, but it can only be assessed after several weeks of therapy. Thus, detection of chemosensitivity at the time of diagnosis would be of great clinical importance. The expression of the proto-oncogene Her-2/neu has been shown to be of predictive value in breast cancer and has also been considered as prognostic marker for osteosarcomas, but reports of mainly immunohistochemical studies are controversial. Therefore, the aim of this study was to investigate Her-2/neu gene expression in laser-microdissected osteosarcoma cells. Laser microdissection enables the precise isolation of morphological defined cells from archival tissue specimens and is in combination with the highly sensitive real-time RT-PCR technique a valuable tool for cell-specific analysis of gene expression. Through optimization of current protocols, we could show that this technique can be successfully applied on formalin-fixed, paraffin-embedded and decalcified osteosarcoma tissue with high sensitivity and reproducibility. In all 17 osteosarcoma biopsies analyzed, we could detect Her-2/neu gene expression. Expression correlated significantly with the response to preoperative chemotherapy, which was assessed histologically according to the six-grade scale of Salzer-Kuntschik. Risk assessment on the basis of increased Her-2/neu gene expression matched the histologic findings in 16 out of 17 cases (94%). These data demonstrate the reliability of laser microdissection in the analysis of gene expression and suggest a possible role of Her-2/neu as prognostic marker for therapy outcome in osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/14631380', 'yes', 'yes', 'NA', 1),
(14633705, 'Matsuyama S, Iwadate M, Kondo M, Saitoh M, Hanyu A, Shimizu K, Aburatani H, Mishima HK, Imamura T, Miyazono K, Miyazawa K', 'SB-431542 and Gleevec inhibit transforming growth factor-beta-induced proliferation of human osteosarcoma cells.', 'Cancer Res', '2003', 'SB-431542 and Gleevec inhibit transforming growth factor-beta-induced proliferation of human osteosarcoma cells. Transforming growth factor-beta (TGF-beta) has growth-stimulating effects on mesenchymal cells and several tumor cell lines. The signaling pathway for this effect is, however, not well understood. We examined how TGF-beta stimulates proliferation of MG63 human osteosarcoma cells. Two distinct type I receptors for TGF-beta, ALK-1 and ALK-5, were expressed and functional in MG63 cells. Of these two receptors, ALK-5 appears to be responsible for the growth stimulation because expression of constitutively active ALK-5, but not ALK-1, stimulated proliferation of MG63 cells. SB-431542 (0.3 microM), a novel inhibitor of ALK4/5/7 kinase, suppressed TGF-beta-induced growth stimulation. DNA microarray analysis as well as quantitative real-time PCR analysis of RNAs from TGF-beta-treated cells demonstrated that several growth factors, including platelet-derived growth factor AA, were induced in response to TGF-beta in MG63 cells. Gleevec (1 microM) as well as AG1296 (5 microM) inhibited TGF-beta-induced growth stimulation of MG63 cells, suggesting that platelet-derived growth factor AA was mainly responsible for the growth-stimulatory effect of TGF-beta. We also examined the mechanisms of perturbation of growth-suppressing signaling in MG63 cells. We found that expression of c-Myc, which is down-regulated by TGF-beta in many other cells, was up-regulated in MG63 cells, suggesting that up-regulation of c-Myc expression may be the mechanism canceling growth-suppressing signaling of TGF-beta in MG63 cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/14633705', 'no', 'no', 'SB-431542|gleevec|AG1296', 4),
(14637192, 'Loennechen T, Mathisen B, Hansen J, Lindstad RI, El-Gewely SA, Andersen K, Maelandsmo GM, Winberg JO', 'Colchicine induces membrane-associated activation of matrix metalloproteinase-2 in osteosarcoma cells in an S100A4-independent manner.', 'Biochem Pharmacol', '2003', 'Colchicine induces membrane-associated activation of matrix metalloproteinase-2 in osteosarcoma cells in an S100A4-independent manner. Like the metastasis-associated protein S100A4, matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) are important in physiological and pathological conditions. Previously, we showed that S100A4 is involved in the regulation of MMPs and TIMPs, and in the present work we have investigated whether the anti-inflammatory and microtubule-disrupting drug colchicine has an effect on the expression of these proteins in osteosarcoma cell lines (OHS) with high and low levels of S100A4. Colchicine treatment of the various OHS cells resulted in an increased expression of MT1-MMP and TIMP-2 mRNA, and a corresponding increase of these two proteins in isolated cell membranes. Colchicine-treated cells produced more of the activated form of MMP-2 than control cells. However, the drug did not affect the amount of MMP-2 and TIMP-1 mRNA or protein, and it reduced the S100A4 mRNA expression. Isolated cell membranes from the colchicine-treated cells were more effective in activating exogenous proMMP-2 than membranes from control cells, and inhibitory studies indicated that it was the colchicine-induced increase in MT1-MMP that caused the increased activation of endogenous MMP-2. A peptide inhibitor of nuclear factor kappaB nuclear translocation, SN50, blocked the colchicine-induced activation of proMMP-2 and reduced the synthesis of MMP-2 in colchicine-treated cells, but not in control cells. It can be concluded that colchicine modulates the expression of MT1-MMP and TIMP-2 and hence the activation of proMMP-2 independently of the S100A4 level in osteosarcoma cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/14637192', 'no', 'no', 'colchicine|SN50', 5),
(14647441, 'Imai T, Adachi S, Nishijo K, Ohgushi M, Okada M, Yasumi T, Watanabe K, Nishikomori R, Nakayama T, Yonehara S, Toguchida J, Nakahata T', 'FR901228 induces tumor regression associated with induction of Fas ligand and activation of Fas signaling in human osteosarcoma cells.', 'Oncogene', '2003', 'FR901228 induces tumor regression associated with induction of Fas ligand and activation of Fas signaling in human osteosarcoma cells. We investigated the antitumor effects of FR901228, a HDAC inhibitor, on human osteosarcoma cells, in vitro and in vivo to explore its possible utility in the treatment of pediatric bone cancers. FR901228 caused marked growth inhibition with a 50% inhibitory concentration of 1.2-7.3 nM and induction of apoptosis in all eight osteosarcoma cell lines tested. These effects of FR901228 were also observed in vivo xenograft models on BALB/c nude mice, and treatment with 5.6 mg/kg/day resulting in a >70% reduction in the mean final tumor volume compared with the mean initial tumor volume. TUNEL assays demonstrated extensive apoptosis in tumor sections of mice treated with FR901228. Induction of apoptosis was preceded by increased expression of Fas ligand (FasL) mRNA, resulting in expression of membrane-bound FasL, which was followed by sequential activation of caspase-8 and -3. The level of apoptosis induction was reduced using a neutralizing anti-FasL antibody and overexpression of either the dominant-negative FADD or the viral FLICE inhibitory protein. Furthermore, treatment with a suboptimal dose of FR901228 greatly sensitized osteosarcoma cells to agonistic anti-Fas antibody-mediated apoptosis. These findings suggest that FR901228 is a highly promising antitumor agent against osteosarcoma, inducing apoptosis by the activation of the Fas/FasL system.', 'http://www.ncbi.nlm.nih.gov/pubmed/14647441', 'no', 'no', 'FR901228', 6),
(14660053, 'Liebau C, Merk H, Schmidt S, Roesel C, Karreman C, Prisack JB, Bojar H, Baltzer AW', 'Interleukin-12 and interleukin-18 change ICAM-I expression, and enhance natural killer cell mediated cytolysis of human osteosarcoma cells.', 'Cytokines Cell Mol Ther', '2002', 'Interleukin-12 and interleukin-18 change ICAM-I expression, and enhance natural killer cell mediated cytolysis of human osteosarcoma cells. Cytokines play important roles in the expression of adhesion molecules and the function of anti-tumor effector cells in the immune system. In this study, the influence of interleukin-12 (IL-12) and IL-18 on the expression of ICAM-1 and natural killer (NK)-cell mediated lysis in a human osteosarcoma cell line (HOS) was evaluated. ICAM-I expression of HOS cells were analyzed by flow cytometry following treatment with IL-12, IL-18 or both, and in co-cultures with peripheral lymphocytes. NK-cell activation in response to IL-12 and IL-18 was investigated by selective flow cytometry using propidium iodide. ICAM-1 expression on HOS cells was significantly enhanced by IL-12, but only when co-cultured in cell-to-cell contact with peripheral lymphocytes. Antibodies to interferon-gamma abrogated this effect. If HOS cells and peripheral lymphocytes were separated in co-cultures, IL-18 could substitute for cell-to-cell contact, facilitating IL-12-mediated enhancement of ICAM-1. Addition of IL-18 also enhanced NK-mediated cytolysis of HOS cells. These findings demonstrate that IL-12 can enhance the expression of ICAM-1 in the presence of IFN-gamma and, with IL-18, enhances NK anti-tumor activity. Immunomodulation via cytokine therapy may lead to improved eradication of chemotherapy-resistant osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/14660053', 'no', 'no', 'NA', 4),
(14669798, 'Nakano T, Tani M, Ishibashi Y, Kimura K, Park YB, Imaizumi N, Tsuda H, Aoyagi K, Sasaki H, Ohwada S, Yokota J', 'Biological properties and gene expression associated with metastatic potential of human osteosarcoma.', 'Clin Exp Metastasis', '2003', 'Biological properties and gene expression associated with metastatic potential of human osteosarcoma. Lung metastasis has a great influence on the prognosis of patients with osteosarcoma. We previously established two high-metastatic sublines, M112 and M132, from the HuO9 human osteosarcoma cell line by in vivo selection. In this study, we newly isolated a high-metastatic subline, H3, and three low-metastatic sublines, L6, L12 and L13, from HuO9 by the dilution plating method. Three high-metastatic sublines produced more than 200 metastatic nodules in the lung, while three low-metastatic sublines produced no or few nodules after injection of 2 x 10(6) cells into the tail vein of nude mice. There were significant differences in the motility and invasiveness between high- and low-metastatic sublines, whereas the growth rates in vitro and the tumorigenicity in vivo showed no correlation with their metastatic abilities. Early adherence to culture plates was significantly lower in two of three low-metastatic sublines, which occupied smaller surface areas on the culture plates than other sublines did. Comparison of the expression of 637 cancer-related genes by cDNA microarray revealed that seven genes were differentially expressed between high- and low-metastatic sublines. Among them, five genes (AXL, TGFA, COLL7A1, WNT5A, and MKK6) were associated with adherence, motility, and/or invasiveness. These results suggest that the differences in motility/invasiveness and adhesive abilities are key determinants of lung metastasis in osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/14669798', 'no', 'no', 'NA', 5),
(14676807, 'Leonard P, Sharp T, Henderson S, Hewitt D, Pringle J, Sandison A, Goodship A, Whelan J, Boshoff C', 'Gene expression array profile of human osteosarcoma.', 'Br J Cancer', '2003', 'Gene expression array profile of human osteosarcoma. The aims of this pilot study were to determine whether needle and open biopsies from osteosarcoma (OS) provide sufficient quality of mRNA for cDNA array analyses to gain insights into the expression profile of OS. A total of 22 samples collected from OS were used for array analyses. A primary cell culture was also established from one of the OS biopsies. Total RNA was extracted and probes were generated for cDNA arrays. cDNA probes were made for all the 22 samples. Two of these samples were needle core bone biopsies. Statistical analysis confirmed the reliability of array data obtained in 16 of the 22 samples. Known genes involved in bone metabolism and osteosarcoma were identified as highly expressed, and the putative new marker Ezrin was also identified. Confirmatory immunohistochemical staining using the Ezrin antibody was performed in a selection of samples.', 'http://www.ncbi.nlm.nih.gov/pubmed/14676807', 'yes', 'no', 'NA', 1),
(14679136, 'Serra M, Reverter-Branchat G, Maurici D, Benini S, Shen JN, Chano T, Hattinger CM, Manara MC, Pasello M, Scotlandi K, Picci P', 'Analysis of dihydrofolate reductase and reduced folate carrier gene status in relation to methotrexate resistance in osteosarcoma cells.', 'Ann Oncol', '2004', 'Analysis of dihydrofolate reductase and reduced folate carrier gene status in relation to methotrexate resistance in osteosarcoma cells. To evaluate the impact of dihydrofolate reductase (DHFR) and reduced folate carrier (RFC) genes on methotrexate (MTX) resistance in osteosarcoma cells in relation to retinoblastoma (RB1) gene status.A series of human osteosarcoma cell lines-either sensitive or resistant to MTX-and 16 osteosarcoma tumour samples were used in this study.In U-2OS MTX-resistant variants, and in other RB1-positive cell lines, MTX resistance was associated with increased levels of DHFR and with a slight decrease of RFC gene expression. In Saos-2 MTX-resistant variants, and in another RB1-negative cell line, development of MTX resistance was associated with a decrease in expression of RFC, without any significant involvement of DHFR. In osteosarcoma clinical samples, amplification of the DHFR gene at clinical onset appeared to be more frequent in RB1-positive compared with RB1-negative tumours.Amplification of the DHFR gene may occur more frequently in the presence of RB1-mediated negative regulation of its activity and can be present at clinical onset in osteosarcoma patients. Simultaneous evaluation of RFC, DHFR and RB1 gene status at the time of diagnosis may become the basis for the identification of potentially MTX-unresponsive osteosarcoma patients, who could benefit from treatment protocols with alternative antifolate drugs.', 'http://www.ncbi.nlm.nih.gov/pubmed/14679136', 'yes', 'yes', 'methotrexate', 3),
(14682477, 'Liang M, Liao EY, Xu X, Luo XH, Xiao XH', 'Effects of progesterone and 18-methyl levonorgestrel on osteoblastic cells.', 'Endocr Res', '2003', 'Effects of progesterone and 18-methyl levonorgestrel on osteoblastic cells. To study the effects of progesterone (P) and 18-methyl levonorgestrel (LNG) on the expression of P receptors (PRA and PRB), c-fos, c-jun, and osteocalcin in the human osteosarcoma cell line MG-63 and human normal osteoblasts in order to understand the mechanism of progestin action on the proliferation and differentiation of osteoblasts.Cell proliferation was tested by MTT assay. The expression of PR, c-fos, c-jun, and osteocalcin was measured by semi-quantitative RT-PCR, Western immunoblot or immunocytochemistry.Progesterone and LNG (10(-10)-10(-6)M) stimulated cell proliferation in a dose-dependent manner. Progesterone and LNG did not affect the expression of PRA and PRB mRNA and protein, but c-fos and c-jun mRNA and protein were upregulated in a dose-dependent manner. The expression of osteocalcin mRNA was also increased in human osteoblasts in a dose and time-dependent manner with greater effects of LNG than P, while the expression of osteocalcin mRNA in MG-63 cells was not changed by P or LNG.Progesterone and LNG promote osteocalcin gene transcription by stimulating the expression of c-fos and c-jun, and result in osteoblast proliferation and differentiation.', 'http://www.ncbi.nlm.nih.gov/pubmed/14682477', 'no', 'no', 'NA', 4),
(14695161, 'McGary EC, Heimberger A, Mills L, Weber K, Thomas GW, Shtivelband M, Lev DC, Bar-Eli M', 'A fully human antimelanoma cellular adhesion molecule/MUC18 antibody inhibits spontaneous pulmonary metastasis of osteosarcoma cells in vivo.', 'Clin Cancer Res', '2003', 'A fully human antimelanoma cellular adhesion molecule/MUC18 antibody inhibits spontaneous pulmonary metastasis of osteosarcoma cells in vivo. The melanoma cellular adhesion molecule, also known as MUC18, is highly expressed on several tumors, including bone sarcomas. The level of MUC18 expression has been found to correlate directly with tumor progression and metastatic potential. These observations have established MUC18 as a candidate mediator of tumor growth and metastasis, and suggest that blockade of MUC18 might be a potential target for immunotherapy against several MUC18-expressing tumors, including human bone sarcomas.To investigate whether blockade of MUC18 might be a potential target for immunotherapy against osteosarcoma, we have recently developed a fully human anti-MUC18 antibody, ABX-MA1. We studied the effect of ABX-MA1 on growth, adhesion, invasion, and metastasis of human osteosaroma cells both in vitro and in vivo.MUC18 was widely expressed on both osteosarcoma and Ewing''s sarcoma cells. ABX-MA1 had no effect on the proliferation of osteosarcoma cells in vitro, nor did it significantly inhibit the growth of KRIB human osteosarcoma cells when they were orthotopically implanted into the tibias of nude mice. However, after 6 weeks, significantly fewer ABX-MA1-treated mice developed spontaneous pulmonary metastases than did IgG-treated control mice. Additionally, ABX-MA1 decreased the invasion of osteosarcoma cells through Matrigel-coated filters and disrupted homotypic adhesion between osteosarcoma cells and their heterotypic interaction with human vascular endothelial cells.Our findings demonstrate that MUC18 plays a central role in the metastasis of osteosarcoma and suggest that targeted inhibition of this antigen by ABX-MA1 may be a novel immunotherapeutic approach in the management of this tumor.', 'http://www.ncbi.nlm.nih.gov/pubmed/14695161', 'no', 'no', 'NA', 1),
(14704791, 'Khanna C, Wan X, Bose S, Cassaday R, Olomu O, Mendoza A, Yeung C, Gorlick R, Hewitt SM, Helman LJ', 'The membrane-cytoskeleton linker ezrin is necessary for osteosarcoma metastasis.', 'Nat Med', '2004', 'The membrane-cytoskeleton linker ezrin is necessary for osteosarcoma metastasis. Metastatic cancers, once established, are the primary cause of mortality associated with cancer. Previously, we used a genomic approach to identify metastasis-associated genes in cancer. From this genomic data, we selected ezrin for further study based on its role in physically and functionally connecting the actin cytoskeleton to the cell membrane. In a mouse model of osteosarcoma, a highly metastatic pediatric cancer, we found ezrin to be necessary for metastasis. By imaging metastatic cells in the lungs of mice, we showed that ezrin expression provided an early survival advantage for cancer cells that reached the lung. AKT and MAPK phosphorylation and activity were reduced when ezrin protein was suppressed. Ezrin-mediated early metastatic survival was partially dependent on activation of MAPK, but not AKT. To define the relevance of ezrin in the biology of metastasis, beyond the founding mouse model, we examined ezrin expression in dogs that naturally developed osteosarcoma. High ezrin expression in dog tumors was associated with early development of metastases. Consistent with this data, we found a significant association between high ezrin expression and poor outcome in pediatric osteosarcoma patients.', 'http://www.ncbi.nlm.nih.gov/pubmed/14704791', 'no', 'yes', 'NA', 2),
(14706926, 'Postiglione L, Di Domenico G, Giordano-Lanza G, Ladogana P, Turano M, Castaldo C, Di Meglio F, Cocozza S, Montagnani S', 'Effect of human granulocyte macrophage-colony stimulating factor on differentiation and apoptosis of the human osteosarcoma cell line SaOS-2.', 'Eur J Histochem', '2003', 'Effect of human granulocyte macrophage-colony stimulating factor on differentiation and apoptosis of the human osteosarcoma cell line SaOS-2. We investigated the effects of human granulocyte macrophage-colony stimulating factor (GM-CSF) on the relation between differentiation and apoptosis in SaOS-2 cells, an osteoblast-like cell line. To determine the relationship between these cellular processes, SaOS-2 cells were treated in vitro for 1, 7 and 14 days with 200 ng/mL GM-CSF and compared with untreated cells. Five nM insulin-like growth factor (IGF-I) and 30 nM okadaic acid were used as negative and positive controls of apoptosis, respectively. Effects on cell differentiation were determined by ECM (extracellular matrix) mineralization, morphology of some typical mature osteoblast differentiation markers, such as osteopontin and sialoprotein II (BSP-II), and production of bone ECM components such as collagen I. The results showed that treatment with GM-CSF caused cell differentiation accompanied by increased production of osteopontin and BSP-II, together with increased ECM deposition and mineralization. Flow cytometric analysis of annexin V and propidium iodide incorporation showed that GM-CSF up-regulated apoptotic cell death of SaOS-2 cells after 14 days of culture in contrast to okadaic acid, which stimulated SaOS-2 apoptosis only during the early period of culture. Endonucleolytic cleavage of genomic DNA, detected by \\Aúladdering analysis\\Aù, confirmed these data. The results suggest that GM-CSF induces osteoblastic differentiation and long-term apoptotic cell death of the SaOS-2 human osteosarcoma cell line, which in turn suggests a possible in vivo physiological role for GM-CSF on human osteoblast cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/14706926', 'no', 'no', 'NA', 3),
(14713104, 'Mathisen B, Lindstad RI, Hansen J, El-Gewely SA, Maelandsmo GM, Hovig E, Fodstad O, Loennechen T, Winberg JO', 'S100A4 regulates membrane induced activation of matrix metalloproteinase-2 in osteosarcoma cells.', 'Clin Exp Metastasis', '2003', 'S100A4 regulates membrane induced activation of matrix metalloproteinase-2 in osteosarcoma cells. To study the role of the metastasis associated protein S100A4, an osteosarcoma cell line (OHS) with a high level of this protein was transfected with a vector containing a ribozyme that degrades S100A4 mRNA and, as controls, OHS cells were transfected with the vector alone. We have followed up our previous investigation (Bjørnland et al. 1999) by a detailed investigation of these cell lines'' synthesis of MMP and TIMP proteins at different cell densities. It is shown that the cell lines with a low S100A4 level produced a reduced amount of immunoreactive MMP-2 at cellular subconfluence, while at confluence there was no difference compared to the control cells. The cell lines with a reduced S100A4 level produced less of the activated form of MMP-2 (62-kDa) and less TIMP-1 than the corresponding control cells, independent of cell density. Isolated cell membranes from cell lines with a reduced S100A4 level contained less MT1-MMP, MMP-2 and TIMP-2 compared to the control cells. Activation of exogenously added proMMP-2 was less effective with the former membrane preparations. It appeared that the mechanism behind the S100A4 dependent activation of proMMP-2 varied with cell density, as SN50, a peptide inhibitor of NF-kappaB nuclear translocation reduced the activation of MMP-2 at low cell density, but had no effect at high cell density. Thus, one of the mechanisms by which S100A4 may exert its effect on metastasis of some tumors is by regulating the MMP-2 activity.', 'http://www.ncbi.nlm.nih.gov/pubmed/14713104', 'no', 'no', 'NA', 5),
(14719070, 'Yamazaki H, Handa A, Nishi M, Tokunaga T, Tomisawa M, Hatanaka H, Abe Y, Kijima H, Ueyama Y, Nakamura M', 'Ribozyme mediated down-regulation of thrombospondin receptor CD36 inhibits the growth of the human osteosarcoma cell line.', 'Oncol Rep', '2004', 'Ribozyme mediated down-regulation of thrombospondin receptor CD36 inhibits the growth of the human osteosarcoma cell line. Thrombospondin-1 (TSP1) is known to possess tumor suppressor functions. In contradiction, TSP1 enhances the stromal vascularization and growth of certain cancers. A cell adhesion receptor, CD36, has been shown to interact with a ligand TSP1. We studied how CD36 affects the growth of the osteosarcoma cell line (HOS) expressing TSP1. We used the anti-CD36 ribozyme to specifically suppress CD36 gene expression in the HOS. The expression of the CD36 mRNA was significantly suppressed in the ribozyme-introduced cell line (HOS/Rz). The transformant HOS/Rz markedly decreased its growth. The growth of the osteosarcoma cell line HOS may be regulated by autocrine or paracrine loop TSP1 and CD36.', 'http://www.ncbi.nlm.nih.gov/pubmed/14719070', 'no', 'no', 'NA', 2),
(14719113, 'Manara MC, Serra M, Benini S, Picci P, Scotlandi K', 'Effectiveness of Type I interferons in the treatment of multidrug resistant osteosarcoma cells.', 'Int J Oncol', '2004', 'Effectiveness of Type I interferons in the treatment of multidrug resistant osteosarcoma cells. P-glycoprotein overexpression is an important adverse prognostic marker for osteosarcoma (OS) patients, which is associated with higher risk for developing metastases as a consequence of the limited responsiveness to standard treatments of P-glycoprotein overexpressing OS cells. The use of cytokines has been advocated as a possible therapeutic approach to overcome multidrug resistance (MDR), being active on cell lines that are resistant to conventional drugs. In this study, we evaluated in vitro effects of interferons (IFNs) on MDR P-glycoprotein overexpressing OS cells. Type I IFNs, but not IFNgamma, showed tangible inhibitory effects on OS cell growth, which were higher in MDR cell lines compared to parental cells. The higher sensitivity of P-glycoprotein overexpressing cells to Type I IFNs correlates with higher expression of the activator of the transcription (STAT)-2 and (STAT)-3, two intracellular mediators of the IFNalpha and IFNbeta signaling pathways, whereas no differences were observed with respect to the expression or activation of the Type I IFN receptor and STAT-1. Exposure of OS MDR cells to Type I IFN decreased the expression of P-glycoprotein. This effect resulted in a significantly increased chemosensitivity of MDR cells to doxorubicin. Therefore, our data support the use of IFNalpha or IFNbeta in the treatment of osteosarcoma patients who overexpress P-glycoprotein in their primary tumors, and respond insufficiently to current therapeutic regimens.', 'http://www.ncbi.nlm.nih.gov/pubmed/14719113', 'no', 'no', 'doxorubicin', 3),
(14735475, 'Hoang BH, Kubo T, Healey JH, Sowers R, Mazza B, Yang R, Huvos AG, Meyers PA, Gorlick R', 'Expression of LDL receptor-related protein 5 (LRP5) as a novel marker for disease progression in high-grade osteosarcoma.', 'Int J Cancer', '2004', 'Expression of LDL receptor-related protein 5 (LRP5) as a novel marker for disease progression in high-grade osteosarcoma. The Wingless-type (Wnt) family of proteins and its coreceptor LRP5 have recently been implicated in human skeletal development. Wnt pathway modulates cell fate and cell proliferation during embryonic development and carcinogenesis through activation of receptor-mediated signaling. Osteosarcoma (OS) is a bone-forming tumor of mesenchymal origin whose growth control has been linked to autocrine or paracrine stimulation by several growth factor families. We examined 4 OS cell lines for WNT1, WNT4, WNT5A, WNT7A, WNT11, FZD1-10 and LRP5 expression by reverse transcription polymerase chain reaction (RT-PCR). In addition, RT-PCR for LRP5 expression was performed in 44 OS patient samples and the findings were correlated with clinical data. Expression profiling of Wnts and their receptors revealed the presence of several isoforms in OS cell lines. Overall, 22/44 (50%) of OS patient samples showed evidence of LRP5 expression. Presence of LRP5 correlated significantly with tumor metastasis (p = 0.005) and the chondroblastic subtype of OS (p = 0.045). In addition, patients whose tumors were positive for LRP5 showed a trend toward decreased event-free survival (p = 0.066). No significant association was found between LRP5 expression and age, gender, site of disease, site of metastasis or degree of chemotherapy-induced tumor necrosis. Sequencing of exon 3 of LRP5 in 10 OS patient-derived cell cultures showed no activating mutation of LRP5. These results showed that expression of LRP5 is a common event in OS and strongly suggest a role for LRP and Wnt signaling in the pathobiology and progression of this disease.Copyright 2003 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/14735475', 'yes', 'yes', 'NA', 1),
(14760101, 'Duan X, Zhou Z, Jia SF, Colvin M, Lafleur EA, Kleinerman ES', 'Interleukin-12 enhances the sensitivity of human osteosarcoma cells to 4-hydroperoxycyclophosphamide by a mechanism involving the Fas/Fas-ligand pathway.', 'Clin Cancer Res', '2004', 'Interleukin-12 enhances the sensitivity of human osteosarcoma cells to 4-hydroperoxycyclophosphamide by a mechanism involving the Fas/Fas-ligand pathway. Cyclophosphamide (CY) and its derivative ifosfamide are alkylating agents used to treat osteosarcoma (OS). The purpose of these studies was to determine whether alkylating agents affect the expression of Fas ligand (FasL) and whether interleukin 12 enhances the sensitivity of human OS cells to alkylating agents. 4-Hydroperoxycyclophosphamide (4-HC), the preactivated CY compound, and 4-hydroperoxydidechlorocloclophosphamide (4-HDC), its nonalkylating analogue, human OS LM6 cells, and a clone of cells derived by transfection with the interleukin 12 gene (LM6-#6) were used for these studies. Incubation of LM6 and LM6-#6 with 10 micro M 4-HC increased the expression of FasL mRNA (2.5- and 3.0-fold, respectively). By contrast, 4-HDC, Adriamycin (ADR), cisplatin (CDP), and methotrexate (MTX) had no effect on FasL mRNA expression. Increased FasL expression after treatment with 4-HC was also demonstrated by immunohistochemistry and flow cytometry. Drug-induced FasL was functional and mediated cell death. We examined the effect of FasL up-regulation by 4-HC on LM6 and LM6-#6 cells. Flow cytometry showed that LM6-#6 cells expressed 2.2-fold more Fas than LM6 cells. Cytotoxicity of 4-HC, 4-HDC, ADR, CDP, and MTX on LM6, LM6-neo, and LM6-#6 were quantified. Colony-forming assay revealed an IC(50) of 2.10 micro M for 4-HC in LM6-neo cells compared with 0.41 micro M in LM6-#6 cells. The IC(50) for 4-HDC, ADR, CDP, and MTX were not significantly different between the two cell lines. We concluded that the increased expression of Fas enhanced LM6-#6 sensitivity to 4-HC. These data indicate that Fas/FasL may be involved in the cytotoxic pathway of CY. Combining biological agents with chemotherapeutic agents that have complementary Fas/FasL pathway actions may offer new therapeutic alternatives.', 'http://www.ncbi.nlm.nih.gov/pubmed/14760101', 'no', 'no', 'cyclophosphamide', 3),
(14973099, 'Barnette P, Scholl R, Blandford M, Ballard L, Tsodikov A, Magee J, Williams S, Robertson M, Ali-Osman F, Lemons R, Keller C', 'High-throughput detection of glutathione s-transferase polymorphic alleles in a pediatric cancer population.', 'Cancer Epidemiol Biomarkers Prev', '2004', 'High-throughput detection of glutathione s-transferase polymorphic alleles in a pediatric cancer population. Polymorphisms of glutathione S-transferase (GST) enzymes have been correlated with altered risk of several cancers, as well as altered response and toxicity from cancer chemotherapy. We report a low cost, highly reproducible and specific PCR-based high-throughput assay for genotyping different GSTs designed for use in large clinical trials. In comparison to an alternative genotyping method (single nucleotide extension), the sensitivity and specificity of the high throughput assay was shown to be 92 and 97%, respectively, depending on the source of genomic DNA. Using the high-throughput assay, we demonstrate by multivariate analysis an increased risk of acute lymphoblastic leukemia, glial brain tumors, and osteosarcoma for patients carrying nonnull alleles of GSTM1 and/or GSTT1.', 'http://www.ncbi.nlm.nih.gov/pubmed/14973099', 'yes', 'yes', 'NA', 2),
(14975209, 'Liu SJ, Hu GF, Liu YJ, Liu SG, Gao H, Zhang CS, Wei YY, Xue Y, Lao WD', 'Effect of human osteopontin on proliferation, transmigration and expression of MMP-2 and MMP-9 in osteosarcoma cells.', 'Chin Med J (Engl)', '2004', 'Effect of human osteopontin on proliferation, transmigration and expression of MMP-2 and MMP-9 in osteosarcoma cells. To explore the effect of human osteopontin (hOPN) on the proliferation, transmigration and expression of matrix metallproteinase-2 (MMP-2) and matrix metallproteinase-9 (MMP-9) in osteosarcoma (OS) cells in vitro.The prokaryotic-expression vector of hOPN was produced. hOPN was then subcloned into E. coli BL21 (DE3) cells and purified with ProBond trade mark Columns. The proliferation, cell cycle and the expression of cyclin A in OS cells were investigated by using MTT assay, flow cytometry and Western blot respectively. The transmigration of OS cells was checked by using transwell cell culture chamber. The micro-pore-filter-membrane system was used to study the chemiotaxis of hOPN to OS cells. The levels of total protein were examined according to Coomassie Brilliant Blue manuals. The expression of MMP-2 and MMP-9 were evaluated by detecting the volume of degradation of gelatin on SDS-PAGE gel.The prokaryotic-expression vector of hOPN and purified hOPN protein were achieved hOPN promoted OS cells proliferation in a dose-dependent manner, and stimulated cyclin A expression in OS cells to accelerate cell division cycle. hOPN facilitated the trans-membrane migration of OS cells. hOPN also enhanced the secretion of MMP-2 and MMP-9 in OS cells.hOPN could stimulate cyclin A expression in OS cells. hOPN has chemiotaxis to OS cells and increases their transmigration. hOPN enhances the secretion of MMP-2 and MMP-9 in OS cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/14975209', 'no', 'no', 'NA', 4),
(14996747, 'Trougakos IP, So A, Jansen B, Gleave ME, Gonos ES', 'Silencing expression of the clusterin/apolipoprotein j gene in human cancer cells using small interfering RNA induces spontaneous apoptosis, reduced growth ability, and cell sensitization to genotoxic and oxidative stress.', 'Cancer Res', '2004', 'Silencing expression of the clusterin/apolipoprotein j gene in human cancer cells using small interfering RNA induces spontaneous apoptosis, reduced growth ability, and cell sensitization to genotoxic and oxidative stress. Clusterin/Apolipoprotein J (CLU) is a heterodimeric ubiquitously expressed secreted glycoprotein that is implicated in several physiological processes and is differentially expressed in many severe physiological disturbances, including tumor formation and in vivo cancer progression. Despite extensive efforts, clarification of CLU''s biological role has been exceptionally difficult and its precise function remains elusive. Short RNA duplexes, referred to as small interfering RNAs (siRNAs), provide a new approach for the elucidation of gene function in human cells. Here, we describe siRNA-mediated CLU gene silencing in osteosarcoma and prostate human cancer cells and illustrate that CLU mRNA is amenable to siRNA-mediated degradation. Our data demonstrate that CLU knockdown in human cancer cells induces significant reduction of cellular growth and higher rates of spontaneous endogenous apoptosis. Moreover, CLU knockdown cancer cells were significantly sensitized to both genotoxic and oxidative stress induced by chemotherapeutic drugs and H(2)O(2), respectively. These effects were more pronounced in cell lines that express high endogenous steady-state levels of the CLU protein and occur through hyperactivation of the cellular apoptotic machinery. Overall, our results reveal that, in the distinct cellular contexts of the osteosarcoma and prostate cancer cells assayed, CLU is a central molecule in cell homeostasis that exerts a cytoprotective function. The described CLU-specific siRNA oligonucleotides that can potently silence CLU gene expression may thus prove valuable agents during antitumor therapy or at other pathological conditions where CLU has been implicated.', 'http://www.ncbi.nlm.nih.gov/pubmed/14996747', 'no', 'no', 'NA', 1);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(14999771, 'Dechant MJ, Fellenberg J, Scheuerpflug CG, Ewerbeck V, Debatin KM', 'Mutation analysis of the apoptotic \\death-receptors\\ and the adaptors TRADD and FADD/MORT-1 in osteosarcoma tumor samples and osteosarcoma cell lines.', 'Int J Cancer', '2004', 'Mutation analysis of the apoptotic \\death-receptors\\ and the adaptors TRADD and FADD/MORT-1 in osteosarcoma tumor samples and osteosarcoma cell lines. Apoptosis is a key mechanism of the organism that regulates embryogenesis and development, maintains homeostasis of the immune system and removes potentially hazardous cells. A dysregulation of apoptosis signaling may thus disturb the balance of cell survival and cell death, leading to the development of several diseases including cancer. In order to determine whether osteosarcomas display an increased frequency of genetic alterations that affect apoptosis signaling, we analyzed the death domains of the death receptor genes CD95/Fas/Apo1, TNFR1, DR3/Apo3/WSL-1/LARD/TRAMP, DR5/TRAIL-R2/TRICK2/KILLER, DR6 and the complete coding sequences of the death receptor gene DR4/TRAIL-R1 and the genes of the adaptors TRADD and FADD/MORT-1. The investigation included 15 osteosarcoma tumor samples, 3 osteosarcoma cell lines (SAOS-2, HOS and MG63) and peripheral blood from 20 donors as controls. We were able to identify 4 different sequence variations within the DR4 gene located on exons 3, 4, 5 and 10 (death-domain). No alterations have been detected in the other genes or exons investigated. Except the sequence variant affecting exon 4, the alterations were homozygous in 15% of the tumor samples and cell lines, whereas the same alterations found in the control group were heterozygous or even not detectable. Three out of 4 alterations are located in the receptor''s extracellular cysteine rich domain, which contains the ligand binding area and 1 on exon 10 coding for the death-domain. They may thus exert influence on ligand-receptor interactions and subsequent apoptosis induction. Our findings suggest that homozygous genetic alterations within the DR4 gene may be implicated in the formation of osteosarcoma.Copyright 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/14999771', 'yes', 'no', 'NA', 1),
(15015580, 'Lu KH, Wang DY, Lue KH, Hsiao YM, Chou MC, Chen YS, Chung JG', 'PCR and flow cytometric analysis of paclitaxel-inhibited arylamine N-acetyltransferase activity and gene expression in human osteogenic sarcoma cells (U-2 OS).', 'Anticancer Res', 'NA', 'PCR and flow cytometric analysis of paclitaxel-inhibited arylamine N-acetyltransferase activity and gene expression in human osteogenic sarcoma cells (U-2 OS). Human epidemiological studies suggest an association between N-acetyltransferase (NAT) activity and the incidence of bladder and colorectal cancers. In this study, paclitaxel was selected to examine the inhibition of arylamine NAT activity, gene expression and 2-aminofluorene-DNA adduct formation in a human osteogenic sarcoma cell line (U-2 OS). The activity of NAT was determined by high performance liquid chromatography (HPLC) assay for the amounts of acetylated 2-aminofluorene (AF) and p-aminobenzoic acid (PABA) and nonacetylated AF and PABA. Human osteogenic sarcoma cell cytosols and intact cells were used to examine the NAT activity, gene expression and AF-DNA adduct formation. The results demonstrated that NAT activity percent of NAT in examined cells, gene expression (NAT1 mRNA) and AF-DNA adduct formation in human osteogenic sarcoma cells were inhibited and decreased by paclitaxel in a dose-dependent manner. The results also demonstrated that paclitaxel decreased the apparent values of Km and Vmax from intact human osteogenic sarcoma cells (U-2 OS). Thus, paclitaxel is an uncompetitive inhibitor of the NAT enzyme.', 'http://www.ncbi.nlm.nih.gov/pubmed/15015580', 'no', 'no', 'paclitaxel', 1),
(15026342, 'Hughes DP, Thomas DG, Giordano TJ, Baker LH, McDonagh KT', 'Cell surface expression of epidermal growth factor receptor and Her-2 with nuclear expression of Her-4 in primary osteosarcoma.', 'Cancer Res', '2004', 'Cell surface expression of epidermal growth factor receptor and Her-2 with nuclear expression of Her-4 in primary osteosarcoma. There is controversy over the role of Her-2 in osteosarcoma, with some investigators reporting association between expression and adverse outcome, whereas others point to the lack of gene amplification and membranous expression by immunohistochemistry (IHC) as inconsistent with biological significance. Her-2 normally requires pairing with epidermal growth factor receptor (EGFR), Her-3, or Her-4, but these have been less well studied in osteosarcoma. We evaluated the expression of each of these receptors in osteosarcoma and their potential to contribute to pathogenesis by examining a panel of low-passage primary osteosarcoma cell lines, comparing these with archival tumor specimens. Her-2 immunoreactivity was seen frequently in the diffuse staining pattern described previously. We observed EGFR in all samples by IHC. Her-3 expression was not observed. Her-4 expression was nuclear in distribution in all tumor samples and many cell line samples, consistent with activation and cleavage of the receptor. Quantified expression of Her-2 and EGFR mRNA by quantitative, real-time PCR in cell lines correlated with IHC for Her-2 but not for EGFR. Western blot identified full-length receptors for EGFR and Her-2 in all expected cell lines and showed Her-4 to be predominantly in the p80 form. Flow cytometry identified cell surface Her-2 and EGFR in all lines with receptor expression by IHC. We conclude that the cell surface expression of Her-2 and EGFR and the nuclear localization of the activated p80 fragment of Her-4 suggest that all three may be contributing to osteosarcoma pathogenesis. Therapy directed against this family of receptors may be beneficial for patients with osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15026342', 'yes', 'no', 'NA', 3),
(15026814, 'Tang HJ, Qian D, Sondak VK, Stachura S, Lin J', 'A modified p53 enhances apoptosis in sarcoma cell lines mediated by doxorubicin.', 'Br J Cancer', '2004', 'A modified p53 enhances apoptosis in sarcoma cell lines mediated by doxorubicin. Mdm2 is frequently overexpressed in sarcoma cells and may contribute to drug resistance by increasing p53 degradation. We investigated the induction of apoptosis in sarcoma cells via adenovirus-mediated gene transfer of wild-type p53 and two modified p53 genes, p53 14/19 and p53 22/23, whose protein products are resistant to Mdm2-mediated degradation. We found that adenovirus-wt p53 (Ad-wt p53) induces significant apoptosis in HT1080 fibrosarcoma cells expressing low levels of Mdm2, but fails to induce apoptosis in SJSA osteosarcoma cells expressing high levels of Mdm2. In contrast, Ad-p53 14/19 induces significant apoptosis in both cell lines. Interestingly, Ad-p53 22/23, a vector encoding a transcription-defective p53 mutant, causes limited apoptosis in both cell lines. We demonstrate that doxorubicin induces phosphorylation of both wt p53 and p53 14/19 protein at multiple sites. We tested the efficacy of doxorubicin and cisplatin with either Ad-wt p53, Ad-p53 22/23 or Ad-p53 14/19. SJSA cells, although harbouring endogenous wt p53, did not undergo significant apoptosis following doxorubicin or cisplatin exposure alone or combined with Ad-wt p53. In contrast, doxorubicin or cisplatin plus Ad-p53 14/19 induced significant apoptosis. Gene transfer of p53 14/19 in combination with the administration of doxorubicin or cisplatin is a potential therapeutic approach for cancers expressing high levels of Mdm2.', 'http://www.ncbi.nlm.nih.gov/pubmed/15026814', 'no', 'no', 'doxorubicin', 2),
(15049011, 'Cheng YY, Huang L, Lee KM, Li K, Kumta SM', 'Alendronate regulates cell invasion and MMP-2 secretion in human osteosarcoma cell lines.', 'Pediatr Blood Cancer', '2004', 'Alendronate regulates cell invasion and MMP-2 secretion in human osteosarcoma cell lines. Osteosarcoma is the most common malignant bone tumor of childhood. Significant proportions of these patients eventually develop pulmonary metastases and succumb to their disease even after conventional multi-agent chemotherapy and surgical excision. Matrix metalloproteinase (MMP)-2 induced degradation of blood vessel basement membranes is an important pre-requisite for tumor invasion and metastasis. Bisphosphonates (BPs) have been known to inhibit tumor growth and metastasis in some tumors such as breast cancer, renal cell carcinoma, and prostate cancer, and may do so through inhibition of MMP secretion. We, therefore, tested the effect of BPs on tumor cell invasion, MMP-2 secretion, and apoptosis of osteosarcoma cell lines.Two osteosarcoma cell lines (SaOS-2, U(2)OS) were treated with alendronate (50, 100, and 150 microM) for 24 and 48 hr. Matrigel invasion assay was used to investigate the invasive potential of osteosarcoma cell lines before and after alendronate treatment. Real-time quantitative RT-PCR was used to determine the mRNA level of MMP-2 with and without alendronate treatment. Enzyme-linked immunosorbent assay (ELISA) was used to quantify the cytokine level of MMP-2 secreted in the condition medium. BP-induced cell apoptosis was evaluated by fluorescent flow cytometric analysis.The results showed that alendronate inhibited cell invasion of both osteosarcoma cell lines in a dose-dependent manner. Alendronate reduced the mRNA level and cellular level of MMP-2 in both cell lines in a time and dose-dependent manner. Alendronate also induced significant apoptosis in both cell lines. Our finding suggests that alendronate downregulates MMP-2 secretion and induces apoptosis in osteosarcoma cells, which may both contribute to the reduction of invasive potential of the tumor cells.Copyright 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15049011', 'no', 'no', 'alendronate', 1),
(15055356, 'Shalita-Chesner M, Glaser T, Werner H', 'Signal transducer and activator of transcription-1 (STAT1), but not STAT5b, regulates IGF-I receptor gene expression in an osteosarcoma cell line.', 'J Pediatr Endocrinol Metab', '2004', 'Signal transducer and activator of transcription-1 (STAT1), but not STAT5b, regulates IGF-I receptor gene expression in an osteosarcoma cell line. The IGF-I receptor (IGF-IR) exhibits potent mitogenic, antiapoptotic, and transforming activities. Previous studies have suggested that the expression of the IGF-IR gene is negatively regulated by certain cytokines, including interferon-gamma (IFN-gamma). The potential involvement of STAT proteins in transcriptional regulation of the IGF-IR gene by IFN-gamma was addressed by transient coexpression of vectors encoding STAT1 and STAT5b, together with an IGF-IR promoter luciferase reporter, in the osteosarcoma-derived cell line Saos-2. Physical interactions between IFN-gamma-induced transcription factors and the IGF-IR promoter region were examined by electrophoretic mobility shift assays (EMSA). The results obtained indicate that the mechanism of action of IFN-gamma involves stimulation of STAT1 which, in turn, binds IFN-gamma activation sites (GAS) in the IGF-IR regulatory region, thus suppressing promoter activity. Taken together, our results suggest that the IGF-IR gene is a novel target for STAT1 action and that at least part of the inhibitory effects of STAT1 may involve repression of the strongly antiapoptotic IGF-IR gene.', 'http://www.ncbi.nlm.nih.gov/pubmed/15055356', 'no', 'no', 'NA', 3),
(15067366, 'Takahashi T, Ogawa Y, Kobayashi T, Sonobe H, Seguchi H, Tani T, Yoshida S', 'The role of selective COX-2 inhibitors in reactive oxygen species formation in osteosarcoma cells after X-ray irradiation.', 'Int J Mol Med', '2004', 'The role of selective COX-2 inhibitors in reactive oxygen species formation in osteosarcoma cells after X-ray irradiation. Selective inhibition of COX-2 is thought to prevent carcinogenesis in some malignant tumors. In this study, in an effort to enhance the effectiveness of osteosarcoma treatment, we investigated the effect of a selective inhibitor of COX-2, with or without irradiation. We also asked whether selective COX-2 inhibitors increase the effect of X-ray irradiation, with regard to reactive oxygen species (ROS) formation in an osteosarcoma cell line. Our results showed that the presence of COX-2 inhibitor without irradiation results in faint spots of ROS formation that do not appear in the absence of COX-2 inhibitor. However, COX-2 inhibitor did not induce ROS formation when combined with irradiation. Thus, radiotherapy with selective COX-2 inhibitions has limitations in the treatment of radioresistant osteosarcoma to obtain the effective achievement, it is indispensable to combine another agent in future studies.', 'http://www.ncbi.nlm.nih.gov/pubmed/15067366', 'no', 'no', 'NA', 1),
(15069760, 'Tsai JY, Aviv H, Benevenia J, Chang VT, Patterson F, Aisner S, Hameed M', 'HER-2/neu and p53 in osteosarcoma: an immunohistochemical and fluorescence in situ hybridization analysis.', 'Cancer Invest', '2004', 'HER-2/neu and p53 in osteosarcoma: an immunohistochemical and fluorescence in situ hybridization analysis. The overexpression of HER-2/neu and p53 has been associated with poor outcome in many neoplasms. Their role in patients with osteosarcoma is unclear. We studied the expression of HER-2/neu and p53 in 22 osteosarcoma samples (from 20 patients--2 had locally recurrent disease) biopsied at the University of Medicine and Dentistry of New Jersey (UMDNJ) from 1996-2000 using both immunohistochemical (IHC) and fluorescence in situ hybridization (FISH) analysis. Fourteen patients (14 samples) presented with Stage II and 6 patients (8 samples) presented with Stage III disease. Median follow-up is two years (range one year to five years). Four of 22 (18%) samples showed focal membranous or cytoplasmic positivity for HER-2/neu and six of 22 samples (27%) showed nuclear positivity for p53 by IHC analysis. In contrast, none of 22 tested samples showed gene amplification for HER-2/neu by FISH analysis. Seven of 13 HER-2/neu and p53 negative patients (54%) are currently disease free (between one year to five years). In this sample of patients, the HER-2/neu oncogene is not overexpressed or amplified in osteosarcoma; six of 22 samples (27%) showed overexpression of p53 by IHC analysis. By FISH, none of the samples demonstrated deletion of p53. Neither HER2/neu nor p53 expression was important for the biology of osteosarcoma in this population.', 'http://www.ncbi.nlm.nih.gov/pubmed/15069760', 'yes', 'no', 'NA', 2),
(15087387, 'Hoang BH, Kubo T, Healey JH, Yang R, Nathan SS, Kolb EA, Mazza B, Meyers PA, Gorlick R', 'Dickkopf 3 inhibits invasion and motility of Saos-2 osteosarcoma cells by modulating the Wnt-beta-catenin pathway.', 'Cancer Res', '2004', 'Dickkopf 3 inhibits invasion and motility of Saos-2 osteosarcoma cells by modulating the Wnt-beta-catenin pathway. Osteosarcoma (OS) is a primary malignancy of bone with a tendency to metastasize early. Despite intensive chemotherapy and surgical resection, approximately 30% of patients still develop distant metastasis. Our previous work using clinical OS samples suggested that expression of the Wnt receptor LRP5 might be associated with tumor metastasis. In the present study, we used a Dickkopf (Dkk) family member and a dominant-negative LRP5 receptor construct to modulate Wnt signaling in OS cells. Saos-2 cells, which ectopically express Dkk-3, do not undergo apoptosis and exhibit enhanced resistance to serum starvation and chemotherapy-induced cytotoxicity. Transfection of Dkk-3 and dominant-negative LRP5 into Saos-2 cells significantly reduces invasion capacity and cell motility. This blockade is associated with changes in cell morphology consistent with a less invasive phenotype. In addition, Dkk-3 and dominant-negative LRP5 also induce changes in beta-catenin localization consistent with an increase in cell-cell adhesion. Taken together, these results support a possible role for Wnt signaling in the pathobiology and progression of human OS.', 'http://www.ncbi.nlm.nih.gov/pubmed/15087387', 'no', 'no', 'NA', 3),
(15093570, 'Anninga JK, van de Vijver MJ, Cleton-Jansen AM, Kristel PM, Taminiau AH, Nooij M, Egeler RM, Hogendoorn PC', 'Overexpression of the HER-2 oncogene does not play a role in high-grade osteosarcomas.', 'Eur J Cancer', '2004', 'Overexpression of the HER-2 oncogene does not play a role in high-grade osteosarcomas. The aim of our study was to determine whether or not the tyrosine kinase receptor, HER2 (also known as ErbB2/Her2/neu), is overexpressed in human osteosarcomas (OS). We studied 15 biopsy and 18 resection specimens at the mRNA and protein levels. HER2 status in the OS specimens was assessed by immunohistochemistry (IHC) and quantitative Real-Time Polymerase chain reaction (PCR). In moderately immunopositive cases fluorescent in situ hybridisation (FISH) analysis was used in order to identify any possible gene amplification. 27 samples were evaluable for IHC and only 1 case showed a moderately positive membrane staining. The remaining samples showed no staining or focal cytoplasmic staining (2 samples). In the moderately positive case, FISH analysis showed no HER-2 gene amplification. There was also no overexpression of HER2 mRNA suggesting this sample was a false-positive immunostain. HER2 mRNA expression was present in all samples at a similar level to that in the breast cancer cell line, MCF7, which does not overexpress HER2 and was used as a negative control. In conclusion, this study shows that HER2 mRNA or membranous HER2 protein overexpression is absent in human OS. We noted various inconsistencies in previous published studies, with regard to methodology and the interpretation of the results based on poor methodology. We therefore conclude that the positive data with regard to HER2 overexpression reported in these previous studies is not reliable. Our results suggest that the monoclonal antibody trastuzumab (Herceptin(R)), directed against the HER2-receptor, is not likely to be an effective therapeutic agent in OS.', 'http://www.ncbi.nlm.nih.gov/pubmed/15093570', 'yes', 'no', 'trastuzumab', 1),
(15095405, 'MacEwen EG, Pastor J, Kutzke J, Tsan R, Kurzman ID, Thamm DH, Wilson M, Radinsky R', 'IGF-1 receptor contributes to the malignant phenotype in human and canine osteosarcoma.', 'J Cell Biochem', '2004', 'IGF-1 receptor contributes to the malignant phenotype in human and canine osteosarcoma. To further define the role of insulin-like growth factor-1 (IGF-1) and its receptor (IGF-1R) in osteosarcoma (OS), human OS cell lines with low (SAOS-2) and high (SAOS-LM2) metastatic potential and three canine OS-derived cell lines were studied. Cell lines were evaluated for: IGF-1R expression; expression of IGF binding proteins (IGFBPs); effect of IGF-1 on tumor cell growth, invasion, expression of urokinase plasminogen activator (uPA), and soluble uPA receptor (suPAR), and; ectopic and orthotopic tumorigenicity of the canine OS cells in athymic mice. All cell lines exhibited steady-state mRNA expression of IGF-1R. The SAOS-2 and SAOS-LM2 cells expressed 9,138 and 10,234 cell-associated binding sites, respectively. Canine OS cells expressed from 1,728 to 3,883 binding sites. Two IGF-1-treated cell lines displayed enhanced proliferation. Two cell lines formed colonies in semisolid media, and IGF-1 increased colony number. Matrigel invasion was enhanced in one cell line following IGF-1 treatment. uPA and suPAR were unchanged in SAOS-2 and SAOS-LM2 cells following IGF-1 treatment, but the highly metastatic OS line SAOS-LM2 expressed five times more suPAR and displayed enhanced invasion compared to the parental, low metastatic SAOS-2. IGFBP-5 was detected in four of five cell lines, and IGFBP-3 was detected in two canine OS cell lines. Two canine OS lines were tumorigenic, and one metastasized spontaneously. In conclusion, OS cells express IGF-1R, which can contribute to their growth and invasion. There is suggestive evidence that increasing receptor number may contribute to in vivo tumorigenesis. Additional studies are needed to determine how IGF-1/IGF-1R interactions contribute to the malignant phenotype of OS.Copyright 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15095405', 'no', 'no', 'NA', 6),
(15112275, 'Ferrari S, Bertoni F, Zanella L, Setola E, Bacchini P, Alberghini M, Versari M, Bacci G', 'Evaluation of P-glycoprotein, HER-2/ErbB-2, p53, and Bcl-2 in primary tumor and metachronous lung metastases in patients with high-grade osteosarcoma.', 'Cancer', '2004', 'Evaluation of P-glycoprotein, HER-2/ErbB-2, p53, and Bcl-2 in primary tumor and metachronous lung metastases in patients with high-grade osteosarcoma. Investigation of the relation between primary tumor and metastatic disease is necessary for the identification of predictive factors for postrecurrence survival (PRS) in patients with recurrent osteosarcoma.Cellular levels of P-glycoprotein, ErbB-2, p53, and Bcl-2 expression were evaluated in primary tumor biopsy and metachronous pulmonary metastasis specimens from 19 patients with high-grade osteosarcoma. Results were analyzed for differences between primary tumor and pulmonary metastases and for correlations between expression patterns and survival.Positive staining in lung metastases was noted in 68%, 53%, 32%, and 84% of patients for P-glycoprotein, ErbB-2, p53, and Bcl-2, respectively. These percentages were higher than those observed in primary tumor specimens for all genetic markers evaluated, with a significant difference in the percentage of patients with positive staining for P-glycoprotein (68% vs. 32%; P = 0.05) and a near-significant difference in the percentage of patients with positive staining for Bcl-2 (84% vs. 53%; P = 0.08). Patients with ErbB-2 expression in the primary tumor were more likely to have multiple metastases and shorter recurrence-free intervals compared with patients in whom ErbB-2 expression was not observed, whereas differences in P-glycoprotein, p53, and Bcl-2 expression were not related to differences in metastatic pattern. PRS was influenced by p53 expression levels in pulmonary metastases, with patients who had negative staining for p53 having a significantly better PRS rate relative to patients with positive staining for p53 (3-year PRS rate: p53-negative, 64%; p53-positive, 17%; P = 0.008).In the current study of patients with high-grade osteosarcoma, most patients exhibited increased cellular expression of P-glycoprotein, ErbB-2, and Bcl-2 in recurrent pulmonary metastases compared with primary tumor. Further studies aimed at investigating the relation between altered p53 expression in lung metastases and postrecurrence survival are recommended.Copyright 2004 American Cancer Society.', 'http://www.ncbi.nlm.nih.gov/pubmed/15112275', 'yes', 'yes', 'NA', 4),
(15138564, 'Chano T, Mori K, Scotlandi K, Benini S, Lapucci C, Manara MC, Serra M, Picci P, Okabe H, Baldini N', 'Differentially expressed genes in multidrug resistant variants of U-2 OS human osteosarcoma cells.', 'Oncol Rep', '2004', 'Differentially expressed genes in multidrug resistant variants of U-2 OS human osteosarcoma cells. Multidrug resistance (MDR) to anticancer agents is a major barrier to the successful treatment of human osteosarcomas. Current understanding of the genes that contribute to the features of MDR is limited, and the mechanisms remain unclear. Here we applied differential display reverse transcription-polymerase chain reaction (DDRT-PCR) to parental and MDR-variants of U-2 OS human osteosarcoma cells, to clarify the genes involved in the MDR cells, and identified five candidate genes. These are BCRP (breast cancer resistance protein) encoding a transmembrane efflux pump; RB1CC1 (RB1-inducible coiled-coil 1), a tumor suppressor regulating RB1 (retinoblastoma 1) expression; a novel transcriptional variant of dUTPase; SSR2 (beta-signal sequence receptor), which is associated with protein translocation across ER membrane; and HSP105 encoding high molecular mass heat shock proteins. Molecular and biological characterization of these genes will yield further insight into the features between MDR and tumor progression in human osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/15138564', 'no', 'no', 'NA', 6),
(15144225, 'Tsai KS, Yang RS, Liu SH', 'Benzo[a]pyrene regulates osteoblast proliferation through an estrogen receptor-related cyclooxygenase-2 pathway.', 'Chem Res Toxicol', '2004', 'Benzo[a]pyrene regulates osteoblast proliferation through an estrogen receptor-related cyclooxygenase-2 pathway. Polycyclic aromatic hydrocarbons (PAHs) have been known as a kind of xenoestrogen. Benzo[a]pyrene, a PAH present in tobacco smoke and tar, has been implicated in the induction of cell proliferation as well as tumors including osteosarcoma. Nevertheless, the literature about the action of benzo[a]pyrene on the bone system is rare. It has been identified that osteoblasts owned the estrogen receptors and estrogen could modulate the osteoblast proliferation. In this study, we found that benzo[a]pyrene was capable of increasing the cell proliferation in cultured rat osteoblasts, human osteosarcoma cell line (MG-63), and estrogen sensitive human cell line (MCF-7) but not in the human estrogen receptor negative cell line (MDA-MB-231). This benzo[a]pyrene-induced osteoblast proliferation could be inhibited by the estrogen receptor antagonist ICI182780 and tamoxifen, PD98059 [extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) inhibitor], and LY294002 [phosphatidylinositol 3-kinase (PI3K) inhibitor] but not alpha-naphthoflavone (aryl hydrocarbon receptor antagonist) and SB203580 (p38 MAPK inhibitor). Western blot analysis showed that benzo[a]pyrene could induce the phosphorylation of ERK1/2 and Akt (PI3K downstream effector) in osteoblasts. The proliferating cell nuclear antigen protein levels in nuclear fraction of osteoblasts were also increased by benzo[a]pyrene. Moreover, cyclooxygenase-2 (COX-2), but not COX-1, expression could be induced in osteoblasts under benzo[a]pyrene treatment. Its upregulation was associated with the induction of prostaglandin E(2) (PGE(2)). COX-2 inhibitors NS398 and aspirin are capable of inhibiting the benzo[a]pyrene-induced osteoblast proliferation. These results indicate that benzo[a]pyrene may modulate the osteoblast proliferation through activation of COX-2 protein.', 'http://www.ncbi.nlm.nih.gov/pubmed/15144225', 'no', 'no', 'benzo[a]pyren|ICI182780|tamoxifen|PD98059|LY294002', 5),
(15145525, 'Kubista B, Erovic BM, Klinger H, Sulzbacher I, Trieb K', 'CD9 expression is not a prognostic factor in human osteosarcoma.', 'Cancer Lett', '2004', 'CD9 expression is not a prognostic factor in human osteosarcoma. CD 9, also known as Motility-Related Protein-1 (MRP-1), is a member of the transmembrane four superfamily and plays a crucial role in cell adhesion, motility and signalling events. Downregulation of CD 9 has been reported to be associated with tumour progression, metastasis and clinical outcome in various kinds of solid tumours. Although prognosis of osteosarcoma has been improved by chemotherapy during the last decades, the problem of non-responders remains. At the present time prognostic factors at diagnosis have not been clearly identified. Furthermore, there is a need for markers that predict the response to chemotherapy at the time of biopsy, allowing stratification of osteosarcoma patients. In this study we investigated the effect of CD9 expression on the response to chemotherapy and survival in osteosarcoma. The expression of CD9 was examined immunohistochemically in 52 patients with high grade osteosarcoma and the results were correlated with histologic response to chemotherapy, 5 year disease free and 5 year overall survival. In patients with osteosarcoma 22 of 52 cases (42%) were positive for CD 9 expression, the rest were negative. CD 9 expression status showed no statistically significant correlation with response to chemotherapy; 41% had a poor response and 59% a good response in the CD9 positive group. In the CD9 negative group 57% had a good and 47% had a bad response. No significant difference was found when comparing disease free survival (58.9% in CD9 positive- versus 69.3% in CD9 negative tumours; P = 0.99) and overall survival of patients (54.0% in CD9 positive- versus 58.1% in CD9 negative tumours; P = 0.90) with CD9 expressing tumours to those with reduced CD9 expression. In conclusion our findings suggest that in contrast to solid tumours, CD9 is unlikely to provide any additional prognostic information for clinical purposes in osteosarcoma patients.', 'http://www.ncbi.nlm.nih.gov/pubmed/15145525', 'yes', 'no', 'NA', 1),
(15167009, 'López-Guerrero JA, López-Ginés C, Pellín A, Carda C, Llombart-Bosch A', 'Deregulation of the G1 to S-phase cell cycle checkpoint is involved in the pathogenesis of human osteosarcoma.', 'Diagn Mol Pathol', '2004', 'Deregulation of the G1 to S-phase cell cycle checkpoint is involved in the pathogenesis of human osteosarcoma. Osteosarcoma (OS) displays complex karyotypes with numerical changes as well as structural abnormalities suggesting that several oncogenes and tumor suppressor genes may be implicated in the biology of OS. The aim of our study was to investigate the possible implication of the molecular alterations of the G1 to S-phase checkpoint genes in the pathogenesis of OS. We analyzed samples from 29 patients and found molecular alterations of the RB and TP53 genes in 6 (21%) and 3 (10%) cases, respectively. Homozygous deletion of the INK4A/ARF locus and methylation of INK4A was detected in 3 (10%) and 2 (7%) cases, respectively. CDK4 and MDM2 co-amplification was observed in 1 case (3%). Cyclin D3 is differentially expressed in a greater proportion than D1- and D2-type cyclins. Cytogenetically, all cases had complex karyotypes being especially significant the losses of the chromosomes 4, 13, and 17. As a whole, 11 of 29 (38%) analyzed OS presented alterations in some of the analyzed G1 to S-phase checkpoint genes. These alterations were more frequently present in adults (P = 0.032). All patients with genetic alterations in the G1/S-phase checkpoint died during their clinical follow-up, whereas more than 53% of the remaining cases were alive in this period (P = 0.007). Hence, in the pathogenesis of human OS, deregulation of the G1/S checkpoint genes, especially RB, TP53, and INK4/ARF locus, plays an important role and defines a subgroup of patients with a poor outcome.', 'http://www.ncbi.nlm.nih.gov/pubmed/15167009', 'yes', 'yes', 'NA', 7),
(15174116, 'Yu Y, Harris RI, Yang JL, Anderson HC, Walsh WR', 'Differential expression of osteogenic factors associated with osteoinductivity of human osteosarcoma cell lines.', 'J Biomed Mater Res A', '2004', 'Differential expression of osteogenic factors associated with osteoinductivity of human osteosarcoma cell lines. Differential expression of multiple osteogenic factors may be responsible for the different osteoinductivity of osteosarcoma cell lines. We compared in vivo osteoinductivity of human osteosarcoma cell lines (Saos-2 vs. U-2 OS) in nude mice, and their in vitro expression of various osteogenic factors of protein level by quantitative immunocytochemistry and mRNA level by RT-PCR and/or in situ hybridization. Saos-2 cells, but not U-2 OS, were osteoinductive in vivo. Significantly higher expression (independent t-test, all p < 0.005) of osteogenic factors were observed in Saos-2 cells compared with U-2 OS, which included bone morphogenetic proteins (particularly BMPs-2, 3, 4, and 7), transforming growth factor-beta (TGF-beta), BMP receptor (BMPR)-1A, receptor-regulated Smads (R-Smads), Smads 1, 2, and 5, and common-mediator Smad (Co-Smad), Smad 4. In contrast, U-2 OS cells expressed higher levels of inhibitory Smad 6 (I-Smad) protein than Saos-2 cells (p < 0.001). These results suggest that a combination of osteogenic factors (BMPs, TGF-beta, BMPRs, and R/Co-Smads) against I-Smad may play important roles in the Saos-2 cell osteoinductivity. This may have a clinical implication in selecting key osteogenic factors for combined therapy for bone defect diseases. The characterized cell lines can be used as positive and negative controls for the assessments of both in vitro and in vivo bone formation capabilities of designed tissues or biomaterials.Copyright 2004 Wiley Periodicals, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15174116', 'no', 'no', 'NA', 11),
(15193436, 'van Dartel M, Redeker S, Bras J, Kool M, Hulsebos TJ', 'Overexpression through amplification of genes in chromosome region 17p11.2 approximately p12 in high-grade osteosarcoma.', 'Cancer Genet Cytogenet', '2004', 'Overexpression through amplification of genes in chromosome region 17p11.2 approximately p12 in high-grade osteosarcoma. Osteosarcomas are malignant tumors of the bone that are characterized by complex genetic changes, including loss and amplification of chromosome regions. Region 17p11.2 approximately p12 is frequently found to be amplified in this tumor, suggesting the presence of an oncogene (or oncogenes) important in osteosarcoma tumorigenesis. We had previously determined amplification profiles for this region. Reasoning that amplification of a causative oncogene in a tumor should result in increased expression of that gene, we have now determined the expression status of genes and expressed sequence tags (ESTs) in 17p11.2 approximately p12. We constructed a 17p11.2 approximately p12-specific macroarray containing 40 genes and 21 ESTs from this region, which was used for expression profiling of 11 osteosarcoma samples (9 tumors and 2 cell lines) and of normal human osteoblasts. Compared to normal osteoblasts, genes with at least threefold increased expression were considered to be overexpressed in the tumor. Genes PMP22 and COPS3, EST AA126939 (encoding part of the hypothetical protein FLJ20343), and two anonymous ESTs (AA918483 and R02360) were found to be most consistently overexpressed after amplification. By real-time reverse transcriptase polymerase chain reaction, we could confirm the overexpression status of PMP22 and COPS3 but not of FLJ20343. We conclude that PMP22 and COPS3, and possibly also the three ESTs, are candidate amplification targets in 17p11.2 approximately p12 in osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15193436', 'yes', 'no', 'NA', 3),
(15198903, 'Liao WM, Zhang CL, Li FB, Zeng BF, Zeng YX', 'p21WAF1/CIP1 gene DNA sequencing and its expression in human osteosarcoma.', 'Chin Med J (Engl)', '2004', 'p21WAF1/CIP1 gene DNA sequencing and its expression in human osteosarcoma. Mutation and expression change of p21WAF1/CIP1 may play a role in the growth of osteosarcoma. This study was to investigate the expression of the p21WAF1/CIP1 gene in human osteosarcoma, p21WAF1/CIP1 gene DNA sequence change and their relationships with the phenotype and clinical prognosis.p21WAF1/CIP1 gene in 10 normal people and the tumours of 45 osteosarcoma patients were examined using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) with silver staining. The PCR product with an abnormal strand was sequenced directly. The p21WAF1/CIP1 gene mRNA and P21 protein of 45 cases of osteosarcoma were investigated by using in situ hybridization and immunohistochemistry, respectively.The occurrence of P21 protein in osteosarcoma was 17.78% (8/45), and p21WAF1/CIP1 mRNA expression in osteosarcoma was 42.22% (19/45). The p21WAF1/CIP1 gene DNA sequencing of amplified production showed that in p21WAF1/CIP1 gene exon 3 of 36 cases of human osteosarcoma, there were 17 cases (47.22%) with C-->T at position 609; 10 normal blood samples'' DNA sequence analysis yielded 8 cases (80.00%) with C-->T at the same position.Along with the increase of malignancy, the expression of p21WAF1/CIP1mRNA and P21 protein in osteosarcoma tends to decrease. It is uncommon for the p21WAF1/CIP1 gene mutation to occur in human osteosarcoma. As a result, the possible existence of tumour subtypes of p21WAF1/CIP1 gene mutation should be investigated. Our research leads to the location of p21WAF1/CIP1 gene polymorphism of Chinese osteosarcoma patients, which can provide a basis for further research.', 'http://www.ncbi.nlm.nih.gov/pubmed/15198903', 'yes', 'no', 'NA', 1),
(15201972, 'Lee WI, Bacchni P, Bertoni F, Maeng YH, Park YK', 'Quantitative assessment of HER2/neu expression by real-time PCR and fluorescent in situ hybridization analysis in low-grade osteosarcoma.', 'Oncol Rep', '2004', 'Quantitative assessment of HER2/neu expression by real-time PCR and fluorescent in situ hybridization analysis in low-grade osteosarcoma. Low-grade osteosarcoma is a rare variant of osteosarcoma. Although malignant, it must be distinguished from conventional osteosarcomas because of its excellent prognosis. Numerous published papers have described the expression of HER2/neu oncogene in osteosarcoma as a poor prognostic factor; however their results are discordant. To address the expression of HER2/neu and to validate the assessment methods of amplification of the HER2/neu oncogene, the authors have employed quantitative real-time PCR and fluorencent in situ hybridization analysis (FISH) in 21 low-grade osteosarcomas. We calculated the quantification of HER2/neu oncogene amplification as the ratio of measured HER2/neu gene/beta-globin reference gene in real-time PCR. All 21 cases had amplified signals in the quantitative real-time PCR. However, in the FISH analysis, HER2/neu oncogene amplification was only identified in 26% (5/19). The exact reasons for the discordance between these two methods are unknown; however, variable histological features might play a potential role. In conclusion, as our study showed amplification of HER2/neu oncogene in low-grade osteosarcoma, we assume that expression of HER2/neu is not a poor prognostic factor in low-grade osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15201972', 'yes', 'no', 'NA', 1),
(15205094, 'Dekairelle AF, Brichard B, Delhez H, Gala JL', 'Identification of a transcriptionally inactive p53 mutant by functional analysis of separated alleles in yeasts (FASAY) in a child osteosarcoma tumor: a case report.', 'Pediatr Hematol Oncol', '2004', 'Identification of a transcriptionally inactive p53 mutant by functional analysis of separated alleles in yeasts (FASAY) in a child osteosarcoma tumor: a case report. Tumor suppressor gene p53 is one of the most specific genetic alterations occurring in osteosarcoma pathogenesis. It is thought to be an early and key step in the tumorigenesis of osteosarcoma. However, whether the p53 status is a marker predictive of response to therapy and a marker of prognostic value remains controversial. The choice of p53 status detection method certainly account for discrepancies. The authors used a simple functional assay (functional analysis of separated alleles in yeast) on the tumor sample of an 8-year-old girl presenting with an osteosarcoma of the tibia. While making it possible to exclude the presence of a germline mutation, FASAY indicated the presence of a somatic p53 mutation lacking transcriptional activity on p21 and bax target genes. FASAY also strongly suggested a loss of heterozygosity p53, which was confirmed by cytogenetic analysis. Sequencing of cDNA extracted from yeast colonies containing mutated p53 identified a 213 stop mutation in exon 6. Despite these p53 alterations, the child is still in complete remission after a follow-up of 48 months.', 'http://www.ncbi.nlm.nih.gov/pubmed/15205094', 'yes', 'yes', 'NA', 1),
(15207713, 'Kim M, Sgagias M, Deng X, Jung YJ, Rikiyama T, Lee K, Ouellette M, Cowan K', 'Apoptosis induced by adenovirus-mediated p14ARF expression in U2OS osteosarcoma cells is associated with increased Fas expression.', 'Biochem Biophys Res Commun', '2004', 'Apoptosis induced by adenovirus-mediated p14ARF expression in U2OS osteosarcoma cells is associated with increased Fas expression. The INK4A/ARF locus on chromosome 9 is a tumor suppressor gene frequently mutated in human cancers. In order to study the effects of p14ARF expression in tumor cells, we constructed a recombinant adenovirus containing p14ARF cDNA (Adp14ARF). Adp14ARF infection of U2OS osteosarcoma cells which has wild type p53 and mutant p14ARF revealed high levels of p14 (ARF) expression within 24h. In addition, Adp14ARF-mediated expressing of p14 (ARF) was associated with increased levels of p53, p21, and mdm2 protein. Growth inhibition assays following Adp14ARF infection demonstrated that the growth of U2OS cells was inhibited relative to infection with control virus. Furthermore, TUNEL analysis as well as PARP cleavage assays demonstrated that Adp14ARF infection was associated with increased apoptosis in U2OS cell line and that it was associated with Adp14ARF induced overexpression of Fas and Fas-L. Addition of Fas-L neutralizing antibody NOK-1 decreased Adp14-mediated cell death, indicating that p14 (ARF) induction of the Fas pathway is associated with increased apoptosis. The finding that Adp14ARF infection did not induce Fas expression in U2OS/E6 and MCF/E6 cells suggests that wild type p53 expression may be necessary for Adp14ARF-mediated induction of Fas. The observation that overexpression of p53 by Adp53 infection in MCF-7 does not induce increased Fas protein levels nor apoptotic cell death suggests that p53 overexpression is required but not sufficient enough for apoptosis. These studies suggest there are other mechanisms other than induction of p53 in ARF-mediated apoptosis and gene therapy using Adp14ARF may be a promising treatment option for human cancers containing wild type p53 and mutant or deleted p14 expression.', 'http://www.ncbi.nlm.nih.gov/pubmed/15207713', 'no', 'no', 'NA', 6),
(15210853, 'Wang D, Luo M, Kelley MR', 'Human apurinic endonuclease 1 (APE1) expression and prognostic significance in osteosarcoma: enhanced sensitivity of osteosarcoma to DNA damaging agents using silencing RNA APE1 expression inhibition.', 'Mol Cancer Ther', '2004', 'Human apurinic endonuclease 1 (APE1) expression and prognostic significance in osteosarcoma: enhanced sensitivity of osteosarcoma to DNA damaging agents using silencing RNA APE1 expression inhibition. Osteosarcoma is the most common highly malignant bone tumor with primary appearance during the second and third decade of life. It is associated with a high risk of relapse, possibly resulting from a developed resistance to chemotherapy agents. As a means to overcome osteosarcoma tumor cell resistance and/or to sensitize tumor cells to currently used chemotherapeutic treatments, we examined the role of human apurinic endonuclease 1 (APE1) in osteosarcoma tumor cell resistance and prognosis. Sixty human samples of archived conventional (intramedullary) osteosarcoma were analyzed. APE1 protein was elevated in 72% of these tissues and among those with a known clinical outcome, there was a significant correlation between high APE1 expression levels and reduced survival times. The remaining 28% of samples showed low expression of APE1. Given that APE1 was overexpressed in osteosarcoma, we decreased APE1 levels using silencing RNA (siRNA) targeting technology in the osteosarcoma cell line, human osteogenic sarcoma (HOS), to enhance chemo- and radiation sensitivity. Using siRNA targeted technology of APE1, protein levels were reduced by more than 90% within 24 hours, remained low for 72 hours, and returned to normal levels at 96 hours. There was also a clear loss of APE1 endonuclease activity following APE1-siRNA treatment. A decrease in APE1 levels in siRNA-treated human osteogenic sarcoma cells led to enhanced cell sensitization to the DNA damaging agents: methyl methanesulfonate, H(2)O(2), ionizing radiation, and chemotherapeutic agents. The findings presented here have both prognostic and therapeutic implications for treating osteosarcoma. The APE1-siRNA results demonstrate the feasibility for the therapeutic modulation of APE1 using a variety of molecules and approaches.', 'http://www.ncbi.nlm.nih.gov/pubmed/15210853', 'yes', 'yes', 'methyl methanesulfonate', 1),
(15211578, 'Gillette JM, Chan DC, Nielsen-Preiss SM', 'Annexin 2 expression is reduced in human osteosarcoma metastases.', 'J Cell Biochem', '2004', 'Annexin 2 expression is reduced in human osteosarcoma metastases. Osteosarcoma is an aggressive primary bone cancer affecting primarily children and young adults. The development of valuable diagnostic indicators and therapeutic agents will be enhanced by the identification and characterization of genes that contribute to its aggressive behavior. We used representational difference analysis to isolate genes differentially expressed between primary human osteosarcoma tumors and subsequent metastatic lung lesions to identify genes potentially involved in metastatic potential. Several genes were differentially expressed between the two tumor populations, including annexin2. The levels of annexin2 mRNA and protein inversely correlated with metastatic potential in a subset of human osteosarcoma tumor specimens, as well as in a human osteosarcoma cell line selected for increased metastatic potential. Annexin2 has been described in several cellular localizations with various functional implications, many of which may be relevant to metastatic potential. Therefore, the subcellular localization of endogenous annexin2 protein was evaluated biochemically by subcellular fractionation and immunologically by flow cytometry and immunofluorescence in osteoblastic cells. Annexin2 was localized to the cytoplasm and intracellular aspect of the plasma membrane, excluded from the nucleus and undetectable on the cell surface or in the conditioned medium. Overexpression of annexin2 in osteosarcoma cells did not alter several in vitro phenotypes often used to assess metastatic potential including motility, adhesion, and proliferation. However, our previous data have implicated annexin2 in the mineralization process of osteoblastic cells in vitro. Consistent with an increase in differentiation-induced mineralization, there was diminished tumorigenicity and experimental metastatic potential of osteosarcoma cells overexpressing annexin2. These data suggest that annexin2 may downregulate osteosarcoma aggressiveness by inducing a more differentiated state in osteoblastic cells.Copyright 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15211578', 'yes', 'no', 'NA', 1),
(15221963, 'Nishijo K, Nakayama T, Aoyama T, Okamoto T, Ishibe T, Yasura K, Shima Y, Shibata KR, Tsuboyama T, Nakamura T, Toguchida J', 'Mutation analysis of the RECQL4 gene in sporadic osteosarcomas.', 'Int J Cancer', '2004', 'Mutation analysis of the RECQL4 gene in sporadic osteosarcomas. Osteosarcoma (OS) is the most prevalent malignant tumor among cases of Rothmund-Thomson syndrome (RTS) with germline mutations of the RECQL4 gene, a member of the RecQ helicase family. We investigated the involvement of the RECQL4 gene in the development of OS unrelated to RTS. RECQL4 mRNA was detected in 9 of 9 OS cell lines by Northern blotting and 26 of 26 OS tumors by RT-PCR. Direct sequencing of the entire coding region along with flanking splice junctions and 13 small (< 100 bp) introns in 71 OS tumors revealed 2 sites with a single-base change causing an amino acid change (G1814A for R355Q and C2474T for P441S) and one site with a 6 bp inframe deletion (4837-42delTGCACC for CT857-8del). Identical genotypes were found in corresponding normal tissues in all cases, and the frequency of each allele was not significantly different between OS and control populations. Our data indicate that the RECQL4 gene is not a frequent target for somatic mutations in sporadic OS unrelated to RTS.Copyright 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15221963', 'yes', 'no', 'NA', 1),
(15237589, 'Park HR, Kim YW, Park JH, Maeng YH, Nojima T, Hashimoto H, Park YK', 'Low expression of p63 and p73 in osteosarcoma.', 'Tumori', 'NA', 'Low expression of p63 and p73 in osteosarcoma. The recent discovery of two p53-related genes, p63 and p73, has revealed an additional level of complexity to the study of p53 function. Both genes encode multiple proteins arising from alternative promoter usage and splicing, with transactivation, DNA-binding, and tetramerization domains. Recent data support a role for p63 in squamous and transitional cell carcinomas, as well as in certain lymphomas and thymomas.To characterize the involvement of p63 and p73 in the development of osteosarcoma, we analyzed the expression and mutation of TAp63 and TAp73 in six osteosarcoma cell lines and twelve osteosarcoma specimens.Semiquantitative DNA/PCR analysis revealed that eight (67%) and six (50%) out of twelve osteosarcoma specimens showed significantly reduced levels of p63 and p73 transcription, respectively. Direct sequencing of the entire coding region detected no mutations in cell lines or osteosarcoma specimens.Our data suggest that low expression of p63 and p73 is relatively common in osteosarcomas and might contribute to their molecular pathogenesis.', 'http://www.ncbi.nlm.nih.gov/pubmed/15237589', 'yes', 'no', 'NA', 2);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(15247015, 'Trougakos IP, Gonos ES', 'Functional analysis of clusterin/apolipoprotein J in cellular death induced by severe genotoxic stress.', 'Ann N Y Acad Sci', '2004', 'Functional analysis of clusterin/apolipoprotein J in cellular death induced by severe genotoxic stress. Clusterin/apolipoprotein J (CLU) is a secreted heterodimeric glycoprotein that is reportedly upregulated during tumorigenesis, as well as during cell injury or death. Despite extensive efforts, CLU function during cellular death remains largely elusive. We are using as a model system to study CLU function three human osteosarcoma (OS) cell lines, namely, Sa OS, KH OS, and U-2 OS cells, induced to die after exposure to severe genotoxic stress mediated by the chemotherapeutic drug doxorubicin (DXR). We initially applied small interfering RNA (siRNA)-mediated specific knockdown of the CLU protein in OS cells. In all three cell lines, CLU knockdown resulted in increased sensitization to DXR-induced apoptosis. Supportively, moderate levels of forced transgene-mediated CLU stable overexpression in KH OS cells could rescue them from DXR-mediated apoptosis. In contrast, stable overexpression of high CLU levels in Sa OS and U-2 OS cells augmented apoptosis induced by cell exposure to severe DXR-mediated genotoxic stress. In summary, our data provide evidence that, although CLU is essential for cellular homeostasis, it may become highly cytotoxic in certain cellular contexts when it accumulates in high amounts intracellularly either by direct synthesis or by uptake from the extracellular milieu.', 'http://www.ncbi.nlm.nih.gov/pubmed/15247015', 'no', 'no', 'doxorubicin', 1),
(15250258, 'Liu Y, Zheng QX, Du JY, Yang SH, Shao ZW, Xiao BJ', 'Effects of TGF beta1 autocrine blockage on osteosarcoma cells.', 'Chin Med Sci J', '2004', 'Effects of TGF beta1 autocrine blockage on osteosarcoma cells. To evaluate the effects of transforming growth factor beta1 (TGFbeta1) autocrine blockage on proliferation activity and drug sensitivity of osteosarcoma. METHODS; Northern blot, MTT determination, and 3H thymidine incorporation were used to investigate the effects of antisense TGF beta1 gene on osteosarcoma.The proliferation of osteosarcoma cells transfected by antisense TGF beta1 gene was suppressed markedly, and adriamycin sensitivity was significantly increased.Blockage of osteosarcoma cells TGF beta1 autocrine loop inhibits cell proliferation and enhances chemotherapy sensitivity.', 'http://www.ncbi.nlm.nih.gov/pubmed/15250258', 'no', 'no', 'NA', 1),
(15262428, 'van Dartel M, Hulsebos TJ', 'Characterization of PMP22 expression in osteosarcoma.', 'Cancer Genet Cytogenet', '2004', 'Characterization of PMP22 expression in osteosarcoma. The peripheral myelin protein (PMP22) gene is highly expressed in peripheral Schwann cells and encodes an important constituent of the myelin sheath. It is also expressed at lower levels in other normal tissues in which the protein is supposed to be involved in cell growth regulation. We recently reported frequent amplification and overexpression of PMP22 in high-grade osteosarcoma. Here, we analyzed PMP22 expression in five osteosarcoma tumors and three osteosarcoma cell lines. In normal Schwann cells, transcription of PMP22 starts at three promoters, P1A, P1B, and P2, which results in the synthesis of three alternatively spliced transcripts that all code for the same protein. We found a comparable expression pattern in normal osteoblasts. However, promoter P1A-driven transcripts were absent in all investigated tumors and cell lines and, compared to normal osteoblasts, the P1B/P2 transcript ratio was found to be increased in two of three cases with PMP22 overexpression and decreased in all five cases without overexpression. In normal Schwann cells and in NIH3T3 cells, PMP22 expression increases upon serum starvation-induced growth arrest. In contrast to this, serum withdrawal caused a considerable decrease of PMP22 expression in the osteosarcoma cell lines. We conclude that the different PMP22 expression in osteosarcoma may result in alternative availability of the PMP22 protein during the cell cycle and aberrant regulation of cell growth control in osteosarcoma tumorigenesis.', 'http://www.ncbi.nlm.nih.gov/pubmed/15262428', 'yes', 'no', 'NA', 1),
(15268905, 'Kapur S, Chen ST, Baylink DJ, Lau KH', 'Extracellular signal-regulated kinase-1 and -2 are both essential for the shear stress-induced human osteoblast proliferation.', 'Bone', '2004', 'Extracellular signal-regulated kinase-1 and -2 are both essential for the shear stress-induced human osteoblast proliferation. Extracellular signal-regulated kinases (Erk)-1 and -2 are key mediators of various mitogenic signaling pathways, including mechanical stress-induced osteoblast proliferation. Mechanical stimuli, such as flow shear stress, simultaneously activate both Erk-1 and -2 in osteoblasts, resulting in stimulation of osteoblast proliferation. This study sought to test whether Erk-1, -2, or both are essential for the fluid flow shear stress-induced osteoblast proliferation. Moloney leukemia virus (MLV)-based vectors expressing wild-type (wt)- or kinase-deficient (kd) Erk-1 and Erk-2, respectively, were constructed and used to transduce human TE85 osteosarcoma cells with an MOI of 30. An MLV-red fluorescent protein (RFP) vector was included as a control. Effects of Erk-1 and -2 overexpression on cell proliferation in response to a 30-min constant fluid flow shear stress at 20 dynes/cm2 were determined with [3H]thymidine incorporation 24 h after the shear stress. The MLV-Erk vector-transduced TE85 cells showed a >10- and approximately 2-fold overexpression of Erk-1 and -2 protein, respectively. The RFP expressing control cells and the parental TE85 cells each showed an approximately twofold increase (P < 0.01) in [3H]thymidine incorporation in response to the shear stress. Cells overexpressing wt-Erk-1 or -2 showed small enhancing effects on the response to the shear stress in the increases in [3H]thymidine incorporation and cell number. Conversely, overexpression of kd-Erk-1 or -2 each alone completely abolished the shear stress-induced osteoblast proliferation. Overexpression of either kd-Erk-1 or kd-Erk-2 alone did not have a significant effect on basal osteoblast proliferation, suggesting that the Erk signaling pathway may not be essential for basal cell proliferation. In summary, this study demonstrates for the first time that Erk-1 and -2 are both required for the mitogenic response to fluid flow shear stress in human osteoblasts and that blocking Erk-1 or -2 each alone is sufficient to completely block the mitogenic response to shear stress-induced proliferation.', 'http://www.ncbi.nlm.nih.gov/pubmed/15268905', 'no', 'no', 'NA', 2),
(15289892, 'Takahashi T, Ogawa Y, Kobayashi T, Sonobe H, Seguchi H, Tani T, Yoshida S', 'A combination of selective COX-2 inhibitors and hydrogen peroxide increase the reactive oxygen species formation in osteosarcoma cells after X-ray irradiation.', 'Int J Mol Med', '2004', 'A combination of selective COX-2 inhibitors and hydrogen peroxide increase the reactive oxygen species formation in osteosarcoma cells after X-ray irradiation. We investigated whether a combination of selective COX-2 inhibitors and hydrogen peroxide increases the effect of X-ray irradiation, with regard to reactive oxygen species (ROS) formation in an osteosarcoma cell line. COX-2 inhibitor did not induce ROS formation when combined with irradiation. A low dose concentration of COX-2 inhibitor in combination with hydrogen peroxide and irradiation did affect ROS formation in the intracellular compartment; however, this same combination of agents at high doses did not modulate the effect of irradiation. Therefore, low doses of COX-2 inhibitor and hydrogen peroxide together, in combination with irradiation, is a potentially useful alternative form of radiotherapy for apoptotic-resistant neoplasms such as osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15289892', 'no', 'no', 'hydrogen peroxide', 1),
(15298715, 'Man TK, Lu XY, Jaeweon K, Perlaky L, Harris CP, Shah S, Ladanyi M, Gorlick R, Lau CC, Rao PH', 'Genome-wide array comparative genomic hybridization analysis reveals distinct amplifications in osteosarcoma.', 'BMC Cancer', '2004', 'Genome-wide array comparative genomic hybridization analysis reveals distinct amplifications in osteosarcoma. Osteosarcoma is a highly malignant bone neoplasm of children and young adults. It is characterized by extremely complex karyotypes and high frequency of chromosomal amplifications. Currently, only the histological response (degree of necrosis) to therapy represent gold standard for predicting the outcome in a patient with non-metastatic osteosarcoma at the time of definitive surgery. Patients with lower degree of necrosis have a higher risk of relapse and poor outcome even after chemotherapy and complete resection of the primary tumor. Therefore, a better understanding of the underlying molecular genetic events leading to tumor initiation and progression could result in the identification of potential diagnostic and therapeutic targets.We used a genome-wide screening method - array based comparative genomic hybridization (array-CGH) to identify DNA copy number changes in 48 patients with osteosarcoma. We applied fluorescence in situ hybridization (FISH) to validate some of amplified clones in this study.Clones showing gains (79%) were more frequent than losses (66%). High-level amplifications and homozygous deletions constitute 28.6% and 3.8% of tumor genome respectively. High-level amplifications were present in 238 clones, of which about 37% of them showed recurrent amplification. Most frequently amplified clones were mapped to 1p36.32 (PRDM16), 6p21.1 (CDC5L, HSPCB, NFKBIE), 8q24, 12q14.3 (IFNG), 16p13 (MGRN1), and 17p11.2 (PMP22 MYCD, SOX1,ELAC27). We validated some of the amplified clones by FISH from 6p12-p21, 8q23-q24, and 17p11.2 amplicons. Homozygous deletions were noted for 32 clones and only 7 clones showed in more than one case. These 7 clones were mapped to 1q25.1 (4 cases), 3p14.1 (4 cases), 13q12.2 (2 cases), 4p15.1 (2 cases), 6q12 (2 cases), 6q12 (2 cases) and 6q16.3 (2 cases).This study clearly demonstrates the utility of array CGH in defining high-resolution DNA copy number changes and refining amplifications. The resolution of array CGH technology combined with human genome database suggested the possible target genes present in the gained or lost clones.', 'http://www.ncbi.nlm.nih.gov/pubmed/15298715', 'yes', 'no', 'NA', 10),
(15310147, 'Park HR, Jung WW, Bertoni F, Bacchini P, Park JH, Kim YW, Park YK', 'Molecular analysis of p53, MDM2 and H-ras genes in low-grade central osteosarcoma.', 'Pathol Res Pract', '2004', 'Molecular analysis of p53, MDM2 and H-ras genes in low-grade central osteosarcoma. Low-grade central osteosarcoma is an uncommon form that is characterized by a long premorbid history, and is compatible with prolonged survival after treatment. However, molecular abnormalities are rare in low-grade central osteosarcomas, whereas p53 mutations occur in approximately 20% of conventional high-grade osteosarcomas. In this study, 21 cases of low-grade central osteosarcoma were analyzed for mutations of the p53 gene, amplification of the MDM2 gene, and mutations of the H-ras gene using formalin-fixed, paraffin-embedded materials. We also examined the expression of p53, MDM2, and p21WAF1 protein immunohistochemically and assessed the proliferation activities using the monoclonal antibody MIB-1. One case (4.7%) showed strong p53 immunoreactivity, whereas p53 gene mutations were not detected at all. Seven cases (33.3%) showed immunoreactivity for MDM2 protein. As for gene alterations, MDM2 amplification was found in four cases (19.0%). p21WAF1 expression was detected in 12 cases (57.1%). MIB-1-LI showed very low levels in all the cases and no significant correlation with p53 or MDM2 immuno-reactivity. None of the tumors showed H-ras mutations. In conclusion, the number of p53 gene alterations in low-grade central osteosarcomas is lower than that in conventional high-grade osteosarcomas. MDM2 alterations and p21WAF1 expression might be involved in the tumorigenesis of low-grade central osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/15310147', 'yes', 'no', 'NA', 4),
(15313187, 'Jacobson A, Johansson S, Branting M, Melhus H', 'Vitamin A differentially regulates RANKL and OPG expression in human osteoblasts.', 'Biochem Biophys Res Commun', '2004', 'Vitamin A differentially regulates RANKL and OPG expression in human osteoblasts. All-trans-retinoic acid (ATRA) induces bone resorption, but the molecular mechanisms are unknown. We have studied the effect of ATRA on osteoprotegerin (OPG) and receptor activator of NF-kappaB ligand (RANKL) expression in human MG-63 osteosarcoma cells and primary osteoblast-like cultures. ATRA dose-dependently down-regulated protein levels of OPG in MG-63 cells, with a maximum (-56%) observed at a dose of 10(-6)M. This effect was confirmed with quantitative real-time PCR, where OPG mRNA was decreased after 4h (-68%) in primary cultures and after 8h (-87%) in MG-63 cells. The reduction in OPG expression was inhibited by a retinoic acid receptor (RAR)-antagonist and was mimicked by a RARbeta,gamma-agonist, indicating that the ATRA effect is mediated by these receptors. In primary cultures we found a threefold induction of RANKL mRNA expression. Thus, the RANKL/OPG ratio was markedly increased, suggesting a potential mechanism of ATRA-induced bone resorption.', 'http://www.ncbi.nlm.nih.gov/pubmed/15313187', 'no', 'no', 'vitamine A', 2),
(15314286, 'Roh MS, Kim CW, Park BS, Kim GC, Jeong JH, Kwon HC, Suh DJ, Cho KH, Yee SB, Yoo YH', 'Mechanism of histone deacetylase inhibitor Trichostatin A induced apoptosis in human osteosarcoma cells.', 'Apoptosis', '2004', 'Mechanism of histone deacetylase inhibitor Trichostatin A induced apoptosis in human osteosarcoma cells. Although histone deacetylase (HDAC) inhibitors are emerging as a promising new treatment strategy in malignancy, how they exert their effect on osteosarcoama cells is as yet unclear. This study was undertaken to investigate the underlying mechanism of a HDAC inhibitor Trichostatin A (TSA)-induced apoptosis in a osteosarcoma cell line HOS. We observed that TSA treatment decreased the viability of the cells and prominently increased acetylation of histone H3. Evidence was obtained indicating that TSA induced apoptosis of HOS cells as follows: (1) Generation of DNA fragmentation; (2) activation of procaspase-3; (3) cleavage of PARP; and (4) increase of DNA hypoploidy. The reduction of MMP and the release of cytochrome c to cytosol were also shown, indicating that TSA induces apoptosis in HOS cells in a histone acetylation- and mitochondria-dependent fashions. We also examined whether TSA can sensitize HOS cells to the action of an antitumor agent genistein. The combination therapy of TSA and genistein showed synergistic anticancer effect indicating that TSA can be considered as a novel therapeutic strategy for osteosarcoma not only from its direct apoptosis-inducing activity but also from the possibility of sensitization to other antitumor agents.', 'http://www.ncbi.nlm.nih.gov/pubmed/15314286', 'no', 'no', 'trIHCostatin A|genistein', 4),
(15318945, 'Pedersen KB, Andersen K, Fodstad Ø, Maelandsmo GM', 'Sensitization of interferon-gamma induced apoptosis in human osteosarcoma cells by extracellular S100A4.', 'BMC Cancer', '2004', 'Sensitization of interferon-gamma induced apoptosis in human osteosarcoma cells by extracellular S100A4. S100A4 is a small Ca2+-binding protein of the S100 family with metastasis-promoting properties. Recently, secreted S100A4 protein has been shown to possess a number of functions, including induction of angiogenesis, stimulation of cell motility and neurite extension.Cell cultures from two human osteosarcoma cell lines, OHS and its anti-S100A4 ribozyme transfected counterpart II-11b, was treated with IFN-gamma and recombinant S100A4 in order to study the sensitizing effects of extracellular S100A4 on IFN-gamma mediated apoptosis. Induction of apoptosis was demonstrated by DNA fragmentation, cleavage of poly (ADP-ribose) polymerase and Lamin B.In the present work, we found that the S100A4-expressing human osteosarcoma cell line OHS was more sensitive to IFN-gamma-mediated apoptosis than the II-11b cells. S100A4 protein was detected in conditioned medium from OHS cells, but not from II-11b cells, and addition of recombinant S100A4 to the cell medium sensitized II-11b cells to apoptosis induced by IFN-gamma. The S100A4/IFN-gamma-mediated induction of apoptosis was shown to be independent of caspase activation, but dependent on the formation of reactive oxygen species. Furthermore, addition of extracellular S100A4 was demonstrated to activate nuclear factor-kappa B (NF-kappa B).In conclusion, we have shown that S100A4 sensitizes osteosarcoma cells to IFN-gamma-mediated induction of apoptosis. Additionally, extracellular S100A4 activates NF-kappa B, but whether these events are causally related remains unknown.', 'http://www.ncbi.nlm.nih.gov/pubmed/15318945', 'no', 'no', 'NA', 3),
(15325100, 'van Dartel M, Hulsebos TJ', 'Amplification and overexpression of genes in 17p11.2 ~ p12 in osteosarcoma.', 'Cancer Genet Cytogenet', '2004', 'Amplification and overexpression of genes in 17p11.2 ~ p12 in osteosarcoma. We summarize and briefly discuss recent findings with respect to the amplification and overexpression of candidate oncogenes in 17p11.2 ~p12 in high-grade osteosarcomas. Amplification of this region occurs in about 25% of cases. The amplification profiles are often complex and suggest the involvement of more than one oncogene. The 17p11.2 ~ p12 region harbors many low-copy repeats (LCRs). We propose LCR-mediated repeated duplication by mitotic nonallelic homologous recombination as mechanism for the generation of the amplifications in this region. Genes PMP22 and COPS3 and three expressed sequence tags from within 17p11.2 ~ p12 have been found to be frequently overexpressed and consistently overexpressed after amplification, which identifies them as candidate oncogenes in this region. Overexpression of COPS3 has been linked to TP53 protein degradation and, being equivalent to TP53 mutation, the induction of genomic instability, which frequently occurs in high-grade osteosarcoma. These findings may serve as a framework for future work aimed to identify the causative oncogenes in 17p11.2 ~p12, to clarify the mechanism of their amplification, and to determine their importance in osteosarcoma tumorigenesis.', 'http://www.ncbi.nlm.nih.gov/pubmed/15325100', 'yes', 'no', 'NA', 3),
(15346048, 'Hoang BH, Dyke JP, Koutcher JA, Huvos AG, Mizobuchi H, Mazza BA, Gorlick R, Healey JH', 'VEGF expression in osteosarcoma correlates with vascular permeability by dynamic MRI.', 'Clin Orthop Relat Res', '2004', 'VEGF expression in osteosarcoma correlates with vascular permeability by dynamic MRI. Dynamic enhanced magnetic resonance imaging has been used to assess tumor angiogenesis in osteosarcoma. Vascular endothelial growth factor has been shown to correlate with pulmonary metastasis and a poor prognosis in osteosarcoma. The purpose of this investigation was to determine whether vascular endothelial growth factor expression in osteosarcoma correlates with vascular permeability detected by dynamic enhanced magnetic resonance imaging and to explore the role of dynamic enhanced magnetic resonance imaging as a noninvasive means of assessing tumor angiogenic activity. Fifty-five osteosarcoma patients with osteosarcoma enrolled in a treatment protocol that included dynamic enhanced magnetic resonance imaging. In 15 patients, tumor tissues were available for vascular endothelial growth factor immunohistochemical studies. A two-compartment model used the exchange rate constants (kep) between the plasma and tumor compartments to quantify vascular permeability during dynamic magnetic resonance imaging studies. Immunohistochemical staining for vascular endothelial growth factor was graded according to the intensity and number of positively stained cells. Vascular endothelial growth factor-positive tumors showed higher mean vascular permeability when compared with vascular endothelial growth factor-negative tumors. Vascular permeability also correlated with increasing vascular endothelial growth factor expression. The preliminary results in this study show an association between vascular endothelial growth factor and dynamic MR signal enhancement in osteosarcoma. Dynamic enhanced magnetic resonance imaging should be investigated as a means to prognosticate osteosarcoma patients with osteosarcoma according to their tumor angiogenic activity.', 'http://www.ncbi.nlm.nih.gov/pubmed/15346048', 'yes', 'yes', 'NA', 1),
(15365076, 'Sanders RP, Drissi R, Billups CA, Daw NC, Valentine MB, Dome JS', 'Telomerase expression predicts unfavorable outcome in osteosarcoma.', 'J Clin Oncol', '2004', 'Telomerase expression predicts unfavorable outcome in osteosarcoma. Osteosarcoma is distinct from most cancers in that the majority of osteosarcomas lack telomerase expression and use the alternative lengthening of telomeres (ALT) mechanism to maintain telomeres. Laboratory studies suggest that compared with ALT, telomerase expression is associated with increased tumor aggressiveness. We evaluated the clinical significance of telomerase expression in human osteosarcoma.Fifty-six osteosarcomas from 51 patients treated at St Jude Children''s Research Hospital between 1982 and 2003 were evaluated for telomerase enzyme activity, mRNA expression of the catalytic component of telomerase (TERT), and presence of the ALT pathway.Outcome analysis was based on TERT mRNA expression in the primary tumor samples from 44 patients. Fourteen primary tumors expressed TERT mRNA (32%; eight TERT only, six TERT and ALT) and 30 did not express TERT mRNA (68%; 29 ALT, one no ALT). Progression-free survival (PFS) was inferior in the TERT-positive group compared with the TERT-negative group (3-year estimates, 21.4% +/- 9.5% v 63.7% +/- 11.1%; P =.014). Likewise, overall survival was inferior in the TERT-positive group compared with the TERT-negative group (3-year estimates, 42.9% +/- 12.2% v 70.0% +/- 9.9%; P =.031). Among 31 patients with nonmetastatic disease at diagnosis, PFS was lower in the TERT-positive group compared with the TERT-negative group (3-year estimates, 33.3% +/- 13.6% v 72.0% +/- 11.5%; P =.092).Telomerase expression in primary tumor samples is associated with decreased PFS and OS in patients with osteosarcoma. Additional studies are warranted to better define the clinical utility of this molecular marker.', 'http://www.ncbi.nlm.nih.gov/pubmed/15365076', 'yes', 'yes', 'NA', 1),
(15370596, 'Ferrari C, Benassi S, Ponticelli F, Gamberi G, Ragazzini P, Pazzaglia L, Balladelli A, Bertoni F, Picci P', 'Role of MMP-9 and its tissue inhibitor TIMP-1 in human osteosarcoma: findings in 42 patients followed for 1-16 years.', 'Acta Orthop Scand', '2004', 'Role of MMP-9 and its tissue inhibitor TIMP-1 in human osteosarcoma: findings in 42 patients followed for 1-16 years. The activity of matrix metalloproteinases (MMPs) in degrading extracellular matrix is controlled by activation of proenzymes and inhibition of MMP tissue inhibitors (TIMPs).To assess the proteolytic cascade imbalance in malignancy progression, tissue expression and serum levels of MMP-2, MMP-9 and of their inhibitors TIMP-2 and TIMP-1 respectively were evaluated in 42 selected patients with high-grade osteosarcoma (OS). MMP-2, MMP-9, TIMP-2 and TIMP-1 were studied in biopsies by immunohistochemistry and in serum by ELISA test. Patients were subdivided into 3 groups according to their follow up: continuously disease-free, diagnosis of metastasis during follow-up, and metastasis at diagnosis.Immunohistochemistry demonstrated an imbalance between MMPs and TIMPs, with a more evident role for MMP-9 than for MMP-2 in tumor progression. TIMP-1 inhibitor in plasma was higher in patients with osteosarcoma than in a control group. This high value of TIMP-1 was particularly evident in the group of patients who later developed metastases and/or local recurrences, and in those with metastases at diagnosis.Our findings confirm the protective action of TIMP-1, as MMP inhibitor, but also show its activity as a growth factor underlining its multifunctional role in OS.Copyright 2004 Taylor & Francis', 'http://www.ncbi.nlm.nih.gov/pubmed/15370596', 'yes', 'no', 'NA', 4),
(15390286, 'Inaba H, Glibetic M, Buck S, Ravindranath Y, Kaplan J', 'Interferon-gamma sensitizes osteosarcoma cells to Fas-induced apoptosis by up-regulating Fas receptors and caspase-8.', 'Pediatr Blood Cancer', '2004', 'Interferon-gamma sensitizes osteosarcoma cells to Fas-induced apoptosis by up-regulating Fas receptors and caspase-8. Osteosarcoma is the third most frequent neoplasm in adolescents. Although chemotherapy, frequently used in pre- and post-operative settings, has resulted in significant improvement in disease-free survival, some patients show little sensitivity to chemotherapy and alternative therapeutic strategies are needed. Because the Fas ligand/Fas receptor (CD95, APO-1) apoptosis pathway is a potential therapeutic target in osteosarcomas, we examined the effect of IFN-gamma on Fas-induced apoptosis in four osteosarcoma cell lines.As measured by flow cytometry, all cell lines expressed cell surface IFN-gamma receptors, and when cultured for 2 days in the presence of IFN-gamma, all cell lines exhibited a significant increase in expression of Fas receptors. By flow cytometric detection of intracellular fragmented DNA as a marker of apoptosis, all cell lines cultured with either IFN-gamma or anti-Fas antibody (clone CH-11) alone showed only moderate apoptosis, whereas significantly high levels of apoptosis occurred in cells cultured with both IFN-gamma and CH-11. Western blotting analysis also revealed that IFN-gamma caused up-regulation of caspase-8 in all cell lines, but no change in Fas-associated death domain protein (FADD/MORT1) or caspase-3. Both caspase-8 and caspase-3 were activated when apoptosis was induced with both IFN-gamma and CH-11. Addition to cultures of z-IETD-fmk, an inhibitor of caspase-8, significantly blocked this apoptosis.IFN-gamma sensitizes osteosarcoma cells to Fas-induced apoptosis through up-regulation of Fas receptor and caspase-8. Combined immunotherapy with IFN-gamma and either anti-Fas monoclonal antibody or cytotoxic T cells that bear Fas ligand might be a useful adjunctive therapy for patients with osteosarcoma.2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15390286', 'no', 'no', 'z-IETD-fmk', 4),
(15390294, 'Khayat CM, Johnston DL', 'Rhabdomyosarcoma, osteosarcoma, and adrenocortical carcinoma in a child with a germline p53 mutation.', 'Pediatr Blood Cancer', '2004', 'Rhabdomyosarcoma, osteosarcoma, and adrenocortical carcinoma in a child with a germline p53 mutation. A child with an unusual association of cancers is described. The patient first presented with a rhabdomyosarcoma of the right scapular muscle, and was successfully treated with chemotherapy. Six years after diagnosis of the first malignancy, the child presented with two synchronous malignancies: osteosarcoma of the jaw and adrenocortical carcinoma. Genetic mutation analysis was performed and revealed a germline p53 mutation of CGT > CAT at codon 273. The family history was negative for any other cancer consistent with the Li-Fraumeni syndrome. This case highlights the need for close surveillance of patients with p53 mutation for malignancy and describes the occurrence of two malignancies synchronously.(c) 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15390294', 'yes', 'no', 'NA', 1),
(15448009, 'Pakos EE, Kyzas PA, Ioannidis JP', 'Prognostic significance of TP53 tumor suppressor gene expression and mutations in human osteosarcoma: a meta-analysis.', 'Clin Cancer Res', '2004', 'Prognostic significance of TP53 tumor suppressor gene expression and mutations in human osteosarcoma: a meta-analysis. Various studies examining the relationship between tumor suppressor protein TP53 overexpression and/or TP53 gene mutations and the response to chemotherapy and clinical outcome in patients with osteosarcoma have yielded inconclusive results. The purpose of the current study was to evaluate the relation of TP53 status with response to chemotherapy and/or clinical outcome in osteosarcoma.We conducted a meta-analysis of 16 studies (n=499 patients) that evaluated the correlation between TP53 status and histologic response to chemotherapy and 2-year survival. Data were synthesized in summary receiver operating characteristic curves and with summary likelihood ratios (LRs) and risk ratios.The quantitative synthesis showed that TP53 status is not a prognostic factor for the response to chemotherapy. The positive LR was 1.21 (95% confidence interval, 0.86-1.71), and the negative LR was 0.91 (95% confidence interval, 0.77-1.07). There was no significant between-study heterogeneity. TP53-positive status tended to be associated with a worse 2-year survival, but the overall results were not formally statistically significant. The association was formally significant in studies that clearly stated that measurements were blinded to outcomes (risk ratio, 2.05; 95% confidence interval, 1.23-3.44), and in studies using reverse transcription-PCR for evaluating TP53 alterations (risk ratio, 1.76; 95% confidence interval, 1.07-2.91).TP53 status is not associated with the histologic response to chemotherapy in patients with osteosarcoma, whereas TP53 gene alterations may be associated with decreased survival.', 'http://www.ncbi.nlm.nih.gov/pubmed/15448009', 'yes', 'yes', 'NA', 1),
(15455381, 'Bagatell R, Beliakoff J, David CL, Marron MT, Whitesell L', 'Hsp90 inhibitors deplete key anti-apoptotic proteins in pediatric solid tumor cells and demonstrate synergistic anticancer activity with cisplatin.', 'Int J Cancer', '2005', 'Hsp90 inhibitors deplete key anti-apoptotic proteins in pediatric solid tumor cells and demonstrate synergistic anticancer activity with cisplatin. Drugs that inhibit the function of heat shock protein 90 (Hsp90) are of interest in the treatment of pediatric cancers because these agents deplete the cellular levels of signaling molecules that are important for the growth and survival of many childhood tumors. To generate preclinical data in anticipation of clinical trials of Hsp90 inhibitors in children, we evaluated the effects of the Hsp90 inhibitor geldanamycin (GA) alone and in combination with cis-platinum (II)-diamine dichloride (cisplatin) in pediatric tumor cells. Immunoblotting demonstrated depletion of the Hsp90 client proteins AKT and the type 1 insulin-like growth factor receptor (IGF1R) in a panel of pediatric tumor cell lines after exposure to GA. Drug exposure also led to a dramatic decrease in cell survival/proliferation in MYCN-amplified and non-amplified neuroblastoma cells. Moderate inhibition of survival/proliferation was observed in RB-deficient and wild-type osteosarcoma cells. Treatment of neuroblastoma and osteosarcoma cell lines with GA in combination with cisplatin resulted in greater than additive inhibition of survival/proliferation based on median dose analysis. Exposure to this drug combination also resulted in a marked increase in nuclear fragmentation as assessed by terminal deoxynucleotide transferase-mediated UTP nick end labeling (TUNEL) analysis. Combined exposure also abrogated the ability of GA to induce a cytoprotective heat shock response and resulted in Hsp90 adduct formation. Our findings suggest that Hsp90 inhibitors may prove useful either alone or as a component of multi-drug regimens in the treatment of neuroblastoma and osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15455381', 'no', 'no', 'geldanamycin|cisplatin', 4),
(15467717, 'Eppert K, Wunder JS, Aneliunas V, Kandel R, Andrulis IL', 'von Willebrand factor expression in osteosarcoma metastasis.', 'Mod Pathol', '2005', 'von Willebrand factor expression in osteosarcoma metastasis. A number of genes are implicated in the initiation and progression of osteosarcoma; however, cytogenetic and comparative genomic hybridization studies indicate the involvement of additional unidentified genes. An examination of gene expression profiles in 22 high-grade osteosarcoma tumor specimens from 15 patients (including paired primary and metastatic samples from five patients) indicated that von Willebrand factor (vWF) mRNA expression may increase during tumor progression. vWF, a large glycoprotein previously considered to be expressed exclusively by endothelial cells and megakaryocytes, is involved in platelet aggregation and adhesion to the subendothelial matrix, processes critical to hematogenous tumor cell metastasis to the lung. Analysis of paired primary and metastatic osteosarcoma tumor samples from 10 patients revealed an increase in vWF gene expression in metastases (P=0.005). Immunohistochemistry showed that, in addition to the endothelial cells, vWF protein was also detected in osteosarcoma cells in vivo in 13 of 29 tumor specimens as well as in SAOS2, an osteosarcoma cell line. The tumor cell staining correlated positively with high vWF expression in the sample (P=0.006). Although vascular endothelial cells contribute to the vWF mRNA detected in the tumor samples, there was neither any correlation between vascular density (VD) and vWF mRNA expression nor between VD and clinical outcome. These findings suggest that vWF expression is deregulated in osteosarcoma tumors, potentially contributing to metastasis.', 'http://www.ncbi.nlm.nih.gov/pubmed/15467717', 'yes', 'no', 'NA', 1),
(15488761, 'Chen W, Cooper TK, Zahnow CA, Overholtzer M, Zhao Z, Ladanyi M, Karp JE, Gokgoz N, Wunder JS, Andrulis IL, Levine AJ, Mankowski JL, Baylin SB', 'Epigenetic and genetic loss of Hic1 function accentuates the role of p53 in tumorigenesis.', 'Cancer Cell', '2004', 'Epigenetic and genetic loss of Hic1 function accentuates the role of p53 in tumorigenesis. The gene hypermethylated in cancer 1 (HIC1) is epigenetically inactivated, but not mutated, in cancer. Here we show that cooperative loss of Hic1 with p53, but not INK4a, yields distinct tumor phenotypes in mice. Germline deletion of one allele of each gene on the opposite chromosome yields breast and ovarian carcinomas and metastatic osteosarcomas with epigenetic inactivation of the wild-type Hic1 allele. Germline deletion of the two genes on the same chromosome results in earlier appearance and increased prevalence and aggressiveness of osteosarcomas with genetic deletion of both wild-type genes. In human osteosarcomas, hypermethylation of HIC1 is frequent only in tumors with p53 mutations. Our results indicate the importance of genes altered only through epigenetic mechanisms in cancer progression in conjunction with genetically modified tumor suppressor genes.', 'http://www.ncbi.nlm.nih.gov/pubmed/15488761', 'yes', 'no', 'NA', 2),
(15492501, 'Bihani T, Mason DX, Jackson TJ, Chen SC, Boettner B, Lin AW', 'Differential oncogenic Ras signaling and senescence in tumor cells.', 'Cell Cycle', '2004', 'Differential oncogenic Ras signaling and senescence in tumor cells. Several studies have shown that forced expression of oncogenic H-ras can induce a senescence-like permanent growth arrest in normal cells. Here we report that expression of oncogenic H-ras in human osteosarcoma U2OS cells also resulted in a senescence-like flat and enlarged cell morphology and permanent growth arrest. In contrast to normal human fibroblasts, U2OS cells were arrested independently of the p16 and ARF tumor suppressors. Treatment with a MEK inhibitor or a p38MAPK inhibitor interrupted oncogenic H-ras-induced growth arrest in U2OS cells, suggesting that activation of MAPK pathways is important. To further determine whether this process is unique to oncogenic H-ras signaling, we examined the effect of oncogenic K-ras on normal cells and human osteosarcoma cells. Similar to oncogenic H-ras, oncogenic K-ras also induced senescence in normal fibroblasts, while transforming immortalized mouse fibroblasts. However, in contrast to oncogenic H-ras, oncogenic K-ras failed to induce a permanent growth arrest in osteosarcoma U2OS cells. Additionally, cells transduced with oncogenic K-ras exhibited distinguishable cellular changes compared to those transduced with oncogenic H-ras. In summary, we report for the first time that oncogenic H-ras signaling can trigger a senescence-like growth arrest in tumor cells, independent of the p16 and ARF tumor suppressors. This result suggests that tumor cells may harbor a senescence-like program that can be activated by ras signaling. Moreover, our study uncovered a cell type-dependent differential response to oncogenic K-ras, as compared to oncogenic H-ras.', 'http://www.ncbi.nlm.nih.gov/pubmed/15492501', 'no', 'no', 'NA', 2),
(15516323, 'Nakayama Y, Kato N, Nakajima Y, Shimizu E, Ogata Y', 'Effect of TNF-alpha on human osteosarcoma cell line Saos2--TNF-alpha regulation of bone sialoprotein gene expression in Saos2 osteoblast-like cells.', 'Cell Biol Int', '2004', 'Effect of TNF-alpha on human osteosarcoma cell line Saos2--TNF-alpha regulation of bone sialoprotein gene expression in Saos2 osteoblast-like cells. Tumor necrosis factor-alpha (TNF-alpha) is a major mediator of inflammatory response in many diseases. It inhibits bone formation and stimulates bone resorption. To determine the molecular mechanisms involved in the regulation of gene expression of osteoblast-like cells, we analyzed the effects of TNF-alpha on the human osteosarcoma cell line Saos2. We used RT-PCR to examine the effects of TNF-alpha on bone sialoprotein (BSP), core binding factor a1 (Cbfa1), osterix, alpha 1 (I) collagen, cyclooxygenase-2 (COX-2), interleukin-6 (IL-6), cathepsin B, cathepsin L and tissue inhibitors of metalloproteinase-1 (TIMP-1). TNF-alpha (10ng/ml) increased BSP, IL-6 and COX-2 mRNA levels after 3h, reaching maximal levels at 12 h. Cbfa1 mRNA levels increased after 3 h, but decreased by 24 h. Osterix, cathepsin B, cathepsin L and TIMP-1 mRNA levels did not change after stimulation with TNF-alpha. On the other hand, alpha 1 (I) collagen mRNA expression was suppressed by TNF-alpha at 24 h. Transient transfection analyses were performed using chimeric constructs of the rat BSP gene promoter linked to a luciferase reporter gene. TNF-alpha (10 ng/ml) had no effect on the promoter activities of BSP transfected into Saos2 cells. The results of gel mobility shift assays using radiolabeled double-stranded cAMP response element (CRE) and FGF2 response element (FRE) oligonucleotides in the proximal promoter of the rat BSP gene showed increased binding of nuclear proteins at 6 h. Gel mobility shift assays with radiolabelled COX-2-CRE and COX-2-NF kappa B oligonucleotides revealed an increase in the binding of nuclear proteins from TNF-alpha-stimulated Saos2 cells. These studies, therefore, showed that TNF-alpha indirectly increased BSP expression, and that it could be mediated through COX-2 and Cbfa1 expression in Saos2 osteoblast-like cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/15516323', 'no', 'no', 'NA', 4),
(15538723, 'Hellwinkel OJ, Müller J, Pollmann A, Kabisch H', 'Osteosarcoma cell lines display variable individual reactions on wildtype p53 and Rb tumour-suppressor transgenes.', 'J Gene Med', '2005', 'Osteosarcoma cell lines display variable individual reactions on wildtype p53 and Rb tumour-suppressor transgenes. One of the most widely studied gene therapeutic strategies for cancer is the introduction of tumour-suppressor genes-generally p53-into the target cells. As the genes of p53 and/or retinoblastoma (Rb) are mutated in the major part of osteosarcomas (OS), we aimed to study the effect of p53 and Rb transgenes on a panel of five different osteosarcoma cell lines.OS cell lines were transduced by adenoviral vectors delivering the transcription units of the wildtype p53 and the Rb gene. Effects of the transgenes alone and at additional cytostatic stress were studied by proliferation, alive/dead and cell cycle assays.The individual cells lines displayed divergent reactions to p53- or Rb-transgene delivery reaching from cell death (SaOs-2, U2OS at p53 transduction) over stopped or lowered cell division (MG-63, K-HOS, SJSA-1 at p53 and Rb transduction) to nearly unhindered cell growth (U2OS at Rb transduction). In those OS cell lines reacting with lowered cell division to p53 or Rb delivery, cytostatics only moderately intensified the transgene effects. Surprisingly, these reactions were apparently not dependent on the functional status of the cellular p53 and/or Rb genes or on differences in the infectability of the cell lines by the adenoviral vectors. Most interestingly, the respective effects of the p53 or Rb transgenes were not multiplied by simultaneous transduction of both tumour-suppressor genes.The application of wildtype tumour-suppressor gene therapy on genetically variable osteosarcomas may be efficient only in yet not identified genetic subgroups of this tumour entity. Hyperactive tumour-suppressor transgenes could be an alternative.Copyright (c) 2004 John Wiley & Sons, Ltd.', 'http://www.ncbi.nlm.nih.gov/pubmed/15538723', 'no', 'no', 'NA', 2),
(15540114, 'Watanabe K, Okamoto K, Yonehara S', 'Sensitization of osteosarcoma cells to death receptor-mediated apoptosis by HDAC inhibitors through downregulation of cellular FLIP.', 'Cell Death Differ', '2005', 'Sensitization of osteosarcoma cells to death receptor-mediated apoptosis by HDAC inhibitors through downregulation of cellular FLIP. Fas-mediated apoptosis plays an important role in elimination of tumor cells in vivo, but some tumor-derived cells are resistant to this mechanism. Here, we show that treatment with the histone deacetylase (HDAC) inhibitor FR901228 renders Fas-resistant osteosarcoma cell lines sensitive to Fas-mediated apoptosis by downregulating expression of cellular FLIP (cellular FLICE-inhibitory protein), an inhibitor of Fas-mediated activation of caspase-8. Moreover, sensitization to Fas-mediated apoptosis was also induced in Fas-resistant osteosarcoma cells by suppressing FLIP expression using FLIP-specific RNA interference. HDAC inhibitors including FR901228 were shown to induce downregulation of cellular FLIP through inhibiting generation of FLIP mRNA, rather than stimulating degradation at either protein or mRNA level, and the inhibition was independent of de novo protein synthesis. These results clearly indicate that some tumor cells exhibit a phenotype resistant to death receptor-mediated apoptosis by expressing cellular FLIP, and that HDAC inhibitors sensitize such resistant tumor cells by directly downregulating cellular FLIP mRNA.', 'http://www.ncbi.nlm.nih.gov/pubmed/15540114', 'no', 'no', 'FR901228', 3),
(15540165, 'Atiye J, Wolf M, Kaur S, Monni O, Böhling T, Kivioja A, Tas E, Serra M, Tarkkanen M, Knuutila S', 'Gene amplifications in osteosarcoma-CGH microarray analysis.', 'Genes Chromosomes Cancer', '2005', 'Gene amplifications in osteosarcoma-CGH microarray analysis. Little is known about the genomic alterations underlying osteosarcoma. We performed a genomewide high-resolution gene copy number analysis of 22 osteosarcoma samples using comparative genomic hybridization on a cDNA microarray that contained cDNA clones of about 13,000 genes. Nineteen of the 22 cases had amplifications that on average spanned more than 1 Mb and contained more than 10 genes. Numerous regions of gain and loss were identified, and their boundaries were defined at high resolution. Novel amplicons were found at 14q11, 17q25, and 22q11-q13. Earlier-known large amplified regions were detected at 12q11-q15, 8q24, 6p12-p13, and 17p11-p13 in 8, 6, 5, and 4 of the 22 samples, respectively. Amplification of 12q was observed more frequently (36% of the cases) than previously reported. Previously known small amplicons at 1p34-p36, 1q21, 19q13, and 21q22 were seen in at least three cases. Our results implicate TOM1L2 and CYP27B1 as having roles as novel targets for the 17p and 12q amplicons, respectively. Details (www.helsinki.fi/cmg) of the amplified genes in each amplicon provide valuable raw data for further in silico studies.(c) 2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15540165', 'yes', 'no', 'NA', 2),
(15547696, 'Cenni V, Maraldi NM, Ruggeri A, Secchiero P, Del Coco R, De Pol A, Cocco L, Marmiroli S', 'Sensitization of multidrug resistant human ostesarcoma cells to Apo2 Ligand/TRAIL-induced apoptosis by inhibition of the Akt/PKB kinase.', 'Int J Oncol', '2004', 'Sensitization of multidrug resistant human ostesarcoma cells to Apo2 Ligand/TRAIL-induced apoptosis by inhibition of the Akt/PKB kinase. Chemotherapeutic agents have been used for the treatment of patients with osteosarcoma (OS). However, inherent or acquired resistance to these agents is a serious problem in the management of OS patients. The emergence of the multidrug resistance (MDR) phenotype in cancer cells is often associated with the overexpression of P-glycoprotein, encoded by the multidrug resistance gene MDR-1. The administration of some of the most common chemotherapeutic agents to these cells becomes ineffective because of their P-gp-driven efflux from the cell. Apo2L/TRAIL is a member of the tumor necrosis factor (TNF) family of cytokines that is considered to induce death of cancer cells but not normal cells. Its powerful apoptotic activity is mediated through its cell surface death domain-containing receptors, TRAIL-R1/DR4 and TRAIL-R2/DR5, which in turn spread the signal in the cytosol through the activation of the caspase cascade. The Akt/PKB kinase is an important cell survival protein which is regulated by D3-phosphoinositides. High Akt expression and activity levels are well documented in many types of tumors, which very often show an altered PI3-K/Akt/PTEN pathway. In this study the U2OS human osteosarcoma cell line and its multidrug resistant (MDR) subline that overexpresses MDR-1 gene, MDR-U2OS, have been analyzed for their responsiveness to TRAIL. In conflict with the presence of active DR4 and DR5 receptors in both clones, U2OS cells exhibited only a low responsiveness to TRAIL, while the MDR-U2OS subline did exhibit a marked TRAIL sensitivity. An analysis of the post-receptor events showed that TRAIL responsiveness correlates with a reduced expression of endogenous Akt. In fact, expression in MDR-U2OS cells of a constitutively active Akt strongly decreased their sensitivity to TRAIL. The identification of Akt as a key modulator of TRAIL responsiveness could help to design TRAIL-based combinations for treatment of osteosarcoma. Moreover, the discovery that multidrug resistant osteosarcomas are highly sensitive to TRAIL-induced apoptosis indicates TRAIL as a new candidate for the treatment of multidrug resistant bone malignancies.', 'http://www.ncbi.nlm.nih.gov/pubmed/15547696', 'no', 'no', 'NA', 6),
(15583032, 'Thomas DM, Johnson SA, Sims NA, Trivett MK, Slavin JL, Rubin BP, Waring P, McArthur GA, Walkley CR, Holloway AJ, Diyagama D, Grim JE, Clurman BE, Bowtell DD, Lee JS, Gutierrez GM, Piscopo DM, Carty SA, Hinds PW', 'Terminal osteoblast differentiation, mediated by runx2 and p27KIP1, is disrupted in osteosarcoma.', 'J Cell Biol', '2004', 'Terminal osteoblast differentiation, mediated by runx2 and p27KIP1, is disrupted in osteosarcoma. The molecular basis for the inverse relationship between differentiation and tumorigenesis is unknown. The function of runx2, a master regulator of osteoblast differentiation belonging to the runt family of tumor suppressor genes, is consistently disrupted in osteosarcoma cell lines. Ectopic expression of runx2 induces p27KIP1, thereby inhibiting the activity of S-phase cyclin complexes and leading to the dephosphorylation of the retinoblastoma tumor suppressor protein (pRb) and a G1 cell cycle arrest. Runx2 physically interacts with the hypophosphorylated form of pRb, a known coactivator of runx2, thereby completing a feed-forward loop in which progressive cell cycle exit promotes increased expression of the osteoblast phenotype. Loss of p27KIP1 perturbs transient and terminal cell cycle exit in osteoblasts. Consistent with the incompatibility of malignant transformation and permanent cell cycle exit, loss of p27KIP1 expression correlates with dedifferentiation in high-grade human osteosarcomas. Physiologic coupling of osteoblast differentiation to cell cycle withdrawal is mediated through runx2 and p27KIP1, and these processes are disrupted in osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15583032', 'no', 'no', 'NA', 3);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(15585647, 'Lafleur EA, Koshkina NV, Stewart J, Jia SF, Worth LL, Duan X, Kleinerman ES', 'Increased Fas expression reduces the metastatic potential of human osteosarcoma cells.', 'Clin Cancer Res', '2004', 'Increased Fas expression reduces the metastatic potential of human osteosarcoma cells. The process of metastasis requires the single tumor cell that seeds the metastatic clone to complete a complex series of steps. Identifying factors responsible for these steps is essential in developing and improving targeted therapy for metastasis. Resistance to receptor-mediated cell death, such as the Fas/Fas ligand pathway, is one mechanism commonly exploited by metastatic cell populations.LM7, a subline of the SAOS human osteosarcoma cell line with low Fas expression, was selected for its high metastatic potential in an experimental nude mouse model. When transfected with the full-length Fas gene (LM7-Fas), these cells expressed higher levels of Fas than the parental LM7 cells or LM7-neo control-transfected cells. These cells were also more sensitive to Fas-induced cell death than controls. When injected intravenously into nude mice, the LM7-Fas cell line produced a significantly lower incidence of tumor nodules than control cell lines. Lung weight and tumor nodule size were also decreased in those mice injected with LM7-Fas. Levels of Fas were quantified in osteosarcoma lung nodules from 17 patients. Eight samples were Fas negative, whereas the remaining 9 were only weakly positive compared with normal human liver (positive control).Our results demonstrate that altering Fas expression can impact the metastatic potential of osteosarcoma cells. We conclude that the increase of Fas on the surface of the LM7 osteosarcoma cells increased their sensitivity to Fas-induced cell death in the microenvironment of the lung, where Fas ligand is constitutively expressed. Thus, loss of Fas expression is one mechanism by which osteosarcoma cells may evade host resistance mechanisms in the lung, increasing metastatic potential. Fas may therefore be a new therapeutic target for osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15585647', 'yes', 'no', 'NA', 1),
(15609301, 'Eppert K, Wunder JS, Aneliunas V, Tsui LC, Scherer SW, Andrulis IL', 'Altered expression and deletion of RMO1 in osteosarcoma.', 'Int J Cancer', '2005', 'Altered expression and deletion of RMO1 in osteosarcoma. In order to increase our understanding of the molecular events underlying osteosarcoma progression, the expression of approximately 950 genes was examined in 24 primary and metastatic osteosarcoma tumor specimens. A gene, RMO1, was isolated with decreased expression in metastatic samples. Real-Time PCR corroborated this pattern, revealing lower expression in the primary sample in 6 of 7 cases for which both primary and metastatic osteosarcoma samples were available from the same patient (p = 0.034). RMO1 is located at 2q33, a region of frequent loss of heterozygosity in cancer, and exhibited loss of heterozygosity in 6 out of 9 primary osteosarcoma tumor samples (67%). Loss of heterozygosity is evident in primary tumors while the decrease in gene expression is seen in the metastatic samples, indicating that these 2 events are separately implicated in cancer progression. Cloning of RMO1 revealed an open reading frame with multiple splice forms with significant homology to GRB7, 10 and 14 and MIG10 in the region containing a Pleckstrin homology domain and a Ras association domain, suggestive of a role in cell signaling and migration. Northern blot analysis indicated that RMO1 mRNA is ubiquitously expressed in tissues except for peripheral blood leukocytes. These data suggest that RMO1 may be a candidate for a protein involved in inhibiting tumor progression.2004 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15609301', 'yes', 'yes', 'NA', 1),
(15625311, 'Morimoto H, Okamura H, Yoshida K, Kitamura S, Haneji T', 'Okadaic acid induces apoptosis through double-stranded RNA-dependent protein kinase/eukaryotic initiation factor-2alpha pathway in human osteoblastic MG63 cells.', 'J Biochem', '2004', 'Okadaic acid induces apoptosis through double-stranded RNA-dependent protein kinase/eukaryotic initiation factor-2alpha pathway in human osteoblastic MG63 cells. Double-stranded RNA-dependent protein kinase (PKR) is a participant in the cellular antiviral response and phosphorylates the alpha-subunit of eukaryotic translation initiation factor 2alpha (eIF-2alpha) to block protein synthesis. Treatment of human osteosarcoma cell line MG63 cells with a serine and threonine protein phosphatase inhibitor, okadaic acid, at the concentration of 100 nM, but not at 20 nM, induced apoptosis. To investigate the functional relationship between phosphatases and apoptosis, we examined the phosphorylation levels of PKR and eIF-2alpha by Western blot analysis. During treatment of cells with it at the higher concentration (100 nM), okadaic acid increased the level of phosphorylated PKR in MG63 cells, this kinase phosphorylating eIF-2alpha. However, at the lower concentration (20 nM), okadaic acid did not affect the level of phosphorylated PKR. In the cells treated with 100 nM okadaic acid, activation of NF-kappaB also occurred. Even though inhibition of translation occurred simultaneously in MG63 cells, the expression of pro-apoptotic proteins Fas and Bax was not affected by 100 nM okadaic acid in these cells. We concluded that the inhibition of translation decreased anti-apoptotic protein expression, thus resulting in apoptosis. Our results also suggest that the inhibition of the protein phosphatase activity by okadaic acid induced apoptosis in MG63 cells through PKR and eIF-2alpha.', 'http://www.ncbi.nlm.nih.gov/pubmed/15625311', 'no', 'no', 'okadaic acid', 3),
(15627479, 'Ahlström M, Pekkinen M, Huttunen M, Lamberg-Allardt C', 'Dexamethasone down-regulates cAMP-phosphodiesterase in human osteosarcoma cells.', 'Biochem Pharmacol', '2005', 'Dexamethasone down-regulates cAMP-phosphodiesterase in human osteosarcoma cells. Cyclic adenosine monophosphate (cAMP) is an important second messenger in the hormonal regulation of bone metabolism. cAMP is inactivated by the cyclic nucleotide phosphodiesterases (PDEs), a superfamily of enzymes divided into 11 known families, designated PDE1-11. Interference with the cAMP signaling pathway has been suggested as one mechanism causing glucocorticoid induced osteoporosis. We speculated that glucocorticoids could affect the cAMP pathway by a down-regulation of PDE-mediated cAMP hydrolysis. The main cAMP hydrolysing enzyme families of human MG-63 and SaOS-2 osteosarcoma cells were identified as PDE1 and PDE4 by assaying the PDE activity of Q-sepharose fractions and cell homogenates with selective inhibitors. Treatment with the glucocorticoid dexamethasone (Dex) decreased cAMP-PDE activity by up to 50%, without affecting cGMP-PDE activity. Dex treatment reduced the sensitivity of the total cAMP-PDE activity towards the PDE4 selective PDE inhibitor rolipram. Forskolin stimulated cAMP accumulation was increased 30-60-fold in the presence of rolipram. Treatment with Dex did not affect the basal or forskolin stimulated cAMP accumulation, but treatment resulted in a reduced effect of rolipram on cAMP accumulation. Expression of the following cAMP-PDE subtypes were detected by reverse transcriptase PCR (RT-PCR): PDE1A, PDE1C, PDE2A, PDE3A, PDE4A, PDE4B, PDE4C, PDE4D, PDE7A, PDE7B, PDE8A, PDE10A and PDE11A. Using semi-quantitative RT-PCR, we detected a 50-70% decrease in the mRNA of PDE4A and PDE4B subtypes following Dex treatment. Further analysis revealed that Dex reduced the PDE4A4 and PDE4B1 isoforms. PDE4A1 PDE4A, PDE4A7, PDE4A10, PDE4B2 were also expressed, but Dex did not affect the transcription of these isoforms. We conclude that Dex treatment could affect the cAMP signaling pathway of human osteosarcoma cells by reducing type 4 cAMP-phosphodiesterase (PDE4).', 'http://www.ncbi.nlm.nih.gov/pubmed/15627479', 'no', 'no', 'dexamethasone', 13),
(15649646, 'Trougakos IP, Lourda M, Agiostratidou G, Kletsas D, Gonos ES', 'Differential effects of clusterin/apolipoprotein J on cellular growth and survival.', 'Free Radic Biol Med', '2005', 'Differential effects of clusterin/apolipoprotein J on cellular growth and survival. The secreted clusterin/apolipoprotein J (CLU) protein form is a ubiquitously expressed heterodimeric glycoprotein which is differentially regulated in many severe physiological disturbance states including cell death, ageing, cancer progression, and various neurological diseases. Despite extensive efforts CLU function remains an enigma, the main cause being the intriguingly distinct and usually opposed functions in various cell types and tissues. In the current report we investigated the effects of CLU on cellular growth and survival in three human osteosarcoma (OS) cell lines, namely KH OS, Sa OS, and U-2 OS that express very low, moderate, and high endogenous steady-state CLU amounts, respectively. We found that exposure of these established OS cell lines or primary OS cells to genotoxic stress results in CLU gene induction at distinct levels that correlate negatively to CLU endogenous amounts. Following CLU-forced overexpression by means of an artificial transgene, we found that although extracellular CLU inhibits cell death in all three OS cell lines, intracellular CLU has different effects on cellular proliferation and survival in these cell lines. Transgenic KH OS cell lines adapted to moderate intracellular CLU levels were growth-retarded and became resistant to genotoxic and oxidative stress. In contrast, transgenic Sa OS and U2 OS cell lines adapted to high intracellular CLU amounts were sensitive to genotoxic and oxidative stress. In these two cell lines, the proapoptotic CLU function could be rescued by caspase inhibition. To monitor the immediate effects of heterologous CLU overexpression prior to cell adaptation, we performed transient transfections in all three OS cell lines. We found that induction of high intracellular CLU amounts increases spontaneous apoptosis in KH OS cells and reduces DNA synthesis in all three cell lines assayed. On the basis of these novel findings we propose that although extracellular CLU as well as intracellular CLU at low/moderate levels is cytoprotective, CLU may become highly cytostatic and/or cytotoxic if it accumulates intracellularly in high amounts either by direct synthesis or by uptake from the extracellular milieu.', 'http://www.ncbi.nlm.nih.gov/pubmed/15649646', 'no', 'no', 'NA', 1),
(15656333, 'Bacci G, Longhi A, Ferrari S, Briccoli A, Donati D, De Paolis M, Versari M', 'Prognostic significance of serum lactate dehydrogenase in osteosarcoma of the extremity: experience at Rizzoli on 1421 patients treated over the last 30 years.', 'Tumori', 'NA', 'Prognostic significance of serum lactate dehydrogenase in osteosarcoma of the extremity: experience at Rizzoli on 1421 patients treated over the last 30 years. The study evaluated the correlation between pretreatment serum lactate dehydrogenase (LDH) levels with the stage of disease and its clinical prognostic value.Pretreatment serum LDH of 1421 patients with osteosarcoma of the extremity were assessed to investigate whether the enzyme correlates with the stage of the tumor. In 860 assessable patients with localized disease, treated according to 10 different protocols of adjuvant (four) and neoadjuvant chemotherapy (six), we also evaluated the correlation between the serum levels of LDH and outcome.According to the stage of disease, the rate of high serum level of LDH was significantly higher in 199 patients with metastatic disease at presentation than in 1222 patients with localized disease (36.6% vs 18.8%; P < 0.0001). In these patients, the 5-year disease-free survival was 39.5% for patients with high LDH levels and 60% for those with normal values. The 5-year disease-free survival correlated with serum level of LDH at univariate and multivariate analysis, although it lost its significance when histologic response to chemotherapy was also considered in the multivarite analysis.Serum LDH has a prognostic value and it should be considered in evaluating the results of therapeutic trials of chemotherapy, as well as defining a category of patients at high-risk of relapse to be treated with a more aggressive regimen.', 'http://www.ncbi.nlm.nih.gov/pubmed/15656333', 'yes', 'yes', 'NA', 1),
(15662138, 'Zhu JJ, Li FB, Zhou JM, Liu ZC, Zhu XF, Liao WM', 'The tumor suppressor p33ING1b enhances taxol-induced apoptosis by p53-dependent pathway in human osteosarcoma U2OS cells.', 'Cancer Biol Ther', '2005', 'The tumor suppressor p33ING1b enhances taxol-induced apoptosis by p53-dependent pathway in human osteosarcoma U2OS cells. p33ING1b can stimulate cell cycle arrest, DNA repair, apoptosis and chemosensitivity. The actions of p33ING1b involve p53-dependent and p53-independent mechanisms. To investigate if the p33ING1b isoform is involved in the chemosensitivity of osteosarcoma cells, p33ING1b was overexpressed in p53+/+ U2OS cells or p53-mutant MG63 cells, and then cell growth arrest and apoptosis were assessed after treatment with taxol. The results showed that p33ING1b markedly increased taxol-induced growth inhibition and apoptosis in p53+/+ U2OS cells, but not in p53-mutant MG63 cells. Moreover, ectopic expression of p33ING1b could obviously upregulate p53, p21WAF1 and bax protein levels and activate caspase-3 in taxol-treated U2OS cells. Taken together, our data demonstrate that p33ING1b enhances taxol-induced apoptosis through p53-dependent pathway in human osteosarcoma cells. p33ING1b may be an important marker and/or therapeutic target in the prevention and treatment of osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15662138', 'no', 'no', 'taxol', 5),
(15670817, 'De Blasio A, Messina C, Santulli A, Mangano V, Di Leonardo E, D''Anneo A, Tesoriere G, Vento R', 'Differentiative pathway activated by 3-aminobenzamide, an inhibitor of PARP, in human osteosarcoma MG-63 cells.', 'FEBS Lett', '2005', 'Differentiative pathway activated by 3-aminobenzamide, an inhibitor of PARP, in human osteosarcoma MG-63 cells. This study describes the molecular mechanism by which treatment with 3-AB, a potent inhibitor of PARP, allows human osteosarcoma MG-63 cells to restrict growth and enter differentiation. Our findings show that in MG-63 cells, aberrant gene expression keeps Rb protein constitutively inactivated through hyperphosphorylation and this promotes uncontrolled proliferation of the cells. After 3-AB-treatment, the poly(ADP-ribosyl)ation of nuclear proteins markedly decreases and this results in an increase in both the hypophosphorylated active form of Rb and pRb/E2F complexes. These effects are accompanied by G1 arrest, downregulation of gene products required for proliferation (cyclin D1, beta-catenin, c-Jun, c-Myc and Id2) and upregulation of those implicated in the osteoblastic differentiation (p21/Waf1, osteopontin, osteocalcin, type I collagen, N-cadherins and alkaline phosphatase). Our study suggests that use of PARP inhibitors may induce a remodeling of chromatin with the reprogramming of gene expression and the activation of differentiation.', 'http://www.ncbi.nlm.nih.gov/pubmed/15670817', 'no', 'no', '3-aminobenzamide', 12),
(15701832, 'Perissinotto E, Cavalloni G, Leone F, Fonsato V, Mitola S, Grignani G, Surrenti N, Sangiolo D, Bussolino F, Piacibello W, Aglietta M', 'Involvement of chemokine receptor 4/stromal cell-derived factor 1 system during osteosarcoma tumor progression.', 'Clin Cancer Res', '2005', 'Involvement of chemokine receptor 4/stromal cell-derived factor 1 system during osteosarcoma tumor progression. Despite intensive chemotherapy and surgery treatment, lung and bone metastasis develop in about 30% of patients with osteosarcoma. Mechanisms for this preferential metastatic behavior are largely unknown. We investigated the role of the chemokine receptor 4 (CXCR4)/stromal cell-derived factor 1 (SDF-1) system to drive the homing of osteosarcoma cells. We analyzed the expression of the CXCR4 and SDF-1 proteins on several osteosarcoma cell lines and the effects of SDF-1 on migration, adhesion, and proliferation of these cancer cells. In vitro assays showed that the migration of osteosarcoma cells expressing CXCR4 receptor follows an SDF-1 gradient and that their adhesion to endothelial and bone marrow stromal cells is promoted by SDF-1 treatment. Moreover, the production of matrix metalloproteinase-9 is increased after SDF-1 exposure. We finally proved in a mouse model our hypothesis of the CXCR4/SDF-1 axis involvement in the metastatic process of osteosarcoma cells. Development of lung metastasis after injection of osteosarcoma cells was prevented by the administration of a CXCR4 inhibitor, the T134 peptide. These data show a possible explanation for the preferential osteosarcoma metastatic development into the lung, where SDF-1 concentration is high, and suggest that molecular strategies aimed at inhibiting the CXCR4/SDF-1 pathway, such as small-molecule inhibitors or anti-CXCR4 antibodies, might prevent the dissemination of osteosarcoma cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/15701832', 'no', 'no', 'T134 peptide', 3),
(15705874, 'Maude SL, Enders GH', 'Cdk inhibition in human cells compromises chk1 function and activates a DNA damage response.', 'Cancer Res', '2005', 'Cdk inhibition in human cells compromises chk1 function and activates a DNA damage response. Cyclin-dependent kinases (Cdk) promote cell proliferation, are often deregulated in human cancers, and are targets of ongoing cancer chemotherapy trials. We show here that Cdk activity is also required in human cells to maintain function of the Chk1 pathway, a key component of the response to DNA damage or stalled replication. Chk1 expression was markedly reduced in primary fibroblasts and U2OS osteogenic sarcoma cells by treatment with small molecule Cdk inhibitors or induction of a dominant-negative mutant of Cdk2. The findings of decreased Chk1 activity and accumulation of Cdc25A, a protein targeted for degradation by Chk1, confirmed that Chk1 function was impaired. Furthermore, Cdk inhibition triggered a DNA damage response, characterized by the accumulation of activated forms of ATM and Chk2 as well as nuclear foci containing phosphorylated substrates of ATM/ATR, including histone H2AX (gammaH2AX). Time course experiments showed that the bulk of ATM activation followed Chk1 down-regulation. Chk1 RNA interference combined with partial inhibition of DNA replication was sufficient to evoke the DNA damage response. Conversely, ectopic expression of Chk1 blunted induction of gammaH2AX foci by Cdk inhibitors, indicating that Chk1 down-regulation was necessary to elicit the full phenotype. Finally, both Cdk and Chk1 inhibitors enhanced the cytotoxity of etoposide, a DNA-damaging agent. These results define a pathway through which Cdk inhibition can mediate DNA damage and potentially enhance the efficacy of extant cancer chemotherapies.', 'http://www.ncbi.nlm.nih.gov/pubmed/15705874', 'no', 'no', 'NA', 5),
(15735124, 'Wunder JS, Gokgoz N, Parkes R, Bull SB, Eskandarian S, Davis AM, Beauchamp CP, Conrad EU, Grimer RJ, Healey JH, Malkin D, Mangham DC, Rock MJ, Bell RS, Andrulis IL', 'TP53 mutations and outcome in osteosarcoma: a prospective, multicenter study.', 'J Clin Oncol', '2005', 'TP53 mutations and outcome in osteosarcoma: a prospective, multicenter study. Mutations of the TP53 gene have been associated with resistance to chemotherapy as well as poor prognosis in many different malignancies. This is the first prospective study of the prognostic value of somatic TP53 mutations in patients with newly diagnosed extremity osteosarcoma.One hundred ninety-six patients with high-grade, nonmetastatic osteosarcoma of the extremities were enrolled from seven tertiary care institutions and observed prospectively for tumor recurrence (median follow-up duration, 44 months). All patients received neoadjuvant or adjuvant chemotherapy and surgery. Tumors were analyzed for the presence of TP53 mutations by polymerase chain reaction single-strand conformation polymorphism analysis and direct DNA sequencing. The association of the status of the TP53 gene with the risk of systemic recurrence was examined using survival analyses with traditional and histologic markers as prognostic factors.Patient age was the only factor that varied with TP53 gene status (P = .05). No relationship was identified between TP53 status and systemic relapse (relative risk, 1.24; P = .41). Analyses based on missense or nonsense mutations gave similar results (P > .10). In multivariate analysis, large (> 9 cm) tumor size (relative risk, 1.9; P = .006) and poor histologic response (< or = 90% necrosis) to chemotherapy (relative risk, 2.14; P = .02) were the only significant independent predictors of systemic outcome.We found no evidence that TP53 mutations predict for development of metastases in patients with high-grade osteosarcoma. Identification of other genes that influence chemotherapy response and clinical outcome in osteosarcoma is needed to facilitate further improvements in patient outcomes.', 'http://www.ncbi.nlm.nih.gov/pubmed/15735124', 'yes', 'no', 'NA', 1),
(15735172, 'Andela VB, Siddiqui F, Groman A, Rosier RN', 'An immunohistochemical analysis to evaluate an inverse correlation between Runx2/Cbfa1 and NF kappa B in human osteosarcoma.', 'J Clin Pathol', '2005', 'An immunohistochemical analysis to evaluate an inverse correlation between Runx2/Cbfa1 and NF kappa B in human osteosarcoma. Dominant negative inhibition of nuclear factor kappa B (NF kappa B) signalling activity in a human osteosarcoma cell line (Saos2) results in malignant reversion and the induction of the osteoblast differentiating transcription factor, Runx2/Cbfa1. This observation suggests that there is an inverse relation between a transcription factor associated with malignant progression and chemoresistance (NF kappa B) and an osteoblast differentiating transcription factor (Runx2/Cbfa1).To assess and correlate Runx2/Cbfa1 and NF kappa B (p65) immunoreactivity in human osteosarcoma.Runx2/Cbfa1 and NFkappaB (p65) immunoreactivity was assessed on 11 paraffin wax embedded archival specimens of human primary osteosarcoma by standard immunohistochemical methods and scored on a scale of 0-3. A Pearson correlation analysis between Runx2/Cbfa1 and NF kappa B (p65) scores was established.Runx2/Cbfa1 was expressed constitutively in all pathology specimens of human osteosarcoma. Of note, a chondroblastic osteosarcoma showed the highest Runx2/Cbfa1 immunoreactivity. A Pearson correlation did not support an inverse correlation between Runx2/Cbfa1 and NF kappa B (p65) scores (r = 0.57) in human osteosarcoma.Runx2/Cbfa1 immunoreactivity does not inversely correlate with NF kappa B immunoreactivity, and thus cannot serve as an indirect measure of NF kappa B activity or an independent predictive or prognostic indicator.', 'http://www.ncbi.nlm.nih.gov/pubmed/15735172', 'yes', 'yes', 'NA', 2),
(15742288, 'Kanno T, Takahashi T, Tsujisawa T, Ariyoshi W, Nishihara T', 'Platelet-rich plasma enhances human osteoblast-like cell proliferation and differentiation.', 'J Oral Maxillofac Surg', '2005', 'Platelet-rich plasma enhances human osteoblast-like cell proliferation and differentiation. Platelet-rich plasma (PRP) is widely used to promote tissue healing. However, there is no concrete evidence for the biological effects of PRP. This study evaluated the biological effects of PRP on the proliferation and differentiation of 2 human osteoblast-like cell lines.Human osteosarcoma cell lines HOS and SaOS-2 were used in this study. PRP was prepared from freshly drawn human venous blood containing a large number of platelets. The MTT assay was used to examine the effects of PRP on osteoblast viability. To evaluate the growth and differentiation, alkaline phosphatase activity was assessed and the expression of procollagen type I, osteopontin, and osteoprotegerin mRNA was measured using semiquantitative reverse transcriptase-polymerase chain reaction. Further, core binding factor alpha 1 (cbfa1/Runx2/AML3/PebpalphaA), a critical regulator of osteoblast differentiation, was also determined.The administration of PRP enhanced the viability of HOS and SaOS-2 cells in a dose-dependent manner. Alkaline phosphatase activity was suppressed during the cell growth phase, but was strongly enhanced when the cells reached confluence. Semiquantitative reverse-transcription polymerase chain reaction analysis showed that PRP enhanced the levels of procollagen type I, osteopontin, osteoprotegerin, and core binding factor alpha 1 (cbfa1) mRNA.These results suggest that PRP has a favorable effect on human osteoblast-like cells, and acts both to enhance bone regeneration and as an activator in wound healing.', 'http://www.ncbi.nlm.nih.gov/pubmed/15742288', 'no', 'no', 'NA', 6),
(15746965, 'Miura K, Uniyal S, Leabu M, Oravecz T, Chakrabarti S, Morris VL, Chan BM', 'Chemokine receptor CXCR4-beta1 integrin axis mediates tumorigenesis of osteosarcoma HOS cells.', 'Biochem Cell Biol', '2005', 'Chemokine receptor CXCR4-beta1 integrin axis mediates tumorigenesis of osteosarcoma HOS cells. It is known that beta1 integrins mediate the migratory response of cells to chemokine stimulation. Also, both beta1 integrins and chemokines have roles in tumor development. In the present study, the beta1 integrin-chemokine axis is assessed using human osteosarcoma (HOS) transfectant cells expressing the CXCR4 receptor for chemokine SDF-1 (CXCL12). We first identified in vitro the specific beta1 integrins that mediated the migratory response to SDF-1 stimulation. Results showed that on collagen type I and laminin, the chemotactic response to SDF-1 was predominantly mediated by alpha2beta1 integrin. On fibronectin, SDF-1-stimulated chemotaxis involved both alpha4beta1 and alpha5beta1 integrins. A comparison of the transfectant clones expressing CXCR4 at low, intermediate, and high levels and the control transfectant revealed that the transfectant clones migratory response in vitro and their ability to form tumors in vivo was related to their levels of CXCR4 expression. In addition, treatment by injection with mAbs to CXCR4, integrin alpha2beta1, or integrin alpha5beta1 effectively inhibited the growth of HOS-CXCR4 transfectant cells in vivo. Therefore, our results show that the beta1 integrins that mediated the migratory response were also functionally linked to the enhanced tumor growth of CXCR4-expressing HOS transfectant cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/15746965', 'no', 'no', 'NA', 6),
(15753370, 'Mintz MB, Sowers R, Brown KM, Hilmer SC, Mazza B, Huvos AG, Meyers PA, Lafleur B, McDonough WS, Henry MM, Ramsey KE, Antonescu CR, Chen W, Healey JH, Daluski A, Berens ME, Macdonald TJ, Gorlick R, Stephan DA', 'An expression signature classifies chemotherapy-resistant pediatric osteosarcoma.', 'Cancer Res', '2005', 'An expression signature classifies chemotherapy-resistant pediatric osteosarcoma. Osteosarcoma is the most common malignant bone tumor in children. Osteosarcoma patients who respond poorly to chemotherapy are at a higher risk of relapse and adverse outcome. Therefore, it was the aim of this study to identify prognostic factors at the time of diagnosis to characterize the genes predictive of poor survival outcome and to identify potential novel therapeutic targets. Expression profiling of 30 osteosarcoma diagnostic biopsy samples, 15 with inferior necrosis following induction chemotherapy (Huvos I/II) and 15 with superior necrosis following induction chemotherapy (Huvos III/IV), was conducted using Affymetrix U95Av2 oligonucleotide microarrays. One hundred and four genes were found to be statistically significant and highly differentially expressed between Huvos I/II and III/IV patients. Statistically significant genes were validated on a small independent cohort comprised of osteosarcoma xenograft tumor samples. Markers of Huvos I/II response predominantly were gene products involved in extracellular matrix (ECM) microenvironment remodeling and osteoclast differentiation. A striking finding was the significant decrease in osteoprotegerin, an osteoclastogenesis inhibitory factor. Additional genes involved in osteoclastogenesis and bone resorption, which were statistically different, include annexin 2, SMAD, PLA2G2A, and TGFbeta1. ECM remodeling genes include desmoplakin, SPARCL1, biglycan, and PECAM. Gene expression of select genes involved in tumor progression, ECM remodeling, and osteoclastogenesis were validated via quantitative reverse transcription-PCR in an independent cohort. We propose that osteosarcoma tumor-driven changes in the bone microenvironment contribute to the chemotherapy-resistant phenotype and offer testable hypotheses to potentially enhance therapeutic response.', 'http://www.ncbi.nlm.nih.gov/pubmed/15753370', 'yes', 'no', 'NA', 8),
(15796961, 'Mendoza S, David H, Gaylord GM, Miller CW', 'Allelic loss at 10q26 in osteosarcoma in the region of the BUB3 and FGFR2 genes.', 'Cancer Genet Cytogenet', '2005', 'Allelic loss at 10q26 in osteosarcoma in the region of the BUB3 and FGFR2 genes. Loss of heterozygosity at 10q26 was mapped using microsatellite markers in 20 osteosarcomas. A four-megabase region centered on marker D10S587 was affected by allelic loss in 60 percent of osteosarcomas. The most frequently lost marker was D10S1723. Around 15 known genes are found in this region. The gene immediately adjacent to D10S1723 encodes BUB3, an element of the spindle assembly mitotic checkpoint. Loss of BUB3 function could contribute to chromosomal instability. The fibroblast growth factor receptor 2 (FGFR2) gene is located 2 Mb from the BUB3 gene and has the potential for a role in cancer. Inherited mutations of the FGFR2 gene result in skeletal dysplasias. FGFR2 alterations have also been implicated in gastric cancer. Human genome project data were used to design primers for amplifying FGFR2 in 18 genomic segments and BUB3 in 7 genomic segments. In each case, the segments encompassed coding exons and flanking intron sequences. The primers were used to search for mutations by polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP). Several shifted bands were detected in the BUB3 exon 3 fragment. Sequencing resolved the BUB3 exon 3 fragment shifts into polymorphisms in intron 2. No mutations of BUB3 or FGFR2 were detected. It remains possible that BUB3 or FGFR2 hemizygosity alone contributes to osteosarcoma, or that one of the genes is cryptically inactivated by a higher-order modification or mutation outside the coding region. There may also be a yet undiscovered tumor suppressor gene in this region.', 'http://www.ncbi.nlm.nih.gov/pubmed/15796961', 'yes', 'no', 'NA', 2),
(15800315, 'Entz-Werlé N, Marcellin L, Gaub MP, Guerin E, Schneider A, Berard-Marec P, Kalifa C, Brugiere L, Pacquement H, Schmitt C, Tabone MD, Jeanne-Pasquier C, Terrier P, Dijoud F, Oudet P, Lutz P, Babin-Boilletot A', 'Prognostic significance of allelic imbalance at the c-kit gene locus and c-kit overexpression by immunohistochemistry in pediatric osteosarcomas.', 'J Clin Oncol', '2005', 'Prognostic significance of allelic imbalance at the c-kit gene locus and c-kit overexpression by immunohistochemistry in pediatric osteosarcomas. Since the recent development of biologic agents targeting oncogenes, increasing attention has been focused on determining the role of tyrosine kinase receptors in the pathogenesis of tumors. Our study was designed to investigate the status of region 4q12, which contains the candidate gene c-kit, and the expression of c-kit by immunohistochemistry (IHC).Paired blood and biopsy specimens of 68 children treated for high-grade primary osteosarcomas were collected. Microsatellite analysis at two genomic sites containing c-kit gene was performed on paired DNA using a sensible fluorescent polymerase chain reaction technology. To confirm the DNA data, we studied c-kit protein expression by IHC in 56 available paraffin-embedded tumor tissues.The frequency of allelic imbalance (AI) at locus 4q12 was 39% in the overall population. In agreement with previous studies, we did not detect microsatellite instability, allowing us to hypothesize that this pathway is not implicated. Furthermore, the normal status at locus 4q12 was associated with a significantly better survival in the whole osteosarcoma population (P = .05). IHC overexpression of c-kit was concordant in all cases presenting an AI. However, normal status at locus 4q12 was correlated to an absence of c-kit protein expression in 19 (65.5%) of 29 informative cases.Allelotyping of locus 4q12, which contains the c-kit gene, could help pediatric osteosarcoma prognostic screening and showed a strong correlation with overexpression of c-kit protein. These results allowed us to hypothesize that, in some cases, a mutation of c-kit gene could lead to a protein overexpression.', 'http://www.ncbi.nlm.nih.gov/pubmed/15800315', 'yes', 'yes', 'NA', 1),
(15814634, 'Laverdiere C, Hoang BH, Yang R, Sowers R, Qin J, Meyers PA, Huvos AG, Healey JH, Gorlick R', 'Messenger RNA expression levels of CXCR4 correlate with metastatic behavior and outcome in patients with osteosarcoma.', 'Clin Cancer Res', '2005', 'Messenger RNA expression levels of CXCR4 correlate with metastatic behavior and outcome in patients with osteosarcoma. To determine if osteosarcoma cells express chemokine receptors and if their presence or absence relates to clinical features.Using fluorescent quantitative real-time PCR, the pattern of 17 chemokine receptors in 3 osteosarcoma cell lines and 68 osteosarcoma patient samples was analyzed.The expression of the chemokine receptors was generally low among the cell lines. In the high-grade osteosarcoma patient samples (n = 47), CXCR4 was the most commonly expressed (63%) and its expression level was inversely correlated to overall survival (P < 0.0001), event-free survival (P < 0.001), and metastasis-free survival (MFS; P = 0.002). There was also a correlation between the expression level of CXCR4 and the presence of metastasis at diagnosis (P = 0.002). CCR7 was expressed in 43% of the samples and its expression level was inversely correlated with overall survival (P = 0.03) and MFS (P = 0.007). CCR10 mRNA expression level was inversely correlated with MFS (P = 0.009). There was no association between the expression of CXCR4, CCR7, and CCR10. Of the 26 samples studied for stromal cell-derived factor-1 expression, 77% expressed it, but there was no correlation with the clinical variables or CXCR4 expression. Multivariate analysis revealed that mRNA expression level of CXCR4 was the only significant variable for overall survival (P = 0.0006), event-free survival (P = 0.004), and MFS (P = 0.025).These data suggest that CXCR4 could be useful as a prognostic factor and as a predictor of potential metastatic development in osteosarcoma. If further studies confirm that it is relevant to metastases in this disease, it could represent a new therapeutic target.', 'http://www.ncbi.nlm.nih.gov/pubmed/15814634', 'yes', 'yes', 'NA', 3),
(15816563, 'Song JS, Park YD, Hyun JW, Kim JH', 'Induction of apoptosis by sodium fluorosilicate treatment in human osteogenic sarcoma (HOS) cells.', 'Anticancer Res', 'NA', 'Induction of apoptosis by sodium fluorosilicate treatment in human osteogenic sarcoma (HOS) cells. Fluorine compounds are widely used for the prevention of caries, and recently sodium fluorosilicate has been used in water fluorination. The cytotoxic effects of sodium fluorosilicate in several osteosarcoma and oral cancer cells were evaluated in this study by measurement of inhibition of cell proliferation. Human osteogenic sarcoma (HOS) cells were the most sensitive to sodium fluorosilicate treatment. Induction of apoptosis, such as nucleosomal DNA fragmentation and the appearance of apoptotic bodies, were observed in HOS cells by agarose gel electrophoresis and by flow cytometric analysis, respectively. The molecular mechanism of apoptosis induction in HOS was investigated by Western blot analysis. The level of Bcl-2 was decreased and consequent release of cytochrome c was increased. Caspase-3 was activated and the cleavage of poly (ADP-ribosyl) polymerase was increased. In conclusion, sodium fluorosilicate induces apoptosis in HOS cells through decrease in Bcl-2, the release of cytochrome c to the cytosol and activation of caspase-3.', 'http://www.ncbi.nlm.nih.gov/pubmed/15816563', 'no', 'no', 'sodium fluorosilicate', 3),
(15827336, 'Nakase M, Inui M, Okumura K, Kamei T, Nakamura S, Tagawa T', 'p53 gene therapy of human osteosarcoma using a transferrin-modified cationic liposome.', 'Mol Cancer Ther', '2005', 'p53 gene therapy of human osteosarcoma using a transferrin-modified cationic liposome. Gene delivery via transferrin receptors, which are highly expressed by cancer cells, can be used to enhance the effectiveness of gene therapy for cancer. In this study, we examined the efficacy of p53 gene therapy in human osteosarcoma (HOSM-1) cells derived from the oral cavity using a cationic liposome supplemented with transferrin. HOSM-1 cells were exposed to transferrin-liposome-p53 in vitro, and the growth inhibition rate, expression of p53 and bax, and induction of apoptosis were measured 48 hours later. Treatment of HOSM-1 cells with transferrin-liposome-p53 resulted in 60.7% growth inhibition. Wild-type p53 expression and an increase in bax expression were observed following transfection with transferrin-liposome-p53, and 20.5% of the treated HOSM-1 cells were apoptotic. In vivo, the HOSM-1 tumor transplanted into nude mice grew to 5 to 6 mm in diameter. Following growth of the tumor to this size, transferrin-liposome-p53 was locally applied to the peripheral tumor (day 0) and then applied once every 5 days for a total of six times. During the administration period, tumor growth did not occur, and the mean tumor volume on the last day of administration (day 25) was 10.0% of that in the saline control group. These results suggest that p53 gene therapy via cationic liposome modification with transferrin is an effective strategy for treatment of osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15827336', 'no', 'no', 'NA', 2),
(15870699, 'Azuma K, Tanaka M, Uekita T, Inoue S, Yokota J, Ouchi Y, Sakai R', 'Tyrosine phosphorylation of paxillin affects the metastatic potential of human osteosarcoma.', 'Oncogene', '2005', 'Tyrosine phosphorylation of paxillin affects the metastatic potential of human osteosarcoma. To acquire information on signal alteration corresponding to the changes in metastatic potential, we analysed protein tyrosine phosphorylation of low- and high-metastatic human osteosarcoma HuO9 sublines, which were recently established as the first metastatic model of human osteosarcoma. Tyrosine phosphorylation of proteins around 60, 70, and 120-130 kDa was enhanced in high-metastatic sublines. Among these proteins, the protein around 70 kDa, which was most remarkably phosphorylated, was identified as paxillin, a scaffold protein in integrin signaling. Activity of Src family kinase correlated well with metastatic potential, and a Src family kinase inhibitor, PP2, not only abolished tyrosine phosphorylation of paxillin but also impaired the motility of high-metastatic sublines. The expression of paxillin was also elevated in high-metastatic sublines, and knocking down of paxillin expression by RNAi method resulted in attenuated motility of high-metastatic cells. We also demonstrated that the phosphorylated form of paxillin is essential for the migration-promoting effect in human osteosarcoma. These findings suggest that enhanced activity of Src family kinases and overexpression of paxillin synergistically contribute to the high metastatic potential of human osteosarcoma through the hyperphosphorylation of paxillin.', 'http://www.ncbi.nlm.nih.gov/pubmed/15870699', 'no', 'no', 'NA', 3),
(15883045, 'Alleva R, Benassi MS, Tomasetti M, Gellert N, Ponticelli F, Borghi B, Picci P, Neuzil J', 'Alpha-tocopheryl succinate induces cytostasis and apoptosis in osteosarcoma cells: the role of E2F1.', 'Biochem Biophys Res Commun', '2005', 'Alpha-tocopheryl succinate induces cytostasis and apoptosis in osteosarcoma cells: the role of E2F1. Alpha-tocopheryl succinate (alpha-TOS), a redox-inactive analog of vitamin E, induces cell cycle arrest, differentiation, and triggers apoptosis. We examined the ability of alpha-TOS to induce cytostasis and/or apoptosis in two human osteosarcoma cell lines, which carry wild-type pRb but differ in the p53 status. In the wt-p53 cells, alpha-TOS induced apoptosis, which was associated with p53 activation and enhanced E2F1 expression. Mutant p53 cells failed to undergo apoptosis when challenged with alpha-TOS. The cell growth arrest after alpha-TOS treatment was associated with a reduced expression of E2F1. Knocking down E2F1 rendered the alpha-TOS-sensitive cells rather resistant to the apoptotic stimulus inducing a marked and prolonged cell growth arrest. We conclude that alpha-TOS induces cell growth arrest or apoptosis involving E2F1.', 'http://www.ncbi.nlm.nih.gov/pubmed/15883045', 'no', 'no', 'Alpha-tocopheryl succinate', 3),
(15900593, 'Entz-Werlé N, Stoetzel C, Berard-Marec P, Kalifa C, Brugiere L, Pacquement H, Schmitt C, Tabone MD, Gentet JC, Quillet R, Oudet P, Lutz P, Babin-Boilletot A, Gaub MP, Perrin-Schmitt F', 'Frequent genomic abnormalities at TWIST in human pediatric osteosarcomas.', 'Int J Cancer', '2005', 'Frequent genomic abnormalities at TWIST in human pediatric osteosarcomas. The identification of genes as markers for chromosome aberrations in specific tumors might facilitate oncogenesis mechanism comprehension, cancer detection, prediction of clinical outcomes, and response to therapy. Previous physiologic and oncologic data identified the TWIST gene as a marker for mesodermal derivative and bone tissue differentiation, but its contribution to bone malignancies has not been investigated. In the present study, search for genomic alterations in high-grade pediatric osteosarcomas was focused on the 7p21 region, and more specifically on the TWIST gene. In a cohort of 74 patients, we observed by allelotyping that 31 of 68 informative tumors were rearranged at the TWIST locus. Among them, analysis by quantitative PCR (QPCR) revealed that, surprisingly, mostly deletions (22/68), but also amplifications (9/68), of the TWIST gene were detected. Furthermore, deletions at TWIST were statistically correlated to other molecular abnormalities, like alterations at the APC or c-kit loci, as well as to clinical features such as a poor outcome. This work shows that the TWIST gene seemed to be involved in high-grade pediatric osteosarcomas and is a new marker with a possible initial predictive value.(c) 2005 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15900593', 'yes', 'yes', 'NA', 3),
(15912538, 'Contardi E, Palmisano GL, Tazzari PL, Martelli AM, Falà F, Fabbi M, Kato T, Lucarelli E, Donati D, Polito L, Bolognesi A, Ricci F, Salvi S, Gargaglione V, Mantero S, Alberghini M, Ferrara GB, Pistillo MP', 'CTLA-4 is constitutively expressed on tumor cells and can trigger apoptosis upon ligand interaction.', 'Int J Cancer', '2005', 'CTLA-4 is constitutively expressed on tumor cells and can trigger apoptosis upon ligand interaction. CTLA-4 (CD152) is a cell surface receptor that behaves as a negative regulator of the proliferation and the effector function of T cells. We have previously shown that CTLA-4 is also expressed on neoplastic lymphoid and myeloid cells, and it can be targeted to induce apoptosis. In our study, we have extended our analysis and have discovered that surface expression of CTLA-4 is detectable by flow cytometry on 30 of 34 (88%) cell lines derived from a variety of human malignant solid tumors including carcinoma, melanoma, neuroblastoma, rhabdomyosarcoma and osteosarcoma (but not in primary osteoblast-like cultures). However, by reverse transcriptase-PCR, CTLA-4 expression was detected in all cell lines. We have also found, by immunohistochemistry, cytoplasmic and surface expression of CTLA-4 in the tumor cells of all 6 osteosarcoma specimens examined and in the tumour cells of all 5 cases (but only weakly or no positivity at all in neighbouring nontumor cells) of ductal breast carcinomas. Treatment of cells from CTLA-4-expressing tumor lines with recombinant forms of the CTLA-4-ligands CD80 and CD86 induced apoptosis associated with sequential activation of caspase-8 and caspase-3. The level of apoptosis was reduced by soluble CTLA-4 and by anti-CTLA-4 scFvs antibodies. The novel finding that CTLA-4 molecule is expressed and functional on human tumor cells opens up the possibility of antitumor therapeutic intervention based on targeting this molecule.Copyright 2005 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/15912538', 'yes', 'no', 'NA', 3),
(15913990, 'Scotlandi K, Manara MC, Hattinger CM, Benini S, Perdichizzi S, Pasello M, Bacci G, Zanella L, Bertoni F, Picci P, Serra M', 'Prognostic and therapeutic relevance of HER2 expression in osteosarcoma and Ewing''s sarcoma.', 'Eur J Cancer', '2005', 'Prognostic and therapeutic relevance of HER2 expression in osteosarcoma and Ewing''s sarcoma. Expression of HER2 was evaluated by immunohistochemical techniques in 84 osteosarcoma (OS) and 113 Ewing''s sarcoma (ES) paraffin-embedded tumour biopsies. HER2 gene status was also assessed in a panel of cell lines as well as in vitro efficacy of trastuzumab (a humanised antibody directed against HER2) as single agent or in combination with the insulin-like growth factor I receptor (IGF-IR) IR3 antibody. Overexpression of HER2 was present in 32% of OS and 16% of ES and was significantly associated with the increased expression of P-glycoprotein, a surface molecule responsible for multidrug resistance. Event-free survival analyses revealed a prognostic value for HER2 and/or P-glycoprotein expression in OS, but not in ES. However, despite its prognostic relevance, no therapeutic effectiveness was observed pre-clinically for trastuzumab-driven therapy, in both OS or ES cell lines, unless the antibody was associated with anti-IGF-IR targeting strategies. Therefore, the therapeutic potential of trastuzumab in these neoplasms may be better exploited in combined treatments with anti-IGF-IR approaches.', 'http://www.ncbi.nlm.nih.gov/pubmed/15913990', 'yes', 'yes', 'trastuzumab', 2),
(15922739, 'Nishida Y, Knudson W, Knudson CB, Ishiguro N', 'Antisense inhibition of hyaluronan synthase-2 in human osteosarcoma cells inhibits hyaluronan retention and tumorigenicity.', 'Exp Cell Res', '2005', 'Antisense inhibition of hyaluronan synthase-2 in human osteosarcoma cells inhibits hyaluronan retention and tumorigenicity. Osteosarcoma is a common malignant bone tumor associated with childhood and adolescence. The results of numerous studies have suggested that hyaluronan plays an important role in regulating the aggressive behavior of various types of cancer cells. However, no studies have addressed hyaluronan with respect to osteosarcomas. In this investigation, the mRNA expression copy number of three mammalian hyaluronan synthases (HAS) was determined using competitive RT-PCR in the osteoblastic osteosarcoma cell line, MG-63. MG-63 are highly malignant osteosarcoma cells with an abundant hyaluronan-rich matrix. The results demonstrated that HAS-2 is the predominant HAS in MG-63. Accumulation of intracellular hyaluronan increased in association with the proliferative phase of these cells. The selective inhibition of HAS-2 mRNA in MG-63 cells by antisense phosphorothioate oligonucleotides resulted in reduced hyaluronan accumulation by these cells. As expected, the reduction in hyaluronan disrupted the assembly of cell-associated matrices. However, of most interest, coincident with the reduction in hyaluronan, there was a substantial decrease in cell proliferation, a decrease in cell motility and a decrease in cell invasiveness. These data suggest that hyaluronan synthesized by HAS-2 in MG-63 plays a crucial role in osteosarcoma cell proliferation, motility, and invasion.', 'http://www.ncbi.nlm.nih.gov/pubmed/15922739', 'no', 'no', 'NA', 1);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(15923123, 'Palmer G, Trolliet S, Talabot-Ayer D, Mézin F, Magne D, Gabay C', 'Pre-interleukin-1alpha expression reduces cell growth and increases interleukin-6 production in SaOS-2 osteosarcoma cells: Differential inhibitory effect of interleukin-1 receptor antagonist (icIL-1Ra1).', 'Cytokine', '2005', 'Pre-interleukin-1alpha expression reduces cell growth and increases interleukin-6 production in SaOS-2 osteosarcoma cells: Differential inhibitory effect of interleukin-1 receptor antagonist (icIL-1Ra1). Interleukin-1 receptor antagonist (IL-1Ra) inhibits the classical biological effects of interleukin (IL)-1 by preventing its binding to IL-1 receptors. There are, however, four different isoforms of IL-1Ra, of which three are intracellular (icIL-1Ra1, 2, 3). Due to their localization, icIL-1Ras cannot interact with cell surface IL-1 receptors and have been suggested to carry out specific functions inside cells. The recent description of nuclear effects of the IL-1alpha precursor (preIL-1alpha) led to the hypothesis that icIL-1Ra variants might antagonize the intracellular actions of preIL-1alpha. The aim of this study was to investigate effects of preIL-1alpha and icIL-1Ra1 in stably transfected SaOS-2 cells. Expression of preIL-1alpha significantly decreased cell growth and this effect was not reversed by addition of exogenous IL-1Ra, suggesting an intracellular mode of action. Transfection of SaOS-2 cells with icIL-1Ra1 did not affect their growth. Furthermore, co-expression of icIL-1Ra1 did not reverse the growth inhibitory effect of preIL-1alpha. In contrast, the production of IL-6 induced by preIL-1alpha was decreased in icIL-1Ra1 co-transfected cells. In conclusion, expression of preIL-1alpha decreased the growth of SaOS-2 cells, likely by an intracellular mechanism. Co-expression of icIL-1Ra1 did not antagonize this effect, indicating that intracellular effects of preIL-1alpha are not necessarily susceptible to inhibition by icIL-1Ra1.', 'http://www.ncbi.nlm.nih.gov/pubmed/15923123', 'no', 'no', 'NA', 3),
(15936998, 'Modrowski D, Orosco A, Thévenard J, Fromigué O, Marie PJ', 'Syndecan-2 overexpression induces osteosarcoma cell apoptosis: Implication of syndecan-2 cytoplasmic domain and JNK signaling.', 'Bone', '2005', 'Syndecan-2 overexpression induces osteosarcoma cell apoptosis: Implication of syndecan-2 cytoplasmic domain and JNK signaling. Syndecans are cell surface heparan sulfate proteoglycans that serve as co-receptors and modulate the actions of a number of extracellular ligands. Syndecans thereby regulate cell adhesion, proliferation, and differentiation. Studies in cancer cells suggest that syndecans may also modulate cell viability. We previously showed that syndecan-2 controls the growth of normal human osteoblastic cells. In this study, we examined the role of syndecan-2 in osteosarcoma cell proliferation and apoptosis. To this goal, MG63 osteosarcoma cells which express low syndecan-2 levels were transfected to overexpress full-length syndecan-2 or truncated syndecan-2 lacking its cytoplasmic domain. Determination of cell growth by cell counting and 3H-thymidine incorporation showed that truncated syndecan-2 inhibited MG63 cell proliferation. Flow cytometry analysis of DNA content and colony forming test revealed that syndecan-2, but not truncated syndecan-2, induced MG63 cell death. We show that characteristic features of apoptosis such as caspase activation, PARP cleavage, cytochrome c release, increased Bax expression, and DNA fragmentation were associated with syndecan-2-induced cell death. We further show that expression of full-length or truncated syndecan-2 induced differential activation of MAPK phosphorylation in MG63 cells. Notably, syndecan-2 but not truncated syndecan-2 overexpression increased JNK phosphorylation. Moreover, SP600125, a specific inhibitor of JNK, suppressed Bax expression induced by syndecan-2 overexpression, indicating that JNK activation mediates syndecan-2-induced apoptosis. The results show that syndecan-2 induces osteoblastic cell apoptosis through the JNK/Bax apoptotic pathway and that the cytoplasmic domain of syndecan-2 is required for this action. This supports a role for syndecan-2 in the regulation of osteosarcoma cell fate and identifies one signaling pathway by which syndecan-2 induces apoptosis in osteosarcoma cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/15936998', 'no', 'no', 'SP600125', 5),
(15939402, 'Hoberg M, Gratz HH, Noll M, Jones DB', 'Mechanosensitivity of human osteosarcoma cells and phospholipase C beta2 expression.', 'Biochem Biophys Res Commun', '2005', 'Mechanosensitivity of human osteosarcoma cells and phospholipase C beta2 expression. Bone adapts to mechanical load by osteosynthesis, suggesting that osteoblasts might respond to mechanical stimuli. We therefore investigated cell proliferation and phospholipase C (PLC) expression in osteoblasts. One Hertz uniaxial stretching at 4000 microstrains significantly increased the proliferation rates of human osteoblast-like osteosarcoma cell line MG-63 and primary human osteoblasts. However, U-2/OS, SaOS-2, OST, and MNNG/HOS cells showed no significant changes in proliferation rate. We investigated the expression pattern of different isoforms of PLC in these cell lines. We were able to detect PLC beta1, beta3, gamma1, gamma2, and delta1 in all cells, but PLC beta2 was only detectable in the mechanosensitive cells. We therefore investigated the possible role of PLC beta2 in mechanotransduction. Inducible antisense expression for 24h inhibited the translation of PLC beta1 in U-2/OS cells by 35% and PLC beta2 in MG-63 by 29%. Fluid shear flow experiments with MG-63 lacking PLC beta2 revealed a significantly higher level of cells losing attachment to coverslips and a significantly lower number of cells increasing intracellular free calcium.', 'http://www.ncbi.nlm.nih.gov/pubmed/15939402', 'no', 'no', 'NA', 1),
(15946696, 'Tanaka H, Tanabe N, Suzuki N, Shoji M, Torigoe H, Sugaya A, Motohashi M, Maeno M', 'Nicotine affects mineralized nodule formation by the human osteosarcoma cell line Saos-2.', 'Life Sci', '2005', 'Nicotine affects mineralized nodule formation by the human osteosarcoma cell line Saos-2. Several in vitro and in vivo studies have indicated that tobacco smoking may be an important risk factor for the development and severity of inflammatory periodontal disease. In the present study, we examined the effect of nicotine on cell proliferation, alkaline phosphatase (ALPase) activity, mineralized nodule formation, and the expression of extracellular matrix proteins in the human osteosarcoma cell line Saos-2. The cells were cultured with Dulbecco''s modified Eagle medium containing 10% fetal bovine serum with 0, 10(-4) M, and 10(-3) M nicotine for up to 14 days. Mineralized nodule formation was examined by alizarin red staining, and the calcium content in mineralized nodules was determined using a calcium E-test kit. The expression of extracellular matrix proteins was estimated by determining the levels of their mRNAs using the real-time polymerase chain reaction. Mineralized nodule formation and calcium content in mineralized nodules were remarkably suppressed by nicotine on days 10 and 14 of culture, respectively. ALPase activity as well as type I collagen and osteopontin expression also decreased in the presence of nicotine after 5, 10, and 14 days of culture, respectively. By contrast, the amount of bone sialoprotein increased during 14 days of culture with nicotine. These results suggest that nicotine suppresses osteogenesis through a decrease in ALPase and type I collagen production by osteoblasts.', 'http://www.ncbi.nlm.nih.gov/pubmed/15946696', 'no', 'no', 'nicotine', 4),
(15965098, 'Yen HC, Tang YC, Chen FY, Chen SW, Majima HJ', 'Enhancement of cisplatin-induced apoptosis and caspase 3 activation by depletion of mitochondrial DNA in a human osteosarcoma cell line.', 'Ann N Y Acad Sci', '2005', 'Enhancement of cisplatin-induced apoptosis and caspase 3 activation by depletion of mitochondrial DNA in a human osteosarcoma cell line. Cisplatin is an anticancer drug that can induce apoptosis. In this study, we investigated the effect of mitochondrial DNA (mtDNA) depletion on cisplatin-induced cell death using a human osteosarcoma cell line (143B) and mtDNA-depleted 143B cells (143B-rho0). Results showed that cisplatin decreased cell survival in 143B-rho0 cells. Moreover, cisplatin induced a greater extent of apoptosis-associated DNA fragmentation and caspase 3 activation in 143B-rho0 cells. The release of mitochondrial cytochrome c into cytosol by cisplatin was enhanced more obviously in 143B cells than in 143B-rho0 cells; however, in the control group of 143B-rho0 cells, it was already dramatically greater. Depletion of mtDNA may increase sensitivity of cells to cisplatin-induced apoptosis by enhancing caspase 3 activation via both cytochrome c-dependent and -independent pathways.', 'http://www.ncbi.nlm.nih.gov/pubmed/15965098', 'no', 'no', 'cisplatin', 2),
(15985268, 'Takenaka K, Yamagishi S, Jinnouchi Y, Nakamura K, Matsui T, Imaizumi T', 'Pigment epithelium-derived factor (PEDF)-induced apoptosis and inhibition of vascular endothelial growth factor (VEGF) expression in MG63 human osteosarcoma cells.', 'Life Sci', '2005', 'Pigment epithelium-derived factor (PEDF)-induced apoptosis and inhibition of vascular endothelial growth factor (VEGF) expression in MG63 human osteosarcoma cells. Pigment epithelium-derived factor (PEDF) has been shown to be the most potent inhibitor of angiogenesis in the mammalian eye, thus suggesting that loss of PEDF is involved in angiogenic eye diseases such as proliferative diabetic retinopathy. Angiogenesis is required for tumor growth and progression as well. We, along with others, have recently found that PEDF could inhibit growth of melanoma and hepatocellular carcinoma in nude mice through its anti-angiogenic effects on tumor endothelial cells. However, the possibility of the direct effect of PEDF on tumor cells has remained. In this study, we investigated the effects of PEDF on growth and vascular endothelial growth factor (VEGF) expression in MG63 human cultured osteosarcoma cells. PEDF decreased viable cell number as well as DNA synthesis in MG63 cells in a dose-dependent manner. Furthermore, PEDF was found to increase caspase-3/7 activity and to subsequently induce apoptotic cell death in MG63 cells. PEDF also inhibited VEGF expression in MG63 cells at both mRNA and protein levels. Our present study provides novel beneficial aspects of PEDF on osteosarcoma cells; one is induction of apoptotic cell death of tumor cells, and the other is the suppression of VEGF expression, which would lead to inhibition of tumor angiogenesis. PEDF therefore might be a promising therapeutic agent for treatment of patients with osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/15985268', 'no', 'no', 'PEDF', 3),
(16007340, 'Amaar YG, Baylink DJ, Mohan S', 'Ras-association domain family 1 protein, RASSF1C, is an IGFBP-5 binding partner and a potential regulator of osteoblast cell proliferation.', 'J Bone Miner Res', '2005', 'Ras-association domain family 1 protein, RASSF1C, is an IGFBP-5 binding partner and a potential regulator of osteoblast cell proliferation. The goal of this study was to identify downstream signaling molecules involved in mediating the IGF-independent effects of IGFBP-5 in osteoblasts. We identified RASSF1C, a member of the RASSF1 gene products, as a IGFBP-5 binding partner and as a potential mediator of IGFBP-5 effects on ERK phosphorylation and cell proliferation.It has been predicted that the intrinsic growth factor action of insulin-like growth factor binding protein (IGFBP)-5 involves either the binding of IGFBP-5 to a putative receptor to induce downstream signaling pathways and/or intracellular translocation of IGFBP-5 to bind to potential signaling molecules involved in osteoblast cell regulation. This study reports the characterization of isoform C of the Ras association family 1 (RASSF1C) gene as an interacting partner of IGFBP-5.IGFBP-5 was used as bait in a yeast two-hybrid screen of a human osteosarcoma cDNA library. Expression levels of RASSF1C were measured by RT-PCR and/or Northern blot. IGFBP-5 effects on ERK phosphorylation were evaluated by immunoblot analysis. The effect of RASSF1C siRNA on cell proliferation was measured by the AlamarBlue assay.One of the clones that interacted strongly with the bait under high stringency conditions corresponded to RASSF1C. The interaction between RASSF1C and IGFBP-5 was confirmed by in vitro co-immunoprecipitation studies. Northern blot and RT-PCR analysis showed that RASSF1C was expressed in a variety of osteoblast cell types that produce IGFBP-5. Addition of synthetic RASSF1C-specific small interfering (si) RNA duplex or use of a RASSF1C-specific si-hairpin plasmid caused a decrease in cell number and abolished IGFBP-5-induced extracellular signal-regulated kinase (ERK)-1/2 phosphorylation but had no effect on IGFBP-5-induced increases in alkaline phosphatase (ALP) activity.We have shown a novel interaction between IGFBP-5 and RASSF1C. Our findings that silencing of RASSF1C results in the reduction of osteoblast cell proliferation and that IGFBP-5 treatment increases phosphorylation of ERK-1/2 raise the possibility that RASSF1C, a Ras effector, could, in part, contribute to mediating the effects of IGFBP-5 on ERK phosphorylation and, consequently, cell proliferation.', 'http://www.ncbi.nlm.nih.gov/pubmed/16007340', 'no', 'no', 'NA', 5),
(16007579, 'Hughes DP, Thomas DG, Giordano TJ, McDonagh KT, Baker LH', 'Essential erbB family phosphorylation in osteosarcoma as a target for CI-1033 inhibition.', 'Pediatr Blood Cancer', '2006', 'Essential erbB family phosphorylation in osteosarcoma as a target for CI-1033 inhibition. The role of erbB tyrosine kinases, especially Her-2, in osteosarcoma has engendered intense debate. Some investigators identified an association between low-level Her-2 expression, compared to none, and poor patient outcome. Others questioned the importance of apparent cytoplasmic expression of Her-2, since membranous overexpression is associated with poor outcome in carcinomas. We previously demonstrated that primary osteosarcoma cells express cell-surface EGFR and Her-2, with the p80 isoform of Her-4 localized to the nucleus. We wished to determine if erbB kinases in osteosarcoma were phosphorylated, and if this was required for growth.We cultured early passage osteosarcoma cell lines in the presence or absence of the pan-erbB inhibitor CI-1033 and examined the phosphorylation status of EGFR, Her-2, and Her-4 by immunohistochemistry, cell-based ELISA, flow cytometry and two dimensional Western blot. We also assessed the impact of CI-1033 upon osteosarcoma growth and survival in vitro.EGFR, Her-2, and Her-4 were constitutively phosphorylated in early passage osteosarcoma cells cultured in vitro. CI-1033 abrogated erbB receptor phosphorylation and caused growth inhibition and apoptosis in a titratible fashion with concentrations of 1 muM or more.EGFR, Her-2, and Her-4 are constitutively phosphorylated in early passage osteosarcoma cells in tissue culture, and erbB signaling provides essential growth and anti-apoptotic signals to osteosarcoma cells. This suggests that erbB overexpression is not required for erbB to promote malignancy, but rather that overexpression is one of several mechanisms that generate unregulated erbB signaling.', 'http://www.ncbi.nlm.nih.gov/pubmed/16007579', 'no', 'no', 'CI-1033', 3),
(16008557, 'de Bock CE, Lin Z, Itoh T, Morris D, Murrell G, Wang Y', 'Inhibition of urokinase receptor gene expression and cell invasion by anti-uPAR DNAzymes in osteosarcoma cells.', 'FEBS J', '2005', 'Inhibition of urokinase receptor gene expression and cell invasion by anti-uPAR DNAzymes in osteosarcoma cells. The urokinase-type plasminogen activator (uPA) receptor (uPAR) has been implicated in signal transduction and biological processes including cancer metastasis, angiogenesis, cell migration, and wound healing. It is a specific cell surface receptor for its ligand uPA, which catalyzes the formation of plasmin from plasminogen, thereby activating the proteolytic cascade that contributes to the breakdown of extracellular matrix, a key step in cancer metastasis. We have synthesized three different DNA enzymes (Dz372, Dz483 and Dz720) targeting uPAR mRNA at three separate purine (A or G)-pyrimidine (U or C) junctions. Two of these DNAzymes, Dz483 and Dz720, cleaved uPAR transcript in vitro with high efficacy and specificity at a molar ratio (uPAR to Dz) as low as 1 : 0.2. When analyzed over 2 h with a 200-fold molar excess of DNAzymes to uPAR transcript, Dz720 and Dz483 were able to decrease uPAR transcript in vitro by approximately 93% and approximately 84%, respectively. They also showed an ability to cleave uPAR mRNA in the human osteosarcoma cell line Saos-2 after transfection. The DNAzyme Dz720 decreased uPAR mRNA within 4 h of transfection, and inhibited uPAR protein concentrations by 55% in Saos-2 cells. The decrease in uPAR mRNA and protein concentrations caused by Dz720 significantly suppressed Saos-2 cell invasion as assessed by an in vitro Matrigel assay. The use of DNAzyme methodology adds a new potential clinical agent for decreasing uPAR mRNA expression and inhibiting cancer invasion and metastasis.', 'http://www.ncbi.nlm.nih.gov/pubmed/16008557', 'yes', 'no', 'NA', 2),
(16010295, 'Ahlström M, Pekkinen M, Huttunen M, Lamberg-Allardt C', 'Cyclic nucleotide phosphodiesterases (PDEs) in human osteoblastic cells; the effect of PDE inhibition on cAMP accumulation.', 'Cell Mol Biol Lett', '2005', 'Cyclic nucleotide phosphodiesterases (PDEs) in human osteoblastic cells; the effect of PDE inhibition on cAMP accumulation. The regulation of the secondary messengers, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), is crucial in the hormonal regulation of bone metabolism. Both cAMP and cGMP are inactivated by cyclic nucleotide phosphodiesterases (PDEs), a superfamily of enzymes divided into 11 families (PDE1-11). We compared the PDEs of cultured human osteoblasts (NHOst) and SaOS-2 osteosarcoma cells. The PDE activity of NHOst cells consisted of PDE1, PDE3 and PDE7, whereas PDE1, PDE7 and PDE4, but no PDE3 activity was detected in SaOS-2 cells. In line with the difference in the PDE profiles, rolipram, a PDE4 inhibitor, increased the accumulation of cAMP in SaOS-2, but not in NHOst cells. Expression of PDE subtypes PDE1C, PDE3A, PDE4A, PDE4B, PDE7A and PDE7B was detected in both cell types. NHOst cells additionally expressed PDE1A.', 'http://www.ncbi.nlm.nih.gov/pubmed/16010295', 'no', 'no', 'rolipram', 5),
(16026999, 'Yang DS, Miao XD, Ye ZM, Feng J, Xu RZ, Huang X, Ge FF', 'Bacterial redox protein azurin induce apoptosis in human osteosarcoma U2OS cells.', 'Pharmacol Res', '2005', 'Bacterial redox protein azurin induce apoptosis in human osteosarcoma U2OS cells. As a low molecular weight redox protein elaborated from the pathogenic bacteria Pseudomonas aeruginosa, azurin is one of representative bacterial products applied in the treatment of tumour. We found that the growth of U2OS cells was significantly inhibited by azurin in a dose-dependent manner with the IC(50) value of 114.54+/-7.65 mgl(-1). But the growth of MG63 cells or L02 cells was almost not inhibited by azurin (P<0.05). Moreover, when treated with azurin, U2OS cells showed typical apoptotic morphological features observed by fluorescent microscopy (AO and Hoechst 33258) and transmission electron microscopy. Typical DNA \\ladder\\ bands were also observed. The apoptosis rate was 35.8% tested by fluorescence-activated cell sorter (Annexin-V-FITC(+)/PI(-)) and the cell-cycle arrested in G(1) phase. But no apoptotic features were observed in control cells. The down-regulation of Bcl-2 (an inhibitor of apoptosis) were detected in U2OS cells when azurin was added for 24h. In contrast, the level of Bax and caspase-3 were significantly up-regulated. So we concluded that azurin could selectively induce apoptosis of human osteosarcoma U2OS cells and the induction of apoptosis by azurin was closely associated with down-regulation of Bcl-2, up-regulation of Bax and activation of caspase-3.', 'http://www.ncbi.nlm.nih.gov/pubmed/16026999', 'no', 'no', 'azurin', 3),
(16035619, 'Duan X, Jia SF, Zhou Z, Langley RR, Bolontrade MF, Kleinerman ES', 'Association of alphavbeta3 integrin expression with the metastatic potential and migratory and chemotactic ability of human osteosarcoma cells.', 'Clin Exp Metastasis', '2004', 'Association of alphavbeta3 integrin expression with the metastatic potential and migratory and chemotactic ability of human osteosarcoma cells. Expression of adhesion molecules such as alphavbeta3 integrin has been associated with the metastatic potential of tumor cells. The purpose of this study was to determine whether alphavbeta3 expression correlated with the metastatic potential of human osteosarcoma cells.We developed a series of sublines (LM2-LM7) from human osteosarcoma SAOS parental cells, with progressively increasing potential to form lung metastases in nude mice after intravenous injection. SAOS parental and LM2 cells were poorly metastatic, but LM7 cells resulted in visible metastatic lung nodules by 6-8 weeks. We quantified alphavbeta3 integrin expression using flow cytometry.alphavbeta3 expression correlated with the metastatic potential of the cells, with LM7 cells showing the highest expression. LM7 cell adhesion to vitronectin decreased after treatment with echistatin, a RGD-containing peptide antagonist of alphavbeta3. LM7 cells demonstrated higher chemotactic activity than SAOS cells to a homogenate made from lung tissue. This chemotactic activity was also inhibited by echistatin. These data indicated that alphavbeta3 was critical for the migration of LM7 cells to the lung homogenate. Chemotaxis to a liver homogenate was the same for LM7 and SAOS cells. Migration of LM7 cells through lung endothelial cells was higher than that through liver endothelial cells, and echistatin again inhibited this migration.alphavbeta3 integrin expression may play a role in the metastatic potential of osteosarcoma cells by enhancing the ability of the cells to migrate specifically to the lung. Alphavbeta3 integrin may therefore be a potential new target for osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16035619', 'no', 'no', 'NA', 2),
(16083752, 'Bjørnland K, Flatmark K, Pettersen S, Aaasen AO, Fodstad O, Maelandsmo GM', 'Matrix metalloproteinases participate in osteosarcoma invasion.', 'J Surg Res', '2005', 'Matrix metalloproteinases participate in osteosarcoma invasion. Osteosarcoma (OS) is a highly malignant bone tumor and is the most frequent malignant bone tumor in children and adolescents. Metastases are the major cause of death, and patients with relapse have poor prognosis. Several solid tumors display enhanced expression of matrix metalloproteinases (MMPs), and recently MMP-inhibitors have entered clinical trials. A disturbance of the MMP system in favor of enhanced proteolytic activity may be suspected in OS because OS growth is accompanied by both enhanced local bone destruction and bone formation, two processes that are dependant on proteolytic enzymes. Thus, the aim of the present study was to evaluate the involvement of MMPs in a panel of human OS cell lines, xenografts and biopsies.Expression of MMPs and their endogenous inhibitors were studied by zymography and Northern blot analyses. In vitro invasion of OS cell lines and effects of MMP-inhibitors (Marimastat and doxycycline) were assessed in the transwell chamber assay.In vitro invasiveness was compared with gelatinase activity, and the most invasive cell line secreted the highest amounts of MMP-2 and MMP-9. Two different MMP-inhibitors significantly reduced OS cell invasion. The majority of the OS xenografts expressed both the inactive and active form of MMP-2 and in some cases also MMP-9. The biopsies from primary and metastatic OS also expressed MMP-2 mRNA. However, MMP-9 levels were higher in the biopsies than in the xenografts.The obtained results support the hypothesis that MMPs and their endogenous inhibitors participate in the invasive process of human OS.', 'http://www.ncbi.nlm.nih.gov/pubmed/16083752', 'no', 'no', 'NA', 2),
(16119739, 'Jung ST, Moon ES, Seo HY, Kim JS, Kim GJ, Kim YK', 'Expression and significance of TGF-beta isoform and VEGF in osteosarcoma.', 'Orthopedics', '2005', 'Expression and significance of TGF-beta isoform and VEGF in osteosarcoma. This article demonstrates that the degree of VEG expression could be used as a clinically signific prognostic factor in osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16119739', 'yes', 'yes', 'NA', 1),
(16140185, 'Lu KH, Lue KH, Chou MC, Chung JG', 'Paclitaxel induces apoptosis via caspase-3 activation in human osteogenic sarcoma cells (U-2 OS).', 'J Orthop Res', '2005', 'Paclitaxel induces apoptosis via caspase-3 activation in human osteogenic sarcoma cells (U-2 OS). Paclitaxel has been found to exhibit cytotoxic and antitumor activity. There is little information regarding the mechanisms of apoptotic-inducing effect of paclitaxel on human osteogenic sarcoma U-2 OS cells. Several key regulatory proteins are involved in the initiation of apoptosis. Caspase-3 plays a direct role in proteolytic cleavage of cellular proteins responsible for progression to apoptosis. We examined the effect of paclitaxel on the cell cycle arrest and apoptosis in U-2 OS cells using flow cytometric analysis and Western blotting. We also measured the inhibition of paclitaxel-induced apoptosis and the caspase-3 activity by the broad-spectrum caspase inhibitor z-VAD-fmk on U-2 OS cells. The increased levels of casapse-3 were also confirmed by cDNA microarray. Our observations were: (1) paclitaxel treatment resulted in G2/M-cycle arrest in U-2 OS cells; (2) time and dose dependent apoptosis of U-2 OS cells was induced by paclitaxel; (3) in U-2 OS cells, z-VAD-fmk blocked the paclitaxel-induced apoptosis and caspase-3 activation. These results suggest that paclitaxel-induced G2/M-cycle arrest of the G2/M phase and apoptosis via a caspase-3 pathway in U-2 OS cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16140185', 'no', 'no', 'paclitaxel', 1),
(16157226, 'Luu HH, Zhou L, Haydon RC, Deyrup AT, Montag AG, Huo D, Heck R, Heizmann CW, Peabody TD, Simon MA, He TC', 'Increased expression of S100A6 is associated with decreased metastasis and inhibition of cell migration and anchorage independent growth in human osteosarcoma.', 'Cancer Lett', '2005', 'Increased expression of S100A6 is associated with decreased metastasis and inhibition of cell migration and anchorage independent growth in human osteosarcoma. While most osteosarcoma patients have metastatic or micrometastatic lesions, less than 15% of them have clinically detectable metastatic diseases at presentation. To identify potential markers that may predict osteosarcoma metastasis, we analyzed the expression of S100A6 in 50 osteosarcoma cases and found that 84% of the analyzed specimens stained positive for S100A6. There is a trend towards decreased clinically evident metastasis with increased S100A6 staining. Overexpression of S100A6 in osteosarcoma cells decreases cell motility and anchorage independent growth on collagen gels. Our findings provide evidence that, while S100A6 is commonly overexpressed in human osteosarcoma, loss of its expression correlates with a metastatic phenotype.', 'http://www.ncbi.nlm.nih.gov/pubmed/16157226', 'yes', 'yes', 'NA', 1),
(16167336, 'Eliseev RA, Zuscik MJ, Schwarz EM, O''Keefe RJ, Drissi H, Rosier RN', 'Increased radiation-induced apoptosis of Saos2 cells via inhibition of NFkappaB: a role for c-Jun N-terminal kinase.', 'J Cell Biochem', '2005', 'Increased radiation-induced apoptosis of Saos2 cells via inhibition of NFkappaB: a role for c-Jun N-terminal kinase. To elucidate the possible effect of NFkappaB on radioresistance, we used the osteosarcoma cell line Saos2, stably expressing the NFkappaB constitutive inhibitor, mIkappaB (Saos2-mIkappaB) or stably transfected with the empty vector (Saos2-EV). Ionizing radiation induced \\intrinsic\\ apoptosis in Saos2-mIkappaB cells but not in Saos2-EV control cells, with intact NFkappaB activity. We find as expected, that this NFkappaB activity was enhanced following irradiation in the Saos2-EV control cells. On the other hand, inhibition of NFkappaB signaling in Saos2-mIkappaB cells led to the upregulation of the pro-apoptotic systems, such as Bax protein and c-Jun N-terminal Kinase (JNK)/c-Jun/AP1 signaling. Inhibition of NFkappaB resulted in decreased expression of the DNA damage protein GADD45beta, a known inhibitor of JNK. Subsequently, JNK activation of c-Jun/AP-1 proteins increased radiation-induced apoptosis in these mutants. Radiation-induced apoptosis in Saos2-mIkappaB cells was inhibited by the JNK specific inhibitor SP600125 as well as by Bcl-2 over-expression. Furthermore, release of cytochrome-c from mitochondria was increased and caspase-9 and -3 were activated following irradiation in Saos2-mIkappaB cells. Antisense inhibition of GADD45beta in Saos2-EV cells significantly enhanced apoptosis following irradiation. Our results demonstrate that radioresistance of Saos2 osteosarcoma cells is due to NFkappaB-mediated inhibition of JNK. Our study brings new insight into the mechanisms underlying radiation-induced apoptosis of osteosarcoma, and may lead to development of new therapeutic strategies against osteosarcoma.Copyright 2005 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/16167336', 'no', 'no', 'NA', 6),
(16196299, 'Yi C, Li X, Xu W, Chen A', 'Relationship between the expression of MTA-1 gene and the metastasis and invasion in human osteosarcoma.', 'J Huazhong Univ Sci Technolog Med Sci', '2005', 'Relationship between the expression of MTA-1 gene and the metastasis and invasion in human osteosarcoma. To compare the expression level of metastasis associated-1 (MTA1) gene in high and low metastatic human osteosarcoma cell lines and examine the relationship of MTA1 expression and the metastasis potentiality of osteosarcoma cells, the expression of MTA1 in MG-63 osteosarcoma cell lines with high and low metastasis potential was detected by semiquantitative TR-PCR. Boyden chamber invasion assay was used to evaluate the invasive capacity in vitro in two osteosarcoma cell lines. The low metastasis MG-63 cells were transfected with MTA1 full-length cDNA expression plasmid by lipofectamine and the changes of MTA1 expression and in vitro invasion potential were examined after the transfection. Our results showed that MG63 cell line with high metastasis potential expressed significantly higher MTA1 than that of MG63 cells with low metastasis as reavealed by RT-PCR. The invasion potential of low metastasis MG63 cell line was increased after MTA1 gene transfection. It is concluded that there may be a relationship between MTA 1 and invasive potentiality of human osteosarcoma cells, and the mechanism of MTA1 in osteosarcoma metastasis and its possible role in associated gene therapy deserve further study.', 'http://www.ncbi.nlm.nih.gov/pubmed/16196299', 'no', 'no', 'NA', 1),
(16206219, 'Roque L, Lacerda A, Rodrigues R, Almeida O, Salgueiro J, Pinto C', 'CGH evaluation of two de novo synchronous tumors in a child with a germline p53 mutation.', 'Pediatr Blood Cancer', '2006', 'CGH evaluation of two de novo synchronous tumors in a child with a germline p53 mutation. We report the case of a child who developed two de novo synchronous tumors: an osteosarcoma and an embryonal rhabdomyosarcoma. The patient was determined to be a de novo carrier of a P53 germline mutation. Comparative genomic hybridization (CGH) analysis revealed that each of the neoplasms was characterized by a specific set of chromosomal imbalances and high-level amplification (HLA) regions. Our CGH findings provide evidence that cancer development is a cellular/organ specific event.', 'http://www.ncbi.nlm.nih.gov/pubmed/16206219', 'yes', 'no', 'NA', 1),
(16211088, 'Ganjavi H, Gee M, Narendran A, Parkinson N, Krishnamoorthy M, Freedman MH, Malkin D', 'Adenovirus-mediated p53 gene therapy in osteosarcoma cell lines: sensitization to cisplatin and doxorubicin.', 'Cancer Gene Ther', '2006', 'Adenovirus-mediated p53 gene therapy in osteosarcoma cell lines: sensitization to cisplatin and doxorubicin. The poor prognosis for patients with metastatic osteosarcoma (OS) indicates that new therapeutic options should be explored. Studies with adenoviral-mediated p53 gene transfer have been conducted in many cancer types including cervical, ovarian, prostatic and head and neck tumors. However, limited work has been carried out with pediatric cancers, including OS. Using three viral constructs containing cDNA for wild-type p53, mutant p53 (Cys135Ser) and lacZ, we studied the effect of adenoviral-mediated gene therapy in four OS cell lines: Saos-2 (p53-/-), HOS (R156P), KHOS/NP (R156P) and MNNG (R156P, F270L). We demonstrated that the virus efficiently enters the cells using the beta-galactosidase assay. Using the MTT assay, we have shown a dose-dependent decrease in cell viability 72 h post-treatment that occurs with Ad-wtp53 but not with Ad-mutp53. We have also shown that treatment with Ad-wtp53 significantly increases sensitivity of the cell lines to cisplatin and doxorubicin, chemotherapeutic agents commonly used in the treatment of OS. Our results indicate that restoration of wt p53 function in OS cells provides a basis for novel approaches to treatment of this disease.', 'http://www.ncbi.nlm.nih.gov/pubmed/16211088', 'yes', 'no', 'doxorubicin|cisplatin', 1),
(16211448, 'Somers GR, Ho M, Zielenska M, Squire JA, Thorner PS', 'HER2 amplification and overexpression is not present in pediatric osteosarcoma: a tissue microarray study.', 'Pediatr Dev Pathol', 'NA', 'HER2 amplification and overexpression is not present in pediatric osteosarcoma: a tissue microarray study. The HER2 gene, located on 17q, encodes a 185-kD transmembrane tyrosine kinase receptor. Amplification of this gene with overexpression of the gene product occurs in about 30% of cases of breast cancer and is considered to be a poor prognostic indicator for this tumor. Results for HER2 expression in osteosarcoma are controversial, with some studies reporting up to 61% of positive cases and others reporting only negative results. Further, expression of HER2 is reported to be a favorable prognostic indicator by some groups and unfavorable by others. The present study used tissue microarrays containing 34 samples of osteosarcoma from 18 patients to analyze HER2 expression by immunohistochemistry and gene copy number by chromogenic in situ hybridization. The microarray included 13 pretreatment biopsies, 11 posttreatment resection specimens, and 10 resected metastases and comprised 18 osteoblastic, 6 chondroblastic, 5 fibroblastic, and 5 mixed subtypes. HER2 protein expression was seen in 4 of 34 (12%) tumor samples that originated from 2 of 18 patients (11%). The staining pattern was consistently weak and focal, and immunohistochemical overexpression of the HER2 protein, defined as complete membrane positivity, was never observed. Further, the presence of HER2 gene amplification was not detected in any osteosarcoma by chromogenic in situ hybridization. Therefore, therapies based on antibodies directed against the HER2 protein are unlikely to have much value in the treatment of pediatric osteosarcomas. From a technical standpoint, this study also demonstrates the value of tissue microarrays in screening tumors at the protein and gene levels using conventional light microscopy.', 'http://www.ncbi.nlm.nih.gov/pubmed/16211448', 'yes', 'no', 'NA', 1),
(16213662, 'Jang HS, Kook SH, Son YO, Kim JG, Jeon YM, Jang YS, Choi KC, Kim J, Han SK, Lee KY, Park BK, Cho NP, Lee JC', 'Flavonoids purified from Rhus verniciflua Stokes actively inhibit cell growth and induce apoptosis in human osteosarcoma cells.', 'Biochim Biophys Acta', '2005', 'Flavonoids purified from Rhus verniciflua Stokes actively inhibit cell growth and induce apoptosis in human osteosarcoma cells. Many studies have suggested that dietary flavonoids are anticancer agents that induce the apoptosis of cancer cells. However, the effects of flavonoids on the induction of apoptosis in osteosarcoma cells are unclear. Previously, a flavonoid fraction, consisting mainly of protocatechuic acid, fustin, fisetin, sulfuretin, and butein, herein named RCMF (the RVS chloroform-methanol fraction), was prepared from a crude acetone extract of Rhus verniciflua Stokes (RVS). This study evaluated the effects of RCMF on the proliferation and apoptosis using human osteosarcoma (HOS) cells. The mechanism of growth inhibition of the HOS cells by the flavonoid fraction, RCMF, was also assessed. The results demonstrated that RCMF exhibited sensitive growth inhibition and induced apoptosis in HOS cells. PARP cleavage was closely associated with the RCMF-induced apoptosis of the HOS cells. Furthermore, the activation of caspase 8 and Bax, the inhibition of Bcl-2 expression, and the release of cytochrome c are believed to be involved in the RCMF-mediated apoptosis. Collectively, these findings suggest that RCMF is an agent which may be capable of inducing sensitive growth inhibition and apoptosis in HOS cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16213662', 'no', 'no', 'Rhus verniciflua Stokes', 5),
(16219634, 'Naruse T, Nishida Y, Hosono K, Ishiguro N', 'Meloxicam inhibits osteosarcoma growth, invasiveness and metastasis by COX-2-dependent and independent routes.', 'Carcinogenesis', '2006', 'Meloxicam inhibits osteosarcoma growth, invasiveness and metastasis by COX-2-dependent and independent routes. Cyclooxygenase-2 (COX-2) inhibitors exert antitumor activity via COX-2-dependent and independent pathways. We wished to evaluate the antitumor activity of meloxicam, a preferential COX-2 inhibitor, in osteosarcoma, the most common primary malignant bone tumor, and determine whether its antitumor effect is COX-2-dependent. COX-2 expression in the osteosarcoma cell lines MG-63, HOS and U2-OS was determined by real-time RT-PCR and western blotting. Subsequently, the inhibitory effects of meloxicam on osteosarcoma cell growth and invasiveness were assayed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and matrigel invasion assays, respectively. Apoptotic activity was evaluated by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling staining and semi-quantification of Bax and Bcl-2 expression by real time RT-PCR and western blotting. Prostaglandin-E(2) (PGE(2)) production in the presence and absence of meloxicam was analyzed by enzyme immunoassay, and to determine whether the effects of meloxicam are COX-2-dependent or independent, PGE(2) was added to see if it reversed the effects of meloxicam. In addition, the effects of meloxicam on tumor growth and metastasis were evaluated in an in vivo mouse model using grafted LM-8 mouse osteosarcoma cells, together with immunohistochemical analysis for vascular endothelial growth factor in lung metastatic lesion. Meloxicam inhibited PGE(2) production, proliferation and invasiveness especially in MG-63 cells, which express relatively high levels of COX-2. Only high concentrations of meloxicam caused apoptosis and upregulated Bax mRNA and protein in MG-63 cell culture. In contrast, meloxicam did not induce apoptosis in HOS and U2-OS cells, expressing relatively low levels of COX-2. Exogenous PGE(2) reduced the effects of meloxicam on cell viability and invasiveness, but not its effect on Bax mRNA. In vivo, high doses of meloxicam suppressed LM-8 tumor growth and lung metastasis. Meloxicam, may have both COX-2-dependent and independent inhibitory actions on osteosarcoma. Its effects are more prominent in osteosarcoma cells that have relatively high levels of COX-2.', 'http://www.ncbi.nlm.nih.gov/pubmed/16219634', 'no', 'no', 'meloxicam', 2),
(16259979, 'Eliseev RA, Schwarz EM, Zuscik MJ, O''Keefe RJ, Drissi H, Rosier RN', 'Smad7 mediates inhibition of Saos2 osteosarcoma cell differentiation by NFkappaB.', 'Exp Cell Res', '2006', 'Smad7 mediates inhibition of Saos2 osteosarcoma cell differentiation by NFkappaB. The transcription factor NFkappaB is constitutively activated in various tumor cells where it promotes proliferation and represses apoptosis. The bone morphogenetic proteins (BMPs) delay cell proliferation and promote differentiation and apoptosis of bone cells through activation of Smad downstream effectors and via Smad-independent mechanisms. Thus, NFkappaB and BMP pathways play opposing roles in regulating osteoblastic cell fate. Here, we show that in osteosarcoma Saos2 osteoblasts, NFkappaB regulates the activity of the BMP/Smad signaling. Inhibition of NFkappaB by overexpression of mIkappaB leads to the induction of osteoblast differentiation. Saos2 cells overexpressing mIkappaB (Saos2-mIkappaB) exhibit higher expression of osteoblast phenotypic genes such as alkaline phosphatase, Runx2 and osteocalcin and are more responsive to BMP2 in comparison to wild-type cells (Saos2-wt) or empty vector infected controls (Saos2-EV). Furthermore, BMP-2 signaling and Smad phosphorylation are significantly increased in Saos2-mIkappaB cells in comparison to Saos2-EV cells. Inhibition of NFkappaB signaling in Saos2-mIkappaB cells is associated with decreased expression of the BMP signaling inhibitor Smad7. While gain of Smad7 function in Saos2-mIkappaB cells results in inhibition of BMP signaling, anti-sense knockdown of Smad7 in Saos2-EV cells leads to upregulation of BMP signaling. We therefore conclude that in osteosarcoma Saos2 cells, NFkappaB represses BMP/Smad signaling and BMP2-induced differentiation through Smad7.', 'http://www.ncbi.nlm.nih.gov/pubmed/16259979', 'no', 'no', 'NA', 6),
(16269403, 'Amaar YG, Tapia B, Chen ST, Baylink DJ, Mohan S', 'Identification and characterization of novel IGFBP5 interacting protein: evidence IGFBP5-IP is a potential regulator of osteoblast cell proliferation.', 'Am J Physiol Cell Physiol', '2006', 'Identification and characterization of novel IGFBP5 interacting protein: evidence IGFBP5-IP is a potential regulator of osteoblast cell proliferation. Insulin-like growth factor binding protein-5 (IGFBP5) is a multifunctional protein, which acts not only as a traditional binding protein, but also functions as a growth factor independent of IGFs to stimulate bone formation. It has been predicted that the intrinsic growth factor action of IGFBP5 involves binding of IGFBP5 to a putative receptor to induce downstream signaling pathways and/or nuclear translocation of IGFBP5 to influence transcription of genes involved in osteoblast cell proliferation/differentiation. Our study indentified proteins that bound to IGFBP5 using IGFBP5 as bait in a yeast two-hybrid screen of the U2 human osteosarcoma cell cDNA library. One of the clones that interacted strongly with the bait under high-stringency conditions corresponded to a novel IGFBP5 interacting protein (IGFBP5-IP) encoded by a gene that resides in mouse chromosome 10. The interaction between IGFBP5-IP and IGFBP5 is confirmed by in vitro coimmunoprecipitation studies that used pFlag and IGFBP5 polyclonal antibody, and cell lysates overexpressing both IGFBP5-IP and IGFBP5. Northern blot and RT-PCR analysis showed that the IGFBP-IP is expressed in both untransformed normal human osteoblasts and in osteosarcoma cell lines, which are known to produce IGFBP5. To determine the roles of IGFBP5-IP, we evaluated the effect of blocking the expression of IGFBP5-IP on osteoblast proliferation. We found that using a IGFBP5-IP-specific small interfering-hairpin plasmid resulted in a decrease in both basal and IGFBP5-induced osteoblast cell proliferation. On the basis of these findings, we predict that IGFBP5-IP may act as intracellular mediator of growth promoting actions of IGFBP5 and perhaps other osteoregulatory agents in bone cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16269403', 'no', 'no', 'NA', 1),
(16273216, 'Meschini S, Marra M, Condello M, Calcabrini A, Federici E, Dupuis ML, Cianfriglia M, Arancia G', 'Voacamine, an alkaloid extracted from Peschiera fuchsiaefolia, inhibits P-glycoprotein action in multidrug-resistant tumor cells.', 'Int J Oncol', '2005', 'Voacamine, an alkaloid extracted from Peschiera fuchsiaefolia, inhibits P-glycoprotein action in multidrug-resistant tumor cells. Multidrug resistance (MDR) in tumor cells is generally associated with increased efflux of the cytotoxic compounds, due to the activation of mechanisms of intracellular transport and to the overexpression of surface proteins, such as P-glycoprotein (Pgp), which act as ATP-dependent molecular pumps. In a previous study, voacamine, a bisindolic alkaloid from Peschiera fuchsiaefolia, was examined for its possible capability of enhancing the cytotoxic effect of doxorubicin (DOX) on resistant human osteosarcoma cells. The effects of voacamine on the cell survival and on accumulation of DOX were investigated on both the parental cell line, U-2 OS-WT, and its resistant counterpart, U-2 OS-R. A differential effect between sensitive and resistant cells on the intracellular DOX concentration and distribution was revealed. In particular, voacamine induced a significant increase of drug retention and intranuclear location in resistant cells. Moreover, the cell survival analysis and the electron microscopic observations revealed an enhancement of the cytotoxic effect of DOX induced by the plant extract. In the present study, a panel of monoclonal antibodies (MAbs), recognizing different and specific structural and functional state of Pgp, was used. By flow cytometry and immunofluorescence confocal microscopy, a dose-dependent increase of the reactivity of Pgp with MAb UIC2, which specifically recognizes an epitope of the drug transporter in its functional conformation, was detected in voacamine-treated U-2 OS-R cells. Conversely, the expression of the epitope recognized by MAb MC57 was downregulated while MAb MM4.17 did not change its binding level to treated and untreated MDR cells. These data suggest that the plant extract reacts with Pgp producing conformational changes with consequent epitope modulation. Taken together, our observations seem to demonstrate that voacamine is a substrate for Pgp and, therefore, interferes with the Pgp-mediated drug export, acting as a competitive antagonist of cytotoxic agents.', 'http://www.ncbi.nlm.nih.gov/pubmed/16273216', 'no', 'no', 'voacamine|doxorubicin', 1),
(16282894, 'Gordon N, Arndt CA, Hawkins DS, Doherty DK, Inwards CY, Munsell MF, Stewart J, Koshkina NV, Kleinerman ES', 'Fas expression in lung metastasis from osteosarcoma patients.', 'J Pediatr Hematol Oncol', '2005', 'Fas expression in lung metastasis from osteosarcoma patients. The authors'' animal studies have shown that the metastatic potential of osteosarcoma (OS) cells correlates inversely with Fas expression-that is, Fas-negative cells metastasize but Fas-positive cells do not. One reason for this in the context of OS lung metastases may be that Fas-positive cells are eliminated by engagement with the Fas ligand (FasL) constitutively expressed on the surface of pneumocytes, whereas Fas-negative tumor cells are not. The purpose of this study was to determine the status of Fas expression in OS lung metastases from patients. Specifically, archived paraffin-embedded specimens of lung metastases from 38 patients with OS were analyzed by immunohistochemistry. Lung nodules from 23 of the 38 patients (60%) were Fas negative, those from 12 patients (32%) were weakly positive, and that from only 1 patient (3%) was strongly positive. Findings in the samples from the remaining two patients (5%) could not be interpreted because of extensive necrosis. Most patients with the weakly positive tumors and the single patient with the strongly positive tumor received chemotherapy prior to lung resection. There was a significant correlation between Fas expression and the administration of preoperative salvage chemotherapy (P = 0.0013). These data indicate that loss of Fas may be one mechanism by which OS cells evade host resistance in the lung. Chemotherapy may induce regression by upregulating Fas.', 'http://www.ncbi.nlm.nih.gov/pubmed/16282894', 'yes', 'yes', 'NA', 1);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(16327977, 'Uchibori M, Nishida Y, Nagasaka T, Yamada Y, Nakanishi K, Ishiguro N', 'Increased expression of membrane-type matrix metalloproteinase-1 is correlated with poor prognosis in patients with osteosarcoma.', 'Int J Oncol', '2006', 'Increased expression of membrane-type matrix metalloproteinase-1 is correlated with poor prognosis in patients with osteosarcoma. Numerous studies have demonstrated a correlation of matrix metalloproteinase (MMP) overexpression with the prognosis of various kinds of cancer. The current study investigated whether the expression of MMPs is correlated with the prognosis of osteosarcoma. Expression levels of MMP-2, -9, MT1-MMP and TIMP-2 were examined immunohistochemically in samples from 47 patients with osteosarcoma. Correlation of the positivity of staining with prognosis was analyzed with the Kaplan-Meier method, and statistically analyzed with log-rank test. Co-localization of MMP-2, MT1-MMP and TIMP-2 was determined by double staining with fluorescence-conjugated antibodies. Activities of gelatinases in representative tissues were examined with gelatin zymography. MMP-2 was expressed strongly in 60% of cases (28/47), and MMP-9, MT1- MMP and TIMP-2 was strongly positive in 61% (29/47), 45% (21/47), and 91% (43/47), respectively. Increased MT1-MMP expression was associated significantly with poor prognosis in overall survival (P=0.0480). In cases of overexpression for both MMP-2 and MT1-MMP, there was a tendency for poor prognosis (P=0.0969). In 36 cases who underwent neoadjuvant chemotherapy, wide resection of the tumors and post-operative adjuvant chemotherapy, increased expression of MT1-MMP resulted in a significant negative prognostic factor for disease-free survival (P=0.0143). Also, co-overexpression of MT1-MMP and MMP-2 showed a tendency to correlate to the reduced disease-free survival (P=0.0502). Increased gelatinase activity was observed in tissues of co-overexpression of MT1-MMP and MMP-2. The results of this study demonstrate the correlation of MT1-MMP expression and the oncological outcome of osteosarcoma patients, suggesting the prognostic significance of these proteins in osteosarcoma patients.', 'http://www.ncbi.nlm.nih.gov/pubmed/16327977', 'yes', 'yes', 'NA', 2),
(16327988, 'Mirandola P, Sponzilli I, Gobbi G, Marmiroli S, Rinaldi L, Binazzi R, Piccari GG, Ramazzotti G, Gaboardi GC, Cocco L, Vitale M', 'Anticancer agents sensitize osteosarcoma cells to TNF-related apoptosis-inducing ligand downmodulating IAP family proteins.', 'Int J Oncol', '2006', 'Anticancer agents sensitize osteosarcoma cells to TNF-related apoptosis-inducing ligand downmodulating IAP family proteins. Although TNF-related apoptosis-inducing ligand (TRAIL) usually induces cell death in tumor cells, there are some tumor cell types that are resistant to its apoptogenic effects. Some chemotherapeutic drugs, however, can sensitize resistant cancer cells to TRAIL by either upregulating surface TRAIL death receptor expression or by modulating intracellular signalling pathways emanating from TRAIL receptors. U2OS human osteosarcoma cells express TRAIL-R2 but are resistant to TRAIL-induced apoptosis. however, the genotoxic drugs, Doxorubicin and Cisplatin, are able to sensitize U2OS cells to TRAIL, without affecting their surface expression of either death or decoy TRAIL receptors. We demonstrate that Doxorubicin and Cisplatin downmodulate X-IAP, while not affecting FLIP levels in U2OS cells. Selective downmodulation of X-IAP protein synthesis by specific small interference RNA transfection induced a sensitization of U2OS cells to TRAIL comparable to that induced by pharmacological treatment with genotoxic drugs. TRAIL-R2 downmodulation by siRNAs completely abolished the TRAIL-induced apoptosis of genotoxin-treated U2OS cells. Our findings demonstrate that Doxorubicin and Cisplatin do not sensitize U2OS osteosarcoma cells to TRAIL by surface receptor modulation but rather by the removal of the intracellular signalling inhibition generated by X-IAP, suggesting a foreseeable relevant advantage to the therapy of these tumors by the combined regimen of genotoxin-based chemotherapy and TRAIL.', 'http://www.ncbi.nlm.nih.gov/pubmed/16327988', 'no', 'no', 'doxorubicin|cisplatin', 2),
(16328036, 'Roncuzzi L, Marti G, Baiocchi D, Del Coco R, Cocchi S, Gasperi-Campani A', 'Effect of Vinorelbine on cell growth and apoptosis induction in human osteosarcoma in vitro.', 'Oncol Rep', '2006', 'Effect of Vinorelbine on cell growth and apoptosis induction in human osteosarcoma in vitro. Vinorelbine (VNR) is a semi-synthetic vinca alkaloid known to exert its antitumour activity by interfering with the polymerisation of tubulin. It has shown a broad spectrum of activity in some advanced carcinomas of lung, breast and ovary. This report demonstrates for the first time the antiproliferative effect of VNR and its molecular mechanism in human osteosarcoma in vitro. TP53 wild-type HOS cells and TP53 mutated MG-63 cells were chosen for this study. In each cell line, VNR caused a significant dose- and time-dependent growth inhibition and induced apoptotic death independent of TP53 status. Phosphorylation and/or alteration of Bcl-2 were not induced by VNR, thereby indicating a new pathway utilised by the drug to induce apoptosis in this tumour in vitro. VNR produced a down-regulation of cyclin D1 and an up-regulation of p53 expression in TP53 wild-type HOS cells, whereas no alteration in cyclin D1 expression was evident in the TP53 negative MG-63 cells. These data suggest a new potential use for Vinorelbine as a therapeutic agent against human osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16328036', 'no', 'no', 'vinorelbine', 3),
(16354601, 'Kawakami T, Shimizu T, Kimura A, Hasegawa H, Siar CH, Ng KH, Nagatsuka H, Nagai N, Kanda H', 'Immunohistochemical examination of cytological differentiation in osteosarcomas.', 'Eur J Med Res', '2005', 'Immunohistochemical examination of cytological differentiation in osteosarcomas. In this immunohistochemical examination, the expression of Runx2, Notch1, Delta and Osteopontin peptides were detected in neoplastic cells in 10 Japanese cases of osteosarcoma. Immunohistochemically, Runx2 peptide expression appeared in the cytoplasm of almost all neoplastic cells of the 10 cases examined. However, Notch1 peptide expression appeared in the cytoplasm of neoplastic cells in the localized and comparatively well-differentiated area of osteosarcoma, which osteoblastic and chondroblastic containing osteoid and/or chondroid tissues. No expression of Notch1 peptide was detected in the fibroblastic and poorly differentiated areas. Delta peptide appearance was nearly the same pattern of Notch1 peptide. Expression of Osteopontin peptide appeared in almost all cells and the strength expression was shown in the area of comparatively well-differentiated tissues. Therefore, these results suggest that Runx2, Notch1, and Delta peptides are closely related to cytological differentiation or acquisition of tissue specific characteristics in neoplastic cells in osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/16354601', 'yes', 'no', 'NA', 2),
(16369996, 'Bransfield KL, Askham JM, Leek JP, Robinson PA, Mighell AJ', 'Phenotypic changes associated with DYNACTIN-2 (DCTN2) over expression characterise SJSA-1 osteosarcoma cells.', 'Mol Carcinog', '2006', 'Phenotypic changes associated with DYNACTIN-2 (DCTN2) over expression characterise SJSA-1 osteosarcoma cells. DYNACTIN-2 (DCTN2) localises to chromosome 12q13-q15, a region prone to stable amplification in several cancers. Transient DCTN2 overexpression has a significant impact on cellular phenotype primarily due to disruption of the DYNEIN-dynactin motor. Changes reported include alterations of microtubule-directed movement of molecular (e.g. TP53) and organelle (e.g. Golgi) cargoes towards the nucleus, centrosome biology, cellular movement and mitosis with a potential predisposition to mitotic block and polyploidy. These changes would be expected to be of relevance to carcinogenesis. To investigate this, we report the first study of DCTN2 genomic amplification and sustained DCTN2 overexpression in cancer cells. QFMPCR was employed to characterise the extent of chromosome 12q13-q15 amplicons in SJSA-1, SJRH30, U373MG and CCF-STTG1 cancer cells. DCTN2 amplification was present in SJSA-1, U373MG and SJRH30 cells, yet was incomplete at the 5''-end in SJRH30 cells. Only SJSA-1 cells were characterised by DCTN2 overexpression on Western blot analyses. Microscopy studies distinguished SJSA-1 cells by greater DCTN2 immunofluorescence and diminished centrosome and 58K protein Golgi-marker focus compared to SJRH30 cells. Indirect evidence derived from the published work of others indicated that TP53 transport into the nucleus was unimpaired. Furthermore, we observed that SJSA-1 cells were easy to propagate. In conclusion, persistent DCTN2 overexpression can be tolerated in SJSA-1 cancer cells despite phenotypic abnormalities predicted from transient overexpression studies. This preliminary study does not support a major role for DCTN2 overexpression in carcinogenesis, although further studies would be necessary to confirm this.', 'http://www.ncbi.nlm.nih.gov/pubmed/16369996', 'no', 'no', 'NA', 2),
(16380506, 'Zhou Z, Lafleur EA, Koshkina NV, Worth LL, Lester MS, Kleinerman ES', 'Interleukin-12 up-regulates Fas expression in human osteosarcoma and Ewing''s sarcoma cells by enhancing its promoter activity.', 'Mol Cancer Res', '2005', 'Interleukin-12 up-regulates Fas expression in human osteosarcoma and Ewing''s sarcoma cells by enhancing its promoter activity. Interleukin-12 (IL-12) has shown significant antitumor activity in several preclinical animal tumor models. Our previous studies showed that IL-12 inhibited tumor growth in human osteosarcoma and Ewing''s sarcoma animal model. Decreased Fas expression in osteosarcoma increased the lung metastatic potential. In this study, we further examined the mechanism of IL-12 antitumor activity and showed that IL-12 significantly increased Fas expression in both human osteosarcoma cells LM7 and Ewing''s sarcoma cells TC71. Up-regulation of Fas expression increased their sensitivity to Fas-induced cell apoptosis. Constructs of the Fas promoter linked to a luciferase reporter gene were used to determine the promoter activity. IL-12 increased Fas promoter activity 4.2- and 4.9-fold in TC71 and LM7 cells, respectively. Time course studies have shown that recombinant IL-12 stimulated Fas promoter activity at 2 hours, reached the peak level at 4 hours, and then declined at 24 hours. To investigate whether IL-12 specifically enhanced Fas promoter activity, we determined whether another gene (E1A) was able to stimulate Fas promoter activity. We also evaluated effect of IL-12 on the topoisomerase IIalpha promoter. The results indicated that E1A but not IL-12 stimulated topoisomerase IIalpha promoter activity. E1A failed to increase Fas promoter activity. We also found that kappaB-Sp1 element at position -295 to -286 in Fas promoter was essential for IL-12-induced activation, and nuclear factor-kappaB transcription factor was activated after IL-12 treatment in TC71 cells. These results indicate that IL-12 up-regulates Fas expression in human osteosarcoma and Ewing''s sarcoma by enhancing Fas promoter activity. Understanding this mechanism may lead to new therapeutic approaches for the treatment of sarcoma involving the use of IL-12.', 'http://www.ncbi.nlm.nih.gov/pubmed/16380506', 'no', 'no', 'NA', 2),
(16394764, 'Oh JH, Kim HS, Kim HH, Kim WH, Lee SH', 'Aberrant methylation of p14ARF gene correlates with poor survival in osteosarcoma.', 'Clin Orthop Relat Res', '2006', 'Aberrant methylation of p14ARF gene correlates with poor survival in osteosarcoma. We studied the methylation status of the CpG promoter regions of the p16 and p14 genes, mutations of four exons of the CDKN2A gene, and expressions of their corresponding proteins. Thirty-two frozen osteosarcoma tissues were used for methylation-specific polymerase chain reaction and sequence analysis. Immunohistochemical staining for p16 and p14 proteins was done. The histologic and clinical data were analyzed to find their prognostic implications. The promoter of p16 gene was methylated in 16%, and p14 in 47%. Poor survival was related to methylation of p14. The methylation of p14 correlated with down-regulation of its protein expression. The methylation of p14 showed the highest hazard ratio by multivariate survival analysis. Our data suggest methylation of the CDKN2A gene is the main mechanism of its protein repression. For the p14 gene, methylation of the promoter region was related to repression of p14 protein and poor prognosis. Level of Evidence: Prognostic study, Level II (retrospective study). See the Guidelines for Authors for a complete description of levels of evidence.', 'http://www.ncbi.nlm.nih.gov/pubmed/16394764', 'yes', 'yes', 'NA', 1),
(16417466, 'Felx M, Guyot MC, Isler M, Turcotte RE, Doyon J, Khatib AM, Leclerc S, Moreau A, Moldovan F', 'Endothelin-1 (ET-1) promotes MMP-2 and MMP-9 induction involving the transcription factor NF-kappaB in human osteosarcoma.', 'Clin Sci (Lond)', '2006', 'Endothelin-1 (ET-1) promotes MMP-2 and MMP-9 induction involving the transcription factor NF-kappaB in human osteosarcoma. In the present study, we have investigated the effect of (i) ET-1 (endothelin-1) and its precursor, big ET-1, on MMP (matrix metalloproteinase)-2 and MMP-9 synthesis and activity in osteosarcoma tissue, and (ii) ET-1 receptor antagonists on cell invasion. Using Western blotting, zymography, RT-PCR (reverse transcription-PCR), immunohistochemistry, immunofluorescence and Northern blotting, we have shown that ET-1 and ET-1 receptors (ET(A) and ET(B)) were expressed in these cells. Additionally, we have demonstrated that ET-1 markedly induced the synthesis and activity of MMP-2, which was significantly increased when compared with MMP-9. Furthermore, inhibition of NF-kappaB (nuclear factor kappaB) activation blocked MMP-2 production and activity, indicating the involvement of NF-kappaB, a ubiquitous transcription factor playing a central role in the differentiation, proliferation and malignant transformation. Since ET-1 acts as an autocrine mediator through gelatinase induction and because inhibition of ET(A) receptor is beneficial for reducing both basal and ET-1-induced osteosarcoma cell invasion, targeting this receptor could be an attractive therapeutic alternative for the successful treatment of osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16417466', 'no', 'no', 'NA', 6),
(16421247, 'Manara MC, Bernard G, Lollini PL, Nanni P, Zuntini M, Landuzzi L, Benini S, Lattanzi G, Sciandra M, Serra M, Colombo MP, Bernard A, Picci P, Scotlandi K', 'CD99 acts as an oncosuppressor in osteosarcoma.', 'Mol Biol Cell', '2006', 'CD99 acts as an oncosuppressor in osteosarcoma. CD99 was recently reported to be under control of the osteoblast-specific transcription factor Cbfa1 (RUNX2) in osteoblasts, suggesting a role in the phato-physiology of these cells. No extensive information is available on the role(s) of this molecule in malignant phenotype, and osteosarcoma, in particular, has never been studied. We report that in 11 different cell lines and 17 clinical samples CD99 expression is either undetectable or very low. Being expressed in the normal counterpart, we tested the hypothesis that CD99 down-regulation may have a role in osteosarcoma development and progression. CD99-forced expression in two osteosarcoma cell lines significantly reduced resistance to anoikis, inhibited growth in anchorage independence as well as cell migration, and led to abrogation of tumorigenic and metastatic ability. Therefore, the molecule acts as a potent suppressor of malignancy in osteosarcoma. CD99 gene transfection induces caveolin-1 up-regulation and the two molecules were found to colocalize on the cell surface. Treatment with antisense oligonucleotides to caveolin-1 abrogates the effects of CD99 on migration. The findings point to an antioncogenic role for CD99 in osteosarcoma, likely through the regulation of caveolin-1 and inhibition of c-Src kinase activity.', 'http://www.ncbi.nlm.nih.gov/pubmed/16421247', 'no', 'no', 'NA', 3),
(16437349, 'Srivastava A, Rock C, Zhang K, Ruan M, Bolander ME, Sarkar G', 'Expression of a novel alternatively spliced UCP-2 transcript in osteogenic sarcoma.', 'J Orthop Sci', '2006', 'Expression of a novel alternatively spliced UCP-2 transcript in osteogenic sarcoma. Development of chemoresistance is common in patients with osteogenic sarcoma (OGS); however, the underlying mechanism is largely unknown. Many anticancer drugs exert their therapeutic action by generating reactive oxygen radicals, which might be countered by the cancer cell through induction of uncoupling protein 2 (UCP-2). UCP-2 has been shown to be able to protect tumor cells from the cytotoxic actions of chemotherapeutic drugs. Because OGS is seldom completely cured by current chemotherapy regimens, we hypothesized that increased expression of UCP-2 underlies this phenomenon. The primary initial interest of our research was to evaluate the level of UCP-2 mRNA in OGS.The level of UCP-2 mRNA was determined by reverse transcriptase polymerase chain reaction (RT-PCR) comparing expression in normal-bone-derived specimens and OGS-derived specimens. Semiquantification of mRNA expression was achieved by radioactive RT-PCR. Nucleotide sequencing was performed using automated instruments.Interestingly, we failed to observe induction of UCP-2 mRNA in OGS tumor specimens and OGS-derived primary cell lines compared to the expression level in normal bone. However, we found expression of a hitherto unknown UCP-2 transcript in eight of eight OGS-derived and one EWS-derived cell lines and in nine of ten OGS biopsy specimens but in only one of six normal bone-derived specimens. Thus, tumor samples express both types (normal and the novel one) of UCP-2 mRNAs, whereas normal bone expresses only the wild-type form. Further experiments identified the novel mRNA species as an alternatively spliced UCP-2 transcript (termed UCP-2as). UCP-2as has a 22-nucleotide insertion from the 3'' end of intron 3 that introduces an early stop codon in exon 4, which theoretically can produce a protein 79 amino acids long.We have identified a hitherto unknown UCP-2 transcript. Expression of the novel transcript appears to be OGS-specific, implying a function advantageous to the tumor.', 'http://www.ncbi.nlm.nih.gov/pubmed/16437349', 'yes', 'no', 'NA', 1),
(16466985, 'Xie XK, Yang DS, Ye ZM, Tao HM', 'Recombinant antisense C-myc adenovirus increase in vitro sensitivity of osteosarcoma MG-63 cells to cisplatin.', 'Cancer Invest', '2006', 'Recombinant antisense C-myc adenovirus increase in vitro sensitivity of osteosarcoma MG-63 cells to cisplatin. C-myc is an oncogene with the important role of cell proliferation controller. It has been found to be amplified and overexpressed in osteosarcoma. Moreover, it can promote cell transformation and induce metastatic features. Some studies showed that overexpression of c-myc could induce resistance in response to antineoplastic agents. Currently, we constructed the recombinant adenovirus (Ad-Asc-myc) encoding antisense c-myc fragment and investigated its effect on the in vitro sensitivity of osteosarcoma MG-63 cells to cisplatin(CDDP). The osteosarcoma MG-63 cells were transfected by the Ad-Asc-myc in vitro, and Western Blot, MTT assay, RT-PCR, flow cytometry (FCM), and transmission electron microscopy (TEM) were used to study expression of c-myc and caspase-3 protein, tumor cell proliferation in vitro, cell apoptotic morphology and cell cycle change. Ad-Asc-myc encoding antisense c-myc fragment was obtained with the titer of 2.0 x 10(9) pfu/ml. Ad-Asc-myc downregulated the expression of c-myc protein after transfected MG-63 cells for 48 hours, combined with the treatment of 2.0, 5.0 microg/ml cisplatin for 2 hours can inhibited tumor cells proliferation in vitro by 33.4 and 54.2 percent, respectively, which had significant difference compared with control recombinant adenovirus (Ad-LacZ) groups (P < 0.05). RT-PCR revealed that Ad-Asc-myc downregulated expression of bcl-2 and upregulated expression of Bax, and no appreciable changes were observed in the expression of E2F-1. Detection of caspase-3 protein TEM, and FCM analysis showed that Ad-Asc-myc could induce apoptosis of transfected cells, which was enhanced by the treatment of cisplatin. Cell cycle analysis showed that obvious G(2)/M phase arrested in transfected cells. In conclusion, Ad-Asc-myc increased the in vitro sensitivity of osteosarcoma MG-63 cells to cisplatin as well as induced apoptosis.', 'http://www.ncbi.nlm.nih.gov/pubmed/16466985', 'no', 'no', 'cisplatin', 4),
(16470222, 'Fromigué O, Haÿ E, Modrowski D, Bouvet S, Jacquel A, Auberger P, Marie PJ', 'RhoA GTPase inactivation by statins induces osteosarcoma cell apoptosis by inhibiting p42/p44-MAPKs-Bcl-2 signaling independently of BMP-2 and cell differentiation.', 'Cell Death Differ', '2006', 'RhoA GTPase inactivation by statins induces osteosarcoma cell apoptosis by inhibiting p42/p44-MAPKs-Bcl-2 signaling independently of BMP-2 and cell differentiation. Osteosarcoma is the most common primary bone tumour in young adults. Despite improved prognosis, resistance to chemotherapy remains responsible for failure of osteosarcoma treatment. The identification of signals that promote apoptosis may provide clues to develop new therapeutic strategies for chemoresistant osteosarcoma. Here, we show that lipophilic statins (atorvastatin, simvastatin, cerivastatin) markedly induce caspases-dependent apoptosis in various human osteosarcoma cells, independently of bone morphogenetic protein (BMP)-2 signaling and cell differentiation. Although statins increased BMP-2 expression, the proapoptotic effect of statins was not prevented by the BMP antagonist noggin, and was abolished by mevalonate and geranylgeranylpyrophosphate, suggesting the involvement of defective protein geranylgeranylation. Consistently, lipophilic statins induced membrane RhoA relocalization to the cytosol and inhibited RhoA activity, which resulted in decreased phospho-p42/p44- mitogen-activated protein kinases (MAPKs) and Bcl-2 levels. Constitutively active RhoA rescued phospho-p42/p44-MAPKs and Bcl-2 and abolished statin-induced apoptosis. Thus, lipophilic statins induce caspase-dependent osteosarcoma cell apoptosis by a RhoA-p42/p44 MAPKs-Bcl-2-mediated mechanism, independently of BMP-2 signaling and cell differentiation.', 'http://www.ncbi.nlm.nih.gov/pubmed/16470222', 'no', 'no', 'atorvastatin|simvastatin|cerivastatin', 6),
(16475708, 'Tenta R, Sourla A, Lembessis P, Koutsilieris M', 'Bone-related growth factors and zoledronic acid regulate the PTHrP/PTH.1 receptor bioregulation systems in MG-63 human osteosarcoma cells.', 'Anticancer Res', 'NA', 'Bone-related growth factors and zoledronic acid regulate the PTHrP/PTH.1 receptor bioregulation systems in MG-63 human osteosarcoma cells. Bisphosphonates are known to inhibit osteoclast-mediated bone resorption and osteoblast differentiation and are currently used in the treatment of Paget''s disease, osteoporosis, metastatic and osteolytic bone disease and hypercalcaemia of malignancy. The parathyroid hormone-related peptide (PTHrP) and type 1 PTH/PTHrP receptor (PTH.1R) bioregulation systems mediate a wide range of local paracrine/autocrine and intracrine functions in various tissues and modify the actions of pharmaceutical agents on target tissues, both in vivo and in vitro. In addition, bone microenvironment-related growth factors, such as insulin-like growth factor 1 (IGF-1), transforming growth factor beta 1 (TGF beta 1), basic fibroblast growth factor (bFGF) and interleukin 6 (IL-6), can modify the actions of various pharmaceutical agents, including cytotoxic drugs in malignant cell lines. Whether IGF-1, TGF beta 1, bFGF, IL-6 and zoledronic acid affect the expressions of PTHrP and PTH.1R in MG-63 osteoblast-like osteosarcoma cells was investigated in this study. Relative quantitative-PCR (expression at mRNA level) and immunofluorescence analysis (localization of the expression at protein level) were employed to assess PTHrP and PTH.IR expressions. Our data showed that primarily IGF-1, TGF beta 1 and IL-6 (up to 25 ng/ml for 48 h) increased PTHrP mRNA expression and modified its perinuclear localization, while zoledronic acid (up to 100 microM for 48 h) inhibited cell proliferation and suppressed PTHrP expression in the MG-63 osteosarcoma cells. These growth factors were incapable of reversing the zoledronic acid decrease of the expression of PTHrP in the MG-63 cells, suggesting that zoledronic acid and the growth factors affect PTHrP expression via an independent intracellular signal transduction pathway in these cells. However, no appreciable modulation of the PTH.1R expression by IGF-1, TGF beta 1, bFGF, IL-6 or zoledronic acid was detected in MG-63 cells. Therefore, we conclude that PTHrP expression possibly mediates the action of bone microenvironment-related growth factors and of zoledronic acid in MG-63 cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16475708', 'no', 'no', 'zoledronic acid', 5),
(16502411, 'Hou P, Ji M, Yang B, Chen Z, Qiu J, Shi X, Lu Z', 'Quantitative analysis of promoter hypermethylation in multiple genes in osteosarcoma.', 'Cancer', '2006', 'Quantitative analysis of promoter hypermethylation in multiple genes in osteosarcoma. Osteosarcoma is the most common solid malignant diseases of childhood, occurring in approximately 6 children per million annually; however, to the authors'' knowledge to date, the cause of osteosarcoma has remained mostly unknown. Genetic alterations of genes that are specific for osteosarcoma have not been identified. Genetic alternations in the status of DNA methylation, known as epigenetic alterations, are the most common molecular alterations in human neoplasia. Aberrant methylation in the promoter region of tumor-related genes is associated closely with epigenetically mediated gene silencing, which is a common feature in human tumors.The authors analyzed CpG islands of 5 different gene loci for aberrant methylation profiles in 30 pairs of osteosarcoma and corresponding normal tissues by using the quantitative methylation-specific polymerase chain reaction method. The objectives of this study were to characterize the methylation changes in osteosarcoma more extensively and to identify epigenetic biomarkers that may be useful in the diagnosis and prevention of osteosarcoma.For the Ras effector homologue (RASSF1A), tissue inhibitor of metalloproteinase 3 (TIMP3), O-6-methylguanine DNA methyltransferase (MGMT), and death-associated protein kinase 1 (DAPK1) genes, significant differences were observed in the degree of hypermethylation between tumors and normal tissues (P < 0.01 and P < 0.001, respectively). Measurement of the cumulative multiple promoter hypermethylation revealed striking differences between tumor specimens and normal tissues (t = 7.31; P < .001). There also was a significant difference in the levels of DNA methylation between the metastatic and nonmetastatic high-grade osteosarcomas (t = 4.57; P < .01). In addition, the methylation levels were associated closely with gender (t = 6.44; P < .001).The results indicated that tumor tissues from patients with osteosarcoma had a significantly higher incidence of hypermethylation for several genes compared with corresponding normal tissues. The epigenetic changes observed in this study may have prognostic importance for patients with osteosarcoma.Copyright 2006 American Cancer Society.', 'http://www.ncbi.nlm.nih.gov/pubmed/16502411', 'yes', 'yes', 'NA', 4),
(16528367, 'Oda Y, Yamamoto H, Tamiya S, Matsuda S, Tanaka K, Yokoyama R, Iwamoto Y, Tsuneyoshi M', 'CXCR4 and VEGF expression in the primary site and the metastatic site of human osteosarcoma: analysis within a group of patients, all of whom developed lung metastasis.', 'Mod Pathol', '2006', 'CXCR4 and VEGF expression in the primary site and the metastatic site of human osteosarcoma: analysis within a group of patients, all of whom developed lung metastasis. The chemokine, CXCL12, and its receptor, CXCR4, have recently been shown to play an important role in metastasis of several kinds of carcinoma. It has also been demonstrated that VEGF regulates both the expression of CXCR4 and invasiveness in breast cancer cell lines. We compared the immunohistochemical expression of CXCR4 and VEGF between the primary site and a concordant pulmonary metastatic site in 30 osteosarcoma patients, all of which had undergone thoracotomy. Microvessel density (MVD) as shown by immunostaining of CD34 and proliferative activity with MIB-1 monoclonal antibody was also evaluated. CXCR4 expression (primary, 33.3% positive vs metastatic, 66.6% positive; P = 0.0097) and MVD (primary, 29.86 +/- 6.87/0.26 mm2 vs metastatic, 43.32 +/- 8.65/0.26 mm2; P = 0.0015) in the metastatic site were both significantly increased compared with those in the primary site, whereas no difference between primary and metastatic sites was observed with regard to VEGF expression. There was a significant positive correlation between immunohistochemical CXCR4 and VEGF expression (P = 0.0269). In total population, the MIB-1-labeling index (LI) was significantly higher in tumors, which showed immunoreactivity for VEGF (MIB-1-LI in VEGF-positive tumors, 24.29 +/- 5.4 vs VEGF-negative tumors, 18.33 +/- 4.16; P = 0.034). Furthermore, those patients with VEGF-positive primary tumors had a significantly worse prognosis compared with the patients with VEGF-negative primary tumors (P = 0.0053). Our results suggested that CXCR4 expression was associated with metastatic progression, and immunohistochemical VEGF expression in the primary site had predictive value for the osteosarcoma patients, who developed lung metastasis.', 'http://www.ncbi.nlm.nih.gov/pubmed/16528367', 'yes', 'yes', 'NA', 2),
(16533764, 'Nadiminty N, Lou W, Lee SO, Mehraein-Ghomi F, Kirk JS, Conroy JM, Zhang H, Gao AC', 'Prostate-specific antigen modulates genes involved in bone remodeling and induces osteoblast differentiation of human osteosarcoma cell line SaOS-2.', 'Clin Cancer Res', '2006', 'Prostate-specific antigen modulates genes involved in bone remodeling and induces osteoblast differentiation of human osteosarcoma cell line SaOS-2. The high prevalence of osteoblastic bone metastases in prostate cancer involves the production of osteoblast-stimulating factors by prostate cancer cells. Prostate-specific antigen (PSA) is a serine protease uniquely produced by prostate cancer cells and is an important serologic marker for prostate cancer. In this study, we examined the role of PSA in the induction of osteoblast differentiation.Human cDNA containing a coding region for PSA was transfected into human osteosarcoma SaOS-2 cells. SaOS-2 cells were also treated with exogenously added PSA. We evaluated changes in global gene expression using cDNA arrays and Northern blot analysis resulting from expression of PSA in human osteosarcoma SaOS-2 cells.SaOS-2 cells expressing PSA had markedly up-regulated expression of genes associated with osteoblast differentiation including runx-2 and osteocalcin compared with the controls. Consistent with these results, the stable clones expressing PSA showed increased mineralization and increased activity of alkaline phosphatase in vitro compared with controls, suggesting that these cells undergo osteoblast differentiation. We also found that osteoprotegerin expression was down-regulated and that the receptor activator of NF-kappaB ligand expression was up-regulated in cells expressing PSA compared with controls.Modulation of the expression of osteogenic genes and alteration of the balance between osteoprotegerin-receptor activator of NF-kappaB ligand by PSA suggests that PSA produced by metastatic prostate cancer cells may participate in bone remodeling in favor of the development of osteoblastic metastases in the heterogeneous mixture of osteolytic and osteoblastic lesions. These findings provide a molecular basis for understanding the high prevalence of osteoblastic bone metastases in prostate cancer.', 'http://www.ncbi.nlm.nih.gov/pubmed/16533764', 'no', 'no', 'NA', 6),
(16540432, 'Nikitovic D, Zafiropoulos A, Katonis P, Tsatsakis A, Theocharis AD, Karamanos NK, Tzanakakis GN', 'Transforming growth factor-beta as a key molecule triggering the expression of versican isoforms v0 and v1, hyaluronan synthase-2 and synthesis of hyaluronan in malignant osteosarcoma cells.', 'IUBMB Life', '2006', 'Transforming growth factor-beta as a key molecule triggering the expression of versican isoforms v0 and v1, hyaluronan synthase-2 and synthesis of hyaluronan in malignant osteosarcoma cells. Versican, a large sized chondroitin-sulphate proteoglycan (PG), and its binding partner, hyaluronan (HA), are extracellular matrix (ECM) components that play an essential role in transformed cell behavior. Expression of certain versican isoforms has been implicated in cell migration and proliferation of cancer cells and, on the other hand, disruption of HA synthesis by inhibiting hyaluronan synthase-2 (HAS2) expression in osteosarcoma cells by suppressing cell proliferation, invasiveness and motility. Considering that growth factors, such as TGF-beta, bFGF and PDGF-BB, are important regulators for the expression of the ECM macromolecules, in this study we examined the effect of these growth factors on the expression of the various versican isoforms, HA synthases as well as HA synthesis by MG-63 osteosarcoma cells and normal human osteoblastic periodontal ligament cells (hPDL). Real-time PCR and metabolic labelling followed by fine HPLC analysis coupled to radiochemical detection were the methods utilized. It was found that, contrary to normal hPDL cells, osteosarcoma MG-63 cells do not constitutively express the versican isoforms V0 and V1. Exogenous addition of TGF-beta2 stimulated the versican transcript levels mainly by forcing osteosarcoma cells to express V1 and V0 isoforms. PDGF-BB and bFGF had only minor effects in these cells. In hPDL cells a strong stimulation of the V3 transcript by all growth factors was observed. TGF-beta2 was also the major stimulator of HAS2 isoform expression as well as hyaluronan synthesis in osteosarcoma cells, while PDGF-BB exerted dominant influence on HAS2 isoform expression and hyaluronan biosynthesis by osteoblasts. The obtained results show for the first time that TGF-beta2 triggers the malignant phenotype pattern of versican and hyaluronan expression in human osteosarcoma cells and indicate that this growth factor may account for the metastatic potential of these cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16540432', 'no', 'no', 'NA', 3),
(16547468, 'Nakatsuka S, Oji Y, Horiuchi T, Kanda T, Kitagawa M, Takeuchi T, Kawano K, Kuwae Y, Yamauchi A, Okumura M, Kitamura Y, Oka Y, Kawase I, Sugiyama H, Aozasa K', 'Immunohistochemical detection of WT1 protein in a variety of cancer cells.', 'Mod Pathol', '2006', 'Immunohistochemical detection of WT1 protein in a variety of cancer cells. WT1 was first identified as a tumor suppressor involved in the development of Wilms'' tumor. Recently, oncogenic properties of WT1 have been demonstrated in various hematological malignancies and solid tumors. Because WT1 has been identified as a molecular target for cancer immunotherapy, immunohistochemical detection of WT1 in tumor cells has become an essential part of routine practice. In the present study, the expression of WT1 was examined in 494 cases of human cancers, including tumors of the gastrointestinal and pancreatobiliary system, urinary tract, male and female genital organs, breast, lung, brain, skin, soft tissues and bone by immunohistochemistry using polyclonal (C-19) and monoclonal (6F-H2) antibodies against WT1 protein. Staining for C-19 and 6F-H2 was found in 35-100 and 5-88% of the cases of each kind of tumor, respectively. WT1-positive tumors included tumor of the stomach, prostate, and biliary and urinary systems, and malignant melanomas. A majority of the positive cases showed diffuse or granular staining in the cytoplasm, whereas ovarian tumors and desmoplastic small round cell tumors frequently showed nuclear staining. Glioblastomas, some of soft tissue sarcomas, osteosarcomas, and malignant melanomas of the skin showed extremely strong cytoplasmic staining as compared with other tumors. Western blot analysis showed that WT1 protein was predominantly expressed in the cytoplasm of the tumor cells in two cases of lung adenocarcinoma, supporting the intracytoplasmic staining for WT1 using immunohistochemistry. Immunohistochemical detection with routinely processed histologic sections could provide meaningful information on the expression of WT1 in cancer cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16547468', 'yes', 'no', 'NA', 1),
(16571651, 'Matsui TA, Sowa Y, Yoshida T, Murata H, Horinaka M, Wakada M, Nakanishi R, Sakabe T, Kubo T, Sakai T', 'Sulforaphane enhances TRAIL-induced apoptosis through the induction of DR5 expression in human osteosarcoma cells.', 'Carcinogenesis', '2006', 'Sulforaphane enhances TRAIL-induced apoptosis through the induction of DR5 expression in human osteosarcoma cells. Sulforaphane (SFN), a naturally occurring isothiocyanate, is an attractive agent because of its potent anticancer effects. SFN suppresses the proliferation of various cancer cells in vitro and in vivo. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is also one of the most promising candidates for cancer therapeutics owing to its ability to selectively induce apoptosis in tumor cells. In this study, we report that SFN enhances TRAIL-induced apoptosis in human osteosarcoma cells, Saos2 and MG63. The apoptosis induced by co-treatment with SFN and TRAIL was markedly blocked by a dominant negative form of the TRAIL receptor or caspase inhibitors. The combined use of SFN and TRAIL effectively induced Bid cleavage and the activation of caspases 8, 10, 9 and 3 at ineffective concentrations for each agent. SFN upregulated the expression of death receptor 5 (DR5), a receptor for TRAIL, at mRNA and protein levels in a dose-dependent manner. In addition, the SFN-mediated sensitization to TRAIL was reduced by DR5 siRNA, suggesting that the sensitization was at least partially mediated through the induction of DR5 expression. Furthermore, SFN sensitized TRAIL-induced apoptosis in a p53-independent manner. On the other hand, SFN neither induced DR5 protein expression or enhanced TRAIL-induced apoptosis in normal human peripheral blood mononuclear cells. Thus, combined treatment with SFN and TRAIL might be a promising therapy for osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16571651', 'no', 'no', 'sulforaphane', 6),
(16596382, 'Nishio J, Gentry JD, Neff JR, Nelson M, Daniels W, Perry D, Gatalica Z, Bridge JA', 'Monoallelic deletion of the p53 gene through chromosomal translocation in a small cell osteosarcoma.', 'Virchows Arch', '2006', 'Monoallelic deletion of the p53 gene through chromosomal translocation in a small cell osteosarcoma. Small cell osteosarcoma is a rare bone tumor of high-grade malignancy that most often arises in the metaphysis of long bones in the second decade of life. Cytogenetic and molecular genetic findings in small cell osteosarcoma are poorly defined. Conventional cytogenetic analysis of a small cell osteosarcoma arising in the proximal tibia of a 9-year-old male revealed a diploid chromosomal complement with complex structural rearrangements involving chromosomes 6, 16, and 17. Immunohistochemical assessment of p53 protein expression revealed nuclear p53 immunoreactivity in approximately 15% of the neoplastic cells. Subsequent fluorescence in situ hybridization (FISH) analyses confirmed loss of the p53 gene locus on the derivative chromosome 17 homolog and were negative for amplification of the MDM2, CDK4, c-MYC, HER-2/neu, CCND1, and COPS3 gene loci. To the best of our knowledge, this represents the first demonstration of monoallelic deletion of p53 in small cell osteosarcoma, suggesting that p53 alterations may play an important role in the development of small cell osteosarcoma as well as conventional osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16596382', 'yes', 'no', 'NA', 7),
(16677779, 'Park HR, Jung WW, Bacchini P, Bertoni F, Kim YW, Park YK', 'Ezrin in osteosarcoma: comparison between conventional high-grade and central low-grade osteosarcoma.', 'Pathol Res Pract', '2006', 'Ezrin in osteosarcoma: comparison between conventional high-grade and central low-grade osteosarcoma. Ezrin is a cytoskeleton linker protein that is actively involved in the regulation of growth and metastatic capacity of cancer cells. Recently, it has been demonstrated that a significant correlation exists between high ezrin expression levels and the poor outcome of pediatric osteosarcoma patients. The expression of ezrin was compared in conventional high-grade and central low-grade osteosarcoma lesions to investigate the role of ezrin overexpression in the metastasis of osteosarcoma. We compared the expression levels of the ezrin protein in 32 cases of high-grade osteosarcomas and 21 cases of low-grade osteosarcomas using immunohistochemistry. Ezrin protein expression levels were examined in three different human osteosarcoma cell lines by Western blotting. In addition, the mRNA expression levels of ezrin in these osteosarcoma cell lines and control fibroblasts were evaluated by real-time quantitative PCR. Ezrin immunoreactivity was present in 43.7% of high-grade osteosarcoma specimens. All low-grade osteosarcomas were negative for ezrin. The expression of ezrin was detected by Western blotting in all three osteosarcoma cell lines. The tested osteosarcoma cell lines showed marked amplification of ezrin mRNA compared to control cells. Taken together, ezrin appears to play a role in the progression of tumors, such as the metastasis of osteosarcoma. However, further data are needed before ezrin can be considered in clinical decision-making about osteosarcoma patients.', 'http://www.ncbi.nlm.nih.gov/pubmed/16677779', 'yes', 'yes', 'NA', 1),
(16683887, 'Willmore-Payne C, Holden JA, Zhou H, Gupta D, Hirschowitz S, Wittwer CT, Layfield LJ', 'Evaluation of Her-2/neu gene status in osteosarcoma by fluorescence in situ hybridization and multiplex and monoplex polymerase chain reactions.', 'Arch Pathol Lab Med', '2006', 'Evaluation of Her-2/neu gene status in osteosarcoma by fluorescence in situ hybridization and multiplex and monoplex polymerase chain reactions. Previous reports suggest that the human epidermal growth factor 2 (HER-2/neu) receptor may be overexpressed in osteosarcoma.To determine whether osteosarcomas have amplifications of the HER-2/neu gene.We studied a series of osteosarcomas by fluorescence in situ hybridization (FISH) and by 2 real-time polymerase chain reaction assays that measure the amount of HER-2/neu DNA relative to a control gene. The HER-2/ neu monoplex and multiplex assays were capable of identifying those cases of breast cancer that were known to overexpress HER-2/neu as assessed by FISH. We initially studied 21 cases of osteosarcoma by FISH analysis (using a technique that included a probe for chromosome 17), 11 of which had their HER-2/neu gene amplification status previously reported.None of these osteosarcoma cases showed HER-2/neu amplification by our FISH analysis and subsequent quantitative (multiplex) polymerase chain reaction. Apparent expression of HER-2/neu protein was observed in several of the cases but the immunoreactivity was localized to the cytoplasm and was not membranous in character. An additional 35 osteosarcoma specimens were subjected to monoplex polymerase chain reaction analysis, and amplifiable DNA was recovered from 19 specimens (54%). None of these samples had HER-2/neu amplification by monoplex PCR analysis and only one case had membranous immunoreactivity graded as 1+.Although a small subset of osteosarcomas had weak noncircumferential membranous immunoreactivity for HER-2/neu protein, no osteosarcomas demonstrated positive (2+ or 3+) immunoreactivity for HER-2/ neu protein and none showed HER-2/neu gene amplification by either FISH or polymerase chain reaction.', 'http://www.ncbi.nlm.nih.gov/pubmed/16683887', 'yes', 'yes', 'NA', 1),
(16696324, 'Liao X, Yang S, Shao Z, Li J, Liu Y, Xiong X, Liu X', 'Effect of exogenous p16ink4a and hRb1 genes on cell cycle regulation of osteosarcoma cell.', 'J Huazhong Univ Sci Technolog Med Sci', '2005', 'Effect of exogenous p16ink4a and hRb1 genes on cell cycle regulation of osteosarcoma cell. To study the effect on regulation of cell cycle of osteosarcoma cell line MG63 tranceduced with exogenous p16ink4a and hRb1 genes, pIRES-p16ink4a-hRb1, pIRES-p16ink4a and pIRES-hRb1 plasmids were constructed by gene recombination technology. The recombinant plasmid was transferred into osteosarcoma cell line MG63 by metafectene, and the resistant clones were selected by G418 selective medium. mRNA and protein expression of osteosarcoma cell line were assayed by RT-PCR and Western-Blot respectively. Cell cycle and apoptosis were analyzed by subG1 flow cytometric. Cell proliferation was tested by MTT. In the genome of these transfected target cells, the expression of p16ink4a and hRb1 mRNA and protein were detected respectively in vitro. It was demonstrated with subG1 flow cytometric analysis and MTT method that p16ink4a and hRb1 genes cooperation more significantly inhibited cell growth and induced a more marked G1 arrest and apoptosis than p16ink4a/hRb1 alone (P < 0.01). Coexpression of exogenous p16ink4a with hRb1 broke the regulatory feedback loop of p16ink4a-cyclinD1 /CDK-hRb1 and played a more significant role in inhibiting cell growth as well as inducing cell apoptosis than p16ink4a or hRb1 did alone in vitro.', 'http://www.ncbi.nlm.nih.gov/pubmed/16696324', 'no', 'no', 'NA', 2),
(16705726, 'Wang W, Luo H, Wang A', 'Expression of survivin and correlation with PCNA in osteosarcoma.', 'J Surg Oncol', '2006', 'Expression of survivin and correlation with PCNA in osteosarcoma. Survivin is a new anti-apoptosis gene with reported expression during fetal development and in cancer tissues. Proliferating cell nuclear antigen (PCNA) is essential for the replication of deoxyribonucleic acid (DNA). In this experiment, we examined the expression of survivin and PCNA in human osteosarcoma tissues and investigating whether survivin and PCNA expressions were correlated with histological grades, histological types, and patients'' clinical characteristics.Sixty-eight osteosarcoma samples were divided into two groups based on the degree of tumor cell differentiation. Immunohistochemical staining was performed to analyze the expression of survinin and PCNA. Four histological types and patients'' clinical characteristics were also recorded.There were abundant levels of survivin and PCNA immunoreactivity in the nucleus and/or cytoplasm of the osteosarcoma cells. All cells essentially revealed the cytoplasmic localization and the nuclear signals of survivin in same cases, while PCNA from the majority of cases predominantly showed nuclear expression. Scoring on the percentage of positive cells indicated that survivin expression was significantly associated with the PCNA-labeling index (LI), which was correlated with the histological grades of osteosarcoma (chi2 = 17.86 and chi2 = 23.75, respectively).The expression of survivin can be used as a useful indicator in grading the malignancy of osteosarcoma.Copyright 2006 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/16705726', 'yes', 'yes', 'NA', 2);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(16734381, 'Goto T, Matsui Y, Fernandes RJ, Hanson DA, Kubo T, Yukata K, Michigami T, Komori T, Fujita T, Yang L, Eyre DR, Yasui N', 'Sp1 family of transcription factors regulates the human alpha2 (XI) collagen gene (COL11A2) in Saos-2 osteoblastic cells.', 'J Bone Miner Res', '2006', 'Sp1 family of transcription factors regulates the human alpha2 (XI) collagen gene (COL11A2) in Saos-2 osteoblastic cells. Genes encoding type XI collagen, normally associated with chondrogenesis, are also expressed by osteoblasts. By studying Saos-2 cells, we showed that the transcription factors, Sp1, Sp3, and Sp7 (Osterix), regulate COL11A2 expression through its proximal promoter. The findings indicate both ubiquitous and osteoblast-specific mechanisms of collagen gene regulation.Type XI collagen is essential for skeletal morphogenesis. Collagen XI gene regulation has been studied in chondrocytes but not in osteoblasts.We cultured Saos-2 cells, a human osteosarcoma-derived line of osteoblasts, and analyzed them for alpha2(XI) protein and COL11A2 regulatory mechanisms.Although types I and V were the dominant collagens deposited by Saos-2 cells, they expressed COL11A2 mRNA, and alpha2(XI) chains were present in the extracellular matrix. The COL11A2 promoter region (from -149 to -40) containing three Sp1 binding sites was required for promoter activity in transient transfection assays. All three Sp1 sites were critical for binding by nuclear proteins in electrophoretic mobility shift assays. Further analysis using consensus oligonucleotides and specific antibodies as well as chromatin immunoprecipitation assay implicated Sp1 and Sp3 in binding to this promoter region. Overexpressing Sp1 or Sp3 significantly increased COL11A2 promoter activity and endogenous COL11A2 gene expression, an effect that was suppressed by the Sp1-binding inhibitor mithramycin A. Further experiments showed that Sp1, Sp3, CREB-binding protein (CBP), p300, and histone deacetylase (HDAC) were physically associated and HDAC inhibitors (trichostatin A or NaB) upregulated COL11A2 promoter activity and endogenous gene expression. Another Sp1 family member, Sp7 (Osterix), was expressed in Saos-2 cells, but not in chondrocytes, and was shown by chromatin immunoprecipitation to occupy the COL11A2 promoter. Overexpressing Sp7 increased COL11A2 promoter activity and endogenous gene expression, an effect also blocked by mithramycin A. Using siRNA to knockdown Sp1, Sp3, or Sp7, it was shown that depression of any of them decreased COL11A2 promoter activity and endogenous gene expression. Finally, primary cultures of osteoblasts expressed COL11A2 and Sp7, upregulated COL11A2 promoter activity and endogenous gene expression when Sp1, Sp3, or Sp7 were overexpressed, and downregulated them when Sp1, Sp3, or Sp7 were selectively depressed. The results establish that Sp1 proteins regulate COL11A2 transcription by binding to its proximal promoter and directly interacting with CBP, p300, and HDAC.', 'http://www.ncbi.nlm.nih.gov/pubmed/16734381', 'no', 'no', 'NA', 6),
(16751779, 'Ternovoi VV, Curiel DT, Smith BF, Siegal GP', 'Adenovirus-mediated p53 tumor suppressor gene therapy of osteosarcoma.', 'Lab Invest', '2006', 'Adenovirus-mediated p53 tumor suppressor gene therapy of osteosarcoma. The clinical outcome for osteosarcoma (OS) remains discouraging despite efforts to optimize treatment using conventional modalities including surgery, radiotherapy and chemotherapy. Novel therapeutic approaches based on our expanding understanding of the mechanisms of tumor cell killing have the potential to alter this situation. Tumor suppressor gene therapy aims to restore the function of a tumor suppressor gene lost or functionally inactivated in cancer cells. One such molecule, the p53 tumor suppressor gene plays a critical role in safeguarding the integrity of the genome and preventing tumorigenesis. Introduction of wild-type (wt) p53 into transformed cells has been shown to be lethal for most cancer cells in vitro, but clinical trials of p53 gene replacement have had limited success. Analysis of these clinical trials highlighted the insufficient efficacy of current vectors and low proapoptotic activity of wt p53 as a single agent in vivo. In this review, a contemporary summarization of the current status of adenovirus-mediated p53 gene therapy of OS is presented. Advancement in our understanding of p53 tumor suppressor activity, the molecular biology of chemoresistant OS, and recent advances in tumor targeting with adenoviral vectors are also addressed. Based on these parameters, prospects for future investigations are proposed.', 'http://www.ncbi.nlm.nih.gov/pubmed/16751779', 'no', 'no', 'NA', 1),
(16759849, 'Díaz-Montero CM, Wygant JN, McIntyre BW', 'PI3-K/Akt-mediated anoikis resistance of human osteosarcoma cells requires Src activation.', 'Eur J Cancer', '2006', 'PI3-K/Akt-mediated anoikis resistance of human osteosarcoma cells requires Src activation. Considerable advances in understanding the mechanisms associated with anoikis resistance of normal and malignant epithelial cells have been made. However, little is still known about the pathways involved in anoikis resistance of non-epithelial cells such as fibroblasts and sarcomas. Our results show that Src activity contributes to anoikis resistance of human osteosarcoma SAOS-2 cells. Src was found to be upregulated in anoikis resistant SAOS cells, and pharmacological inhibition of its activity resulted in the restoration of anoikis sensitivity. A normal pattern of dephosphorylation of FAK was observed upon cell detachment of both anoikis sensitive and resistant SAOS-2 cells, suggesting that FAK activity during anoikis resistance is not essential. The activity of Akt was found to be upregulated in anoikis resistant SAOSar cells and the pharmacological inhibition of PI3-K activity restored sensitivity to anoikis resistant cells, reconfirming the critical role of PI3-K/Akt pathway in cell survival. Furthermore, pharmacological inhibition of Src resulted in a decrease of Akt phosphorylation at Ser473. Altogether, these studies indicated a survival pathway mediated by the Src-dependent activation of the PI3-K/Akt pathway in a manner independent of FAK activity.', 'http://www.ncbi.nlm.nih.gov/pubmed/16759849', 'no', 'no', 'NA', 2),
(16764926, 'Müller CI, Miller CW, Hofmann WK, Gross ME, Walsh CS, Kawamata N, Luong QT, Koeffler HP', 'Rare mutations of the PIK3CA gene in malignancies of the hematopoietic system as well as endometrium, ovary, prostate and osteosarcomas, and discovery of a PIK3CA pseudogene.', 'Leuk Res', '2007', 'Rare mutations of the PIK3CA gene in malignancies of the hematopoietic system as well as endometrium, ovary, prostate and osteosarcomas, and discovery of a PIK3CA pseudogene. Lipid kinase PIK3CA mutations have been described in several cancers. They clustered in two ''hot spots'' located in helical (exon 9) and kinase (exon 20) domains associated with increased kinase activity strongly suggesting oncogenic potential. Mutational analysis of previously unexamined tumors showed an amino acid change from threonine to alanine (T1025A) in exon 20 in one of 28 endometrial cancer samples and 6 endometrial cell lines. Additionally, a silent polymorphism (T1025T) was found in two of 20 MDS samples, one of 43 NHL samples, two of 40 osteosarcoma samples and Ishikawa. The polymorphism was established by identifying two of 92 normal samples with the same change. No PIK3CA mutations were found in acute myeloid leukemia (AML), myelodysplastic syndromes (MDS) and non-Hodgkin lymphomas (NHL) as well as in osteosarcomas, prostate and ovarian cancer samples. Additionally, a previously unidentified PIK3CA pseudogene spanning exons 9-13 on chromosome 22 was discovered.', 'http://www.ncbi.nlm.nih.gov/pubmed/16764926', 'yes', 'no', 'NA', 1),
(16773198, 'Tofuku K, Yokouchi M, Murayama T, Minami S, Komiya S', 'HAS3-related hyaluronan enhances biological activities necessary for metastasis of osteosarcoma cells.', 'Int J Oncol', '2006', 'HAS3-related hyaluronan enhances biological activities necessary for metastasis of osteosarcoma cells. Several studies have suggested that increased production of hyaluronan (HA) is associated with metastatic behavior in various malignant tumors. To our knowledge, HA molecular weights required for metastasis are still unsolved in osteosarcoma. We examined the size of HA and hyaluronan synthase (HAS) isoforms related to biological functions required for metastasis in the LM8 stably highly metastatic osteosarcoma cell line. We found that HA of molecular weight which HAS3 produces enhanced biological functions related to metastasis such as cell proliferation, invasion, and degradation of extracellular matrix. Moreover, cell proliferation and invasion were inhibited by suppressing the activity of HAS3 expressed in LM8 cells, using hyaluronan synthase suppressor, 4-methylumbelliferone (MU). HA with the molecular weight related to HAS2 was the most adherent to CD44 in LM8 cells, suggesting that HAS2 may play an important role in pericellular coat formation. These results suggest that HAS3-related HA enhances crucial biological activities necessary for metastasis and that HAS2-related HA offers an advantageous environment for osteosarcoma cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/16773198', 'no', 'no', '4-methylumbelliferone', 3),
(16786118, 'Ek ET, Ojaimi J, Kitagawa Y, Choong PF', 'Does the degree of intratumoural microvessel density and VEGF expression have prognostic significance in osteosarcoma?', 'Oncol Rep', '2006', 'Does the degree of intratumoural microvessel density and VEGF expression have prognostic significance in osteosarcoma? The role of angiogenesis as a prognostic indicator in cancer has been extensively studied in recent times with several studies demonstrating a positive correlation for various malignant tumours. However, the role of angiogenesis in osteosarcoma remains a topic of debate. In this study, we aim to evaluate the significance of intratumoural microvessel density (MVD) and the degree of vascular epithelial growth factor (VEGF) expression as markers of angiogenesis and correlate this with disease outcome. Archival paraffin-embedded pre-treatment biopsy tissue of patients treated at St. Vincent''s Hospital, Melbourne, with non-metastatic osteosarcoma at initial diagnosis was reviewed. Tissue was processed for immunofluorescent staining of the microvascular endothelial cells with antibodies directed against CD31 and CD34. The degree of angiogenesis was assessed, as determined by the microvessel density (MVD). Further histological examination was performed to assess the degree of VEGF expression. Histological observations were correlated with various clinicopathological factors and patient outcome in terms of recurrence, metastasis and death. Twenty-five cases were reviewed, 15 were male and 10 were female, and the median age was 26 years (range, 13-85). The mean follow-up was 21.5 months (range, 3-75 months). The median MVD was 43 microvessels/0.26 mm2 (range, 25-54) and 46 microvessels/0.26 mm2 (range, 30-58) for CD31 and CD34, respectively. Despite the moderate to high vascularity, there was no significant difference noted between the MVD and disease outcome factors for both CD31 and CD34. There was a trend towards a higher MVD in patients aged > 40 years compared to those < 40 years (p = 0.110 for CD31 and p = 0.097 for CD34). In terms of VEGF expression, 24 of 25 cases demonstrated either moderate or strong expression; however, no prognostic significance was determined. In this study, we were able to demonstrate that osteosarcoma is a relatively vascular tumour; however, the degree of MVD and VEGF expression does not provide prognostic information. It is likely that angiogenesis plays a key role in the pathogenesis of osteosarcoma and is, therefore, a potential target for novel anti-angiogenic therapies.', 'http://www.ncbi.nlm.nih.gov/pubmed/16786118', 'yes', 'no', 'NA', 1),
(16795036, 'Ozaki A, Morimoto H, Tanaka H, Okamura H, Yoshida K, Amorim BR, Haneji T', 'Okadaic acid induces phosphorylation of p65NF-kappaB on serine 536 and activates NF-kappaB transcriptional activity in human osteoblastic MG63 cells.', 'J Cell Biochem', '2006', 'Okadaic acid induces phosphorylation of p65NF-kappaB on serine 536 and activates NF-kappaB transcriptional activity in human osteoblastic MG63 cells. Nuclear factor-kappa B (NF-kappaB) is an essential transcription factor in the control of expression of genes involved in cell growth, differentiation, inflammation, and neoplastic transformation. Previously, we reported that okadaic acid (OA), which is a specific inhibitor of serine/threonine protein phosphatases, induced apoptosis in cells of human osteosarcoma cell line MG63. However, to date, it is not clear whether the phosphorylation status of NF-kappaB could be affected by the treatment with OA. In this report, we demonstrate that treatment of MG63 cells with OA enhanced the phosphorylation level of NF-kappaB, as judged from the results of Western blot analysis and a lambda protein phosphatase dephosphorylation assay. The phosphorylation level of NF-kappaB was enhanced in both time- and dose-dependent manners. In the cells treated with 100 nM OA for 3 h, consequential translocation of NF-kappaB from the cytosol to the nucleus occurred. Western blotting experiments with an anti-phospho-p65NF-kappaB antibody disclosed that the NF-kappaB was phosphorylated on serine 536. Furthermore, OA stimulated the transcriptional activity of NF-kappaB in MG63 cells, as judged from the results of a luciferase assay. Our findings indicate that OA elicit phosphorylation of NF-kappaB on serine 536 in MG63 cells, resulting in the translocation of phospho-NF-kappaB to the nucleus, thereby promoting transcriptional activity of genes.2006 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/16795036', 'no', 'no', 'okadaic acid', 1),
(16797629, 'Hafeez BB, Ahmed S, Wang N, Gupta S, Zhang A, Haqqi TM', 'Green tea polyphenols-induced apoptosis in human osteosarcoma SAOS-2 cells involves a caspase-dependent mechanism with downregulation of nuclear factor-kappaB.', 'Toxicol Appl Pharmacol', '2006', 'Green tea polyphenols-induced apoptosis in human osteosarcoma SAOS-2 cells involves a caspase-dependent mechanism with downregulation of nuclear factor-kappaB. Development of chemotherapy resistance and evasion from apoptosis in osteosarcoma, a primary malignant bone tumor, is often correlated with constitutive nuclear factor-kappaB (NF-kappaB) activation. Here, we investigated the ability of a polyphenolic fraction of green tea (GTP) that has been shown to have antitumor effects on various malignant cell lines to inhibit growth and induce apoptosis in human osteosarcoma SAOS-2 cells. Treatment of SAOS-2 cells with GTP (20-60 microg/ml) resulted in reduced cell proliferation and induction of apoptosis, which correlated with decreased nuclear DNA binding of NF-kappaB/p65 and lowering of NF-kappaB/p65 and p50 levels in the cytoplasm and nucleus. GTP treatment of cells reduced IkappaB-alpha phosphorylation but had no effect on its protein expression. Furthermore, GTP treatment resulted in the inhibition of IKK-alpha and IKK-beta, the upstream kinases that phosphorylate IkappaB-alpha. The increase in apoptosis in SAOS-2 cells was accompanied with decrease in the protein expression of Bcl-2 and concomitant increase in the levels of Bax. GTP treatment of SAOS-2 cells also resulted in significant activation of caspases as was evident by increased levels of cleaved caspase-3 and caspase-8 in these cells. Treatment of SAOS-2 cells with a specific caspase-3 inhibitor Ac-Asp-Glu-Val-Asp-CHO (Ac-DEVD-CHO) and general caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp (OMe)-fluoromethyl ketone (Z-VAD-FMK) rescued SAOS-2 cells from GTP-induced apoptosis. Taken together, these results indicate that GTP is a candidate therapeutic for osteosarcoma that mediates its antiproliferative and apoptotic effects via activation of caspases and inhibition of NF-kappaB.', 'http://www.ncbi.nlm.nih.gov/pubmed/16797629', 'no', 'no', 'polyphenolic fraction of green tea', 6),
(16798617, 'Indovina P, Ferrante A, Rainaldi G, Santini MT', 'Hypoxia and ionizing radiation: changes in adhesive properties and cell adhesion molecule expression in MG-63 three-dimensional tumor spheroids.', 'Cell Commun Adhes', 'NA', 'Hypoxia and ionizing radiation: changes in adhesive properties and cell adhesion molecule expression in MG-63 three-dimensional tumor spheroids. The effects of chemically induced hypoxia and ionizing radiation on the adhesive properties of MG-63 human osteosarcoma three-dimensional spheroids were investigated. Hypoxia was induced by addition of CoCl2 to small, nonhypoxic spheroids and verified by HIF-1alpha expression. In addition, the possible role of important cell adhesion molecules involved in tumor dissemination in inducing adhesive changes were also studied. In particular, two key integrins (i.e., the alpha chain of the fibronectin receptor, alpha5, and the alpha chain of the collagen receptor, alpha2), an important member of the immunoglobulin superfamily (CD54 or ICAM-1) and the strategic molecule CD44 (H-CAM, the principal receptor for hyaluronan) were examined. Because of the important role of fibronectin in adhesive processes, variations in this extracellular matrix component were also examined. The results seem to indicate that CoCl2-induced hypoxia greatly increases adhesion of MG-63 spheroids to both tissue culture plates and plates coated with fibronectin or collagen when compared to controls, while ionizing radiation induces a great decrease in this attachment. Furthermore, chemically induced hypoxia also partially inhibits the effects of ionizing radiation. The data also show that these adhesive changes are accompanied by concomitant variations in the expression of alpha5 and alpha2 integrins, CD44, and CD54 and fibronectin.', 'http://www.ncbi.nlm.nih.gov/pubmed/16798617', 'no', 'no', 'CoCl2', 6),
(16814113, 'Alkhalaf M, Jaffal S', 'Potent antiproliferative effects of resveratrol on human osteosarcoma SJSA1 cells: Novel cellular mechanisms involving the ERKs/p53 cascade.', 'Free Radic Biol Med', '2006', 'Potent antiproliferative effects of resveratrol on human osteosarcoma SJSA1 cells: Novel cellular mechanisms involving the ERKs/p53 cascade. The chemopreventive activity of resveratrol (RSVL) has been demonstrated in several types of cancer. However, its effects and the underling mechanisms remain poorly understood. In this study, we investigated the involvement of the mitogen activated protein kinase (MAPK)/p53 signal transduction mechanism in RSVL-induced growth inhibition using a human osteosarcoma cell line. We demonstrate that RSVL reduces cell viability and growth of SJSA1 osteosarcoma cells. Morphological profiles and 4,6-diamidino-2-phenylindole nuclear staining of RSVL-treated cells indicated marked nuclear fragmentation. Cleavage of the (116-kDa) poly(ADP-ribose) polymerase protein into an 89-kDa fragment (a proapoptotic marker system) was substantially augmented by RSVL treatment. RSVL-dependent growth impairment was preceded by enhanced phosphorylation of extracellular signal-regulated kinase (ERK)1/2 (at Thr202/Tyr204). Likewise, RSVL increased the phosphorylation of p53 tumor suppressor protein (at Ser15). The effects of RSVL on ERKs and on p53 phosphorylation were abrogated by either the MAPK inhibitor PD98059 or the p53 inhibitor pifithrine-alpha. The present study indicates that RSVL antiproliferative effects on osteosarcoma cells are mediated by the activation of the ERKs/p53 signaling pathway and therefore identifies new targets for strategies to treat and/or prevent osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/16814113', 'no', 'no', 'resveratrol|PD98059|pifithrine-alpha', 4),
(16818639, 'Xu Z, Choudhary S, Voznesensky O, Mehrotra M, Woodard M, Hansen M, Herschman H, Pilbeam C', 'Overexpression of COX-2 in human osteosarcoma cells decreases proliferation and increases apoptosis.', 'Cancer Res', '2006', 'Overexpression of COX-2 in human osteosarcoma cells decreases proliferation and increases apoptosis. Overexpression of cyclooxygenase-2 (COX-2) is generally considered to promote tumorigenesis. To investigate a potential role of COX-2 in osteosarcoma, we overexpressed COX-2 in human osteosarcoma cells. Saos-2 cells deficient in COX-2 expression were retrovirally transduced or stably transfected with murine COX-2 cDNA. Functional expression of COX-2 was confirmed by Northern and Western analyses and prostaglandin production. Overexpression of COX-2 reduced cell numbers by 50% to 70% compared with controls. Decreased proliferation in COX-2-overexpressing cells was associated with cell cycle prolongation in G(2)-M. Apoptosis, measured by both Annexin V binding assay and terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling staining, was increased in cells overexpressing COX-2, and the increase was not reversed by treatment with NS-398, indicating that the effects were not mediated by prostaglandins. Retroviral COX-2 overexpression in two other human osteosarcoma cell lines, U2OS and TE85, also decreased cell viability. However, in the human colon carcinoma HCT-116 cell line, which is deficient in COX-2, retroviral overexpression of COX-2, at similar efficiency as in Saos-2 cells, increased resistance to apoptosis. Reactive oxygen species (ROS), measured by flow cytometry, were increased by COX-2 overexpression in Saos-2 cells but not in HCT-116 cells. Inhibition of peroxidase activity, but not of COX activity, blocked the ROS increase. Antioxidants blocked the increase in ROS and the increase in apoptosis due to COX-2 overexpression in Saos-2 cells. Our results suggest that (a) COX-2 overexpression in osteosarcoma cells may increase resistance to tumorigenesis by increasing ROS to levels that decrease cell viability and (b) the effects of COX-2 overexpression are cell type/tissue dependent.', 'http://www.ncbi.nlm.nih.gov/pubmed/16818639', 'no', 'no', 'NA', 1),
(16822878, 'Hosono A, Yamaguchi U, Makimoto A, Endo M, Watanabe A, Shimoda T, Kaya M, Matsumura T, Sonobe H, Kusumi T, Yamaguchi T, Hasegawa T', 'Utility of immunohistochemical analysis for cyclo-oxygenase 2 in the differential diagnosis of osteoblastoma and osteosarcoma.', 'J Clin Pathol', '2007', 'Utility of immunohistochemical analysis for cyclo-oxygenase 2 in the differential diagnosis of osteoblastoma and osteosarcoma. To study the immunoexpression of cyclo-oxygenase (COX) 2 in osteoblastomas (OBs) and osteosarcomas (OSs), and to assess the utility of immunohistochemical analysis for COX 2 in the differential diagnosis of the two tumour forms.The immunohistochemical features of COX 2 were studied in 11 OBs and 30 OSs, including 26 high-grade OSs (16 osteoblastic, 7 chondroblastic, and 3 fibroblastic) and 4 low-grade OSs.Tumour cells from all 11 OBs unequivocally showed diffuse, intense and cytoplasmic immunoreactivity for COX 2. Strong cytoplasmic expression of COX 2 was observed in 5 of 26 (19%) high-grade OSs, all chondroblastic. In one osteoblastic-type OS, COX 2 was expressed in the chondroblastic component, but this tumour was considered to be COX 2 negative. No COX 2 expression was noted in atypical osteoblastic cells. Staining in the four low-grade OSs was negative.The results of immunohistochemical analysis of COX 2 suggest that in addition to the routine histopathological evaluation, COX 2 is a valuable diagnostic marker in the distinction between OB and OS.', 'http://www.ncbi.nlm.nih.gov/pubmed/16822878', 'yes', 'yes', 'NA', 1),
(16826116, 'Charity RM, Foukas AF, Deshmukh NS, Grimer RJ', 'Vascular endothelial growth factor expression in osteosarcoma.', 'Clin Orthop Relat Res', '2006', 'Vascular endothelial growth factor expression in osteosarcoma. Vascular endothelial growth factor expression by osteosarcomas at biopsy has been shown to have a negative influence on survival. Whether it continues to be expressed and influence outcome in tumors after neoadjuvant chemotherapy has not been determined. To determine if a high-degree of vascular endothelial growth factor expression by the surviving osteosarcoma cells in the resected tumor has a negative influence on prognosis, we retrospectively reviewed 52 patients with Stage IIB osteosarcoma around the knee, 48 of whom received neoadjuvant chemotherapy. Vascular endothelial growth factor expression by the surviving tumor cells in the surgically resected specimens was determined by immunohistochemical staining. Followup was for a minimum of 92 months. The presence of vascular endothelial growth factor expression in greater than 25% of tumor cells was independently associated with a reduced overall and disease-free survival rate.Prognostic study, Level II (retrospective study). See the Guidelines for Authors for a complete description of levels of evidence.', 'http://www.ncbi.nlm.nih.gov/pubmed/16826116', 'yes', 'yes', 'NA', 1),
(16827134, 'Kim SG, Kim HH, Kim HK, Kim CH, Chun HS, Kanai Y, Endou H, Kim do K', 'Differential expression and functional characterization of system L amino acid transporters in human normal osteoblast cells and osteogenic sarcoma cells.', 'Anticancer Res', 'NA', 'Differential expression and functional characterization of system L amino acid transporters in human normal osteoblast cells and osteogenic sarcoma cells. The amino acid transport system L is a major nutrient transport system responsible for Na(+)-independent transport of neutral amino acids, including several essential amino acids. The system L is divided into two major subgroups, the L-type amino acid transporter 1 (LAT1) and the L-type amino acid transporter 2 (LAT2). In malignant tumors, the LAT1 is highly expressed to support tumor cell growth. In the present study, the expressions and functions of the system L amino acid transporters were examined and compared in both FOB human osteoblast cells and Saos2 human osteogenic sarcoma cells.The expressions and functions of the system L amino acid transporters in both FOB and Saos2 cells were examined using RT-PCR, Western blot analysis and amino acid transport measurement.RT-PCR and Western blot analysis revealed that the FOB and Saos2 cells expressed LAT1 and LAT2, together with their associated protein 4F2hc, but the expression of LAT2 in the Saos2 cells was very weak. The uptakes of [14C]L-leucine by FOB and Saos2 cells were Na(+)-independent and were completely inhibited by the system L selective inhibitor, 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH). The affinity and the inhibition profiles of [14C]L-leucine uptake by various amino acids in the FOB and Saos2 cells were comparable with those for the LAT2 and LAT1 expressed in Xenopus oocytes, respectively. The majority of [14C]L-leucine uptake is, therefore, mediated by LAT2 and LAT1 in FOB and Saos2 cells, respectively.These results suggest that the transport of neutral amino acids, including several essential amino acids into the FOB and Saos2 cells, are mainly mediated by LAT2 and LAT1, respectively. Moreover, the specific inhibition of LAT1 in tumor cells might be a new rationale for antitumor therapy.', 'http://www.ncbi.nlm.nih.gov/pubmed/16827134', 'no', 'no', 'NA', 2),
(16849574, 'Russo AJ, Magro PG, Hu Z, Li WW, Peters R, Mandola J, Banerjee D, Bertino JR', 'E2F-1 overexpression in U2OS cells increases cyclin B1 levels and cdc2 kinase activity and sensitizes cells to antimitotic agents.', 'Cancer Res', '2006', 'E2F-1 overexpression in U2OS cells increases cyclin B1 levels and cdc2 kinase activity and sensitizes cells to antimitotic agents. The E2F transcription factors play a critical role in coordinating transcription of specific genes essential for G1-S transition. In early G1, the retinoblastoma protein (pRB) becomes phosphorylated by cyclin-dependent kinases, disrupting pRB binding to E2F-1-3, allowing \\free\\ E2F to regulate genes involved in proliferation. In the present study, we used a tetracycline E2F-1 inducible U2OS osteosarcoma cell line to investigate the effect of increasing levels of E2F-1 on the cytotoxicity of various chemotherapeutic drugs. Upon overexpression of E2F-1, there was no detectable change in cytotoxicity to doxorubicin, cisplatin, 5-fluorouracil, or etoposide. In contrast, overexpression of E2F-1 resulted in a marked increase in sensitivity to vinblastine and paclitaxel, drugs that are known to be more effective against cells in M phase. Therefore, we investigated the effect of E2F-1 overexpression on proteins regulating the G2-M transition and M phase, in particular cyclin B1 and cdc2 kinase. Cyclin B1 mRNA and protein levels increased within 24 hours of E2F1 induction together with an increase in associated cdc2 kinase activity. Overexpression of cyclin B1 also resulted in a specific increase in sensitivity to paclitaxel and an increase in the cellular growth rate. Knockdown of cyclin B1 using an RNA interference oligo resulted in a slower cellular growth rate and an increase in resistance to paclitaxel. These studies add support to recent reports that show E2F regulates genes involved in mitotic entry and exit and allow the suggestion that mitotic inhibitors may have selective effects in tumors that overexpress E2F-1.', 'http://www.ncbi.nlm.nih.gov/pubmed/16849574', 'no', 'no', 'paclitaxel', 3),
(16857797, 'Srivastava A, Fuchs B, Zhang K, Ruan M, Halder C, Mahlum E, Weber K, Bolander ME, Sarkar G', 'High WT1 expression is associated with very poor survival of patients with osteogenic sarcoma metastasis.', 'Clin Cancer Res', '2006', 'High WT1 expression is associated with very poor survival of patients with osteogenic sarcoma metastasis. Although metastasis is the primary determinant of poor survival of patients with osteogenic sarcoma, some patients live much longer than others, indicating metastatic heterogeneity underlying survival outcome. The purpose of the investigation was to identify genes underlying survival outcome of patients with osteogenic sarcoma metastasis.We have used microarray to first compare mRNA expression between normal bone and osteogenic sarcoma specimens, identified genes overexpressed in osteogenic sarcoma, and compared expression of the selected gene between a poorly metastatic (SAOS) and two highly metastatic cell lines (LM8 and 143B). Finally, expression of the selected gene was assessed by immunostaining of osteogenic sarcoma samples with known survival outcome.Microarray analysis revealed 5.3-fold more expression of WT1 mRNA in osteogenic sarcoma compared with normal bone and >2-fold overexpression in 143B and LM8 cells compared with SAOS. Furthermore, WT1 mRNA was absent in normal bone (10 of 10) by reverse transcription-PCR but present in osteogenic sarcoma-derived cell lines (5 of 8). One hundred percent (42 of 42) of low-grade osteogenic sarcoma specimens expressed no WT1 as determined by immunostaining; however, 24% (12 of 49) of the high-grade specimens showed intense staining. Mean survival of patients with high-grade metastatic osteogenic sarcoma but low WT1 staining (27 of 37) was 96.5 +/- 129.3 months, whereas mean survival of patients with high-grade metastatic osteogenic sarcoma having intense staining (10 of 37) was 18.3 +/- 12.3 months (P > 0.0143). All splice variants of WT1 mRNA, including a hitherto unknown variant (lacking exons 4 and 5), were found to be expressed in osteogenic sarcoma.WT1 seems to be associated with very poor survival of patients with osteogenic sarcoma metastasis.', 'http://www.ncbi.nlm.nih.gov/pubmed/16857797', 'yes', 'yes', 'NA', 1),
(16878156, 'de Nigris F, Botti C, Rossiello R, Crimi E, Sica V, Napoli C', 'Cooperation between Myc and YY1 provides novel silencing transcriptional targets of alpha3beta1-integrin in tumour cells.', 'Oncogene', '2007', 'Cooperation between Myc and YY1 provides novel silencing transcriptional targets of alpha3beta1-integrin in tumour cells. We show that human osteosarcoma cells (Saos-2) have downregulation of alpha3beta1-integrin compared to normal bone cells; this was further described in human osteosarcomas and in a primary murine sarcoma. The alpha3 gene was silenced in Saos-2 cells causing a low expression of alpha3beta1-integrin and reduction in collagen attachment with increasing migratory capacity. Chromatin immunoprecipitation assay performed on alpha3 promoter established that Myc and Yin Yang protein (YY1) cooperate in tandem to downregulate the alpha3 gene. This silencing mechanism involves the binding of Myc and YY1 to DNA and formation of complexes among Myc/Max, YY1, CREB-binding protein and deacetylation activity. The promoter containing deletions of E-boxes or YY1 cassettes failed to downregulate the transcription of a reporter gene as well as the inhibition of deacetylation activity. Overexpression of both Myc and YY1 was necessary to determine the alpha3-integrin promoter downregulation in normal osteoblasts. This downregulation of alpha3beta1-integrin can contribute to the acquisition of a more aggressive phenotype. YY1 regulated negatively the Myc activity through a direct interaction with the Myc/Max and deacetylase complexes. This represents a novel silencing mechanism with broad implications in the transcription machinery of tumours.', 'http://www.ncbi.nlm.nih.gov/pubmed/16878156', 'no', 'no', 'NA', 6),
(16933368, 'Trieb K, Blahovec H, Kubista B', 'Effects of hyperthermia on heat shock protein expression, alkaline phosphatase activity and proliferation in human osteosarcoma cells.', 'Cell Biochem Funct', 'NA', 'Effects of hyperthermia on heat shock protein expression, alkaline phosphatase activity and proliferation in human osteosarcoma cells. Hyperthermia can be used as a possible adjuvant therapy in treatment of cancer patients. In this study, the direct effect of hyperthermia on osteosarcoma derived cell lines HOS85, MG-63 and SaOS-2 was investigated. Heat shock at 42 degrees C inhibited proliferation significantly in all three cell lines tested. Furthermore a sub-lethal heat shock (42 degrees C, 1 h) decreases alkaline phosphatase activity, the absolute marker for osteoblast-like cells, in all of the three cell lines. Hsp70 was expressed constitutively and was found to be upregulated in a time-dependent manner; by up to 150% in Western blot analysis. The results of this study indicate that heat shock has an inhibitory effect on human osteosarcoma cells. These data suggest that hyperthermia has an anti-tumour effect on cancers of the bone and might, therefore, become an adjuvant treatment option.Copyright (c) 2006 John Wiley & Sons, Ltd.', 'http://www.ncbi.nlm.nih.gov/pubmed/16933368', 'no', 'no', 'NA', 2),
(16938571, 'Yan T, Wunder JS, Gokgoz N, Seto KK, Bell RS, Andrulis IL', 'hCDC4 variation in osteosarcoma.', 'Cancer Genet Cytogenet', '2006', 'hCDC4 variation in osteosarcoma. The hCDC4 gene (also known as Fbw7 or Archipelago) encodes an F-box protein that is responsible for targeting cyclin E for Skp1-cullin-F box protein (SCF) ubiquitination and proteosomal degradation. Disruption of this pathway has been associated with chromosomal instability and aneuploidy in several cancer cell lines and primary tumors. This study aimed to examine whether hCDC4 mutations contribute to aneuploidy in osteosarcoma. We analyzed 147 primary high-grade osteosarcoma specimens and 6 osteosarcoma cell lines. The protein truncation test (PTT) and single-strand conformation polymorphism (SSCP) analysis with subsequent sequencing were performed to detect alterations of the hCDC4 gene. All specimens exhibited the same PTT pattern of normal bands with less intense common bands. Two shifts were detected by SSCP, and subsequent DNA analysis identified one in-frame three-base GAG (424-426) deletion and one silent nucleotide substitution (C1261T). We conclude that somatic hCDC4 mutations are infrequent in osteosarcoma, and are unlikely to play an important role in aneuploidy of this tumor.', 'http://www.ncbi.nlm.nih.gov/pubmed/16938571', 'yes', 'no', 'NA', 1),
(16944980, 'Leavey PJ, Timmons C, Frawley W, Lombardi D, Ashfaq R', 'KAI-1 expression in pediatric high-grade osteosarcoma.', 'Pediatr Dev Pathol', 'NA', 'KAI-1 expression in pediatric high-grade osteosarcoma. Recent evidence implicates cell surface proteins of the tetraspanin superfamily in the process of metastasis whereas the downregulation of KAI-1, a member of the tetraspanin family, is associated with an aggressive clinical phenotype in several types of human cancers. To determine if expression of KAI-1-1 is associated with any known prognostic marker or clinical outcome in high-grade osteosarcoma, we examined 91 nondecalcified archival samples from 47 patients for the expression of KAI-1. Archival, paraffin-embedded, and decalcified pathologic samples were examined by immunohistochemistry and results were correlated to clinical outcomes and known prognostic markers. There were 46 samples from diagnostic biopsies (1 diagnostic sample was not available), 32 tumor resection samples, and 13 metastasis samples. Thirty-three percent (n=30) of the samples expressed KAI-1 (16 biopsies, 9 resections, and 5 metastasis). KAI-1 expression was not significantly related to known prognostic markers or to either tumor necrosis after neoadjuvant therapy or the incidence of metastasis at diagnosis. KAI-1 expression was not significantly different between paired diagnostic tumor samples and either resection or metastasis tumor samples. Twenty-five patients remain alive at a median follow-up of 95 months. The overall and progression-free survival percentages at 5 years were 62% and 47% for KAI-1-positive patients and 49% and 38% for KAI-1-negative patients, respectively. This difference was not statistically significant. We conclude that KAI-1 is expressed in a proportion of high-grade osteosarcoma but is not of clinical significance and cannot be used to stratify treatment groups for these patients.', 'http://www.ncbi.nlm.nih.gov/pubmed/16944980', 'yes', 'no', 'NA', 1),
(16956425, 'Wiontzek M, Matziolis G, Schuchmann S, Gaber T, Krocker D, Duda G, Burmester GR, Perka C, Buttgereit F', 'Effects of dexamethasone and celecoxib on calcium homeostasis and expression of cyclooxygenase-2 mRNA in MG-63 human osteosarcoma cells.', 'Clin Exp Rheumatol', 'NA', 'Effects of dexamethasone and celecoxib on calcium homeostasis and expression of cyclooxygenase-2 mRNA in MG-63 human osteosarcoma cells. Glucocorticoids and selective COX-2 inhibitors are potent anti-inflammatory agents. They are also suggested to influence bone physiology and remodeling. Here we searched for effects of dexamethasone and celecoxib on crucial parameters of bone physiology that could be therapeutically relevant.The human osteosarcoma cell line MG-63 was used to measure effects of these drugs on (i) intracellular calcium concentration ([Ca2+]i) using a microfluorometric technique, (ii) alkaline phosphatase and osteocalcin levels (EIA) and (iii) the expression of cox-2 mRNA (quantitative real time PCR). Measurements were performed in Vitamine D-incubated quiescent cells and in cells stimulated with TNF-alpha and IL-1beta.We found the cytokine-stimulation to increase [Ca2+]i which was prevented by dexamethasone already after 30 min and still after 48 h. Dexamethasone was without any effect on [Ca2+]i in quiescent cells. Celecoxib had no measurable short-term or long-term effects neither in quiescent nor in stimulated cells. Vitamin D stimulated the expression of cox-2 mRNA which was further enhanced by TNF-alpha/IL-1beta. Dexamethasone did not have any measurable effects on COX-2 expression after 30 min, but a pronounced inhibition was seen after 48 h. In contrast, celecoxib had no effect on COX-2 expression. Neither of the drugs had any effect on the secretion of alkaline phosphatase and osteocalcin.We found dexamethasone to inhibit the [Ca2+]i increase in MG-63 cells following stimulation and to reduce considerably COX-2 expression via the genomic pathway. In contrast, celecoxib did not show any measurable short-term or long-term effects on the parameters of bone physiology measured.', 'http://www.ncbi.nlm.nih.gov/pubmed/16956425', 'no', 'no', 'dexamethasone|celecoxib', 3),
(16962318, 'de Nigris F, Botti C, de Chiara A, Rossiello R, Apice G, Fazioli F, Fiorito C, Sica V, Napoli C', 'Expression of transcription factor Yin Yang 1 in human osteosarcomas.', 'Eur J Cancer', '2006', 'Expression of transcription factor Yin Yang 1 in human osteosarcomas. The transcription factor Yin Yang 1 (YY1) is known to be present in some human cancer cell lines and its expression correlates with immune-mediated apoptosis. By using Western blot analysis, we have shown that the YY1 protein is strongly expressed in human osteosarcoma cells and localised mainly in the nucleus. Moreover, by using immunohistochemistry and RT-PCR techniques, we have analysed the expression of YY1 protein in biopsies from human osteosarcomas. The YY1 protein was not detectable by immunohistochemistry in osteoid tissue. However, its expression was restricted to osteosarcoma tissues. These data were confirmed by densitometric analysis of RT-PCR for YY1 expression. Thus, YY1 gene activation appears to be an early event in the process of osteoblastic transformation and its detection may represent, together with the analysis of other established markers, a useful diagnostic tool in human osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/16962318', 'yes', 'yes', 'NA', 1),
(16982772, 'Cai D, Zhang X, Shapiro GI', 'Combined depletion of cell cycle and transcriptional cyclin-dependent kinase activities induces apoptosis in cancer cells.', 'Cancer Res', '2006', 'Combined depletion of cell cycle and transcriptional cyclin-dependent kinase activities induces apoptosis in cancer cells. Selective cyclin-dependent kinase (cdk) 2 inhibition is readily compensated. However, reduced cdk2 activity may have antiproliferative effects in concert with other family members. Here, inducible RNA interference was used to codeplete cdk2 and cdk1 from NCI-H1299 non-small cell lung cancer and U2OS osteosarcoma cells, and effects were compared with those mediated by depletion of either cdk alone. Depletion of cdk2 slowed G1 progression of NCI-H1299 cells and depletion of cdk1 slowed G2-M progression in both cell lines, with associated endoreduplication in U2OS cells. However, compared with the incomplete cell cycle blocks produced by individual depletion, combined depletion had substantial consequences, with G2-M arrest predominating in NCI-H1299 cells and apoptosis the primary outcome in U2OS cells. In U2OS cells, combined depletion affected RNA polymerase II expression and phosphorylation, causing decreased expression of the antiapoptotic proteins Mcl-1 and X-linked inhibitor of apoptosis (XIAP), effects usually mediated by inhibition of the transcriptional cdk9. These events do not occur after individual depletion of cdk2 and cdk1, suggesting that reduction of cdk2, cdk1, and RNA polymerase II activities all contribute to apoptosis in U2OS cells. The limited cell death induced by combined depletion in NCI-H1299 cells was significantly increased by codepletion of cdk9 or XIAP or by simultaneous treatment with the cdk9 inhibitor flavopiridol. These results show the potency of concomitant compromise of cell cycle and transcriptional cdk activities and may guide the selection of clinical drug candidates.', 'http://www.ncbi.nlm.nih.gov/pubmed/16982772', 'no', 'no', 'NA', 5),
(17001316, 'Deshpande AM, Akunowicz JD, Reveles XT, Patel BB, Saria EA, Gorlick RG, Naylor SL, Leach RJ, Hansen MF', 'PHC3, a component of the hPRC-H complex, associates with E2F6 during G0 and is lost in osteosarcoma tumors.', 'Oncogene', '2007', 'PHC3, a component of the hPRC-H complex, associates with E2F6 during G0 and is lost in osteosarcoma tumors. Polyhomeotic-like 3 (PHC3) is a ubiquitously expressed member of the polycomb gene family and part of the human polycomb complex hPRC-H. We found that in normal cells PHC3 associated with both hPRC-H complex components and with the transcription factor E2F6. In differentiating and confluent cells, PHC3 and E2F6 showed nuclear colocalization in a punctate pattern that resembled the binding of polycomb bodies to heterochromatin. This punctate pattern was not seen in proliferating cells suggesting that PHC3 may be part of an E2F6-polycomb complex that has been shown to occupy and silence target promoters in G(0). Previous loss of heterozygosity (LoH) analyses had shown that the region containing PHC3 underwent frequent LoH in primary human osteosarcoma tumors. When we examined normal bone and human osteosarcoma tumors, we found loss of PHC3 expression in 36 of 56 osteosarcoma tumors. Sequence analysis revealed that PHC3 was mutated in nine of 15 primary osteosarcoma tumors. These findings suggest that loss of PHC3 may favor tumorigenesis by potentially disrupting the ability of cells to remain in G(0).', 'http://www.ncbi.nlm.nih.gov/pubmed/17001316', 'yes', 'no', 'NA', 2),
(17012855, 'Zhang HZ, Wang Y, Gao P, Lin F, Liu L, Yu B, Ren JH, Zhao H, Wang R', 'Silencing stathmin gene expression by survivin promoter-driven siRNA vector to reverse malignant phenotype of tumor cells.', 'Cancer Biol Ther', '2006', 'Silencing stathmin gene expression by survivin promoter-driven siRNA vector to reverse malignant phenotype of tumor cells. Stathmin gene overexpression has been shown to play an important role in maintenance of malignant phenotype in tumor cells, and the blocking efficacy and tumor specificity of this target has been concerned in clinical trails. In this report, we designed survivin promoter-driven siRNA eukaryotic expression vector that expressed the small interfering RNA targeting stathmin gene to selectively knock down the stathmin gene expression in two different kinds of tumor cell lines while sparing normal cell lines. The therapeutic potential of this recombinant vector was tested in human cervical cancer Hela cells and osteosarcoma SSOP-9607 cells, and in human umbilical vein endothelial cell line ECV304 cells as control. The siRNA vector- transfected Hela cells and SSOP-9607 cells revealed marked inhibition of stathmin expression and a dramatic growth inhibition comparing with ECV304 cells, parental-vector transfected cells and untransfected cells. Cell cycle analysis of siRNA vector transfected tumor cells by Flow Cytometry showed G(2)/M phase block, while morphologic analysis by TURNEL staining method showed marked increase of apoptosis. Our study indicates that survivin gene promoter-driven stathmin siRNA expression vector may have potential use in tumor gene therapy with targeted tumor gene silencing effect.', 'http://www.ncbi.nlm.nih.gov/pubmed/17012855', 'no', 'no', 'NA', 1),
(17014383, 'Shogren KL, Turner RT, Yaszemski MJ, Maran A', 'Double-stranded RNA-dependent protein kinase is involved in 2-methoxyestradiol-mediated cell death of osteosarcoma cells.', 'J Bone Miner Res', '2007', 'Double-stranded RNA-dependent protein kinase is involved in 2-methoxyestradiol-mediated cell death of osteosarcoma cells. We studied the involvement of interferon-regulated, PKR on 2-ME-mediated actions in human osteosarcoma cells. Our results show that PKR is activated by 2-ME treatment and is necessary for 2-ME-mediated induction of osteosarcoma cell death.Osteosarcoma is the most common primary bone tumor and most frequently develops during adolescence. 2-Methoxyestradiol (2-ME), a metabolite of 17beta-estradiol, induces interferon gene expression and apoptosis in human osteosarcoma cells. In this report, we studied the role of interferon-regulated double-stranded (ds)RNA-dependent protein kinase (PKR) protein on 2-ME-mediated cell death in human osteosarcoma cells.Western blot analyses were used to measure PKR protein and phosphorylation levels. Cell survival and apoptosis assays were measured using trypan blue exclusion and Hoechst dye methods, respectively. A transient transfection protocol was used to express the dominant negative PKR mutants.PKR was increased in 2-ME-treated MG63 cells, whereas 17beta-estradiol, 4-hydroxyestradiol, and 16alpha-hydroxyestradiol, which do not induce cell death, had no effect on PKR protein levels. Also, 2-ME treatment induced PKR kinase activity as indicated by increased autophosphorylation and phosphorylation of the endogenous substrate, eukaryotic initiation factor (eIF)-2alpha. dsRNA poly (I).poly (C), an activator of PKR protein, increased cell death when osteosarcoma cells were treated with a submaximal concentration of 2-ME. In contrast, a serine-threonine kinase inhibitor SB203580 and a specific PKR inhibitor 2-aminopurine (2-AP) blocked the 2-ME-induced cell death in MG63 cells. A dominant negative PKR mutant protein conferred resistance to 2-ME-induced cell death to MG63 osteosarcoma and 2-ME-mediated PKR regulation did not require interferon gene expression. PKR protein is activated in cell free extracts by 2-ME treatment, resulting in autophosphorylation and in the phosphorylation of the substrate eIF-2alpha. We conclude from these results that PKR is regulated by 2-ME independently of interferon and is essential for 2-ME-mediated cell death in MG63 osteosarcoma cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/17014383', 'no', 'no', '2-methoxyestradiol', 1);


INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(17493842, 'Yan XB, Yang DS, Gao X, Feng J, Shi ZL, Ye Z', 'Caspase-8 dependent osteosarcoma cell apoptosis induced by proteasome inhibitor MG132.', 'Cell Biol Int', '2007', 'Caspase-8 dependent osteosarcoma cell apoptosis induced by proteasome inhibitor MG132. Many researchers have reported that proteasome inhibitors could induce apoptosis in a variety of cancer cells, such as breast cancer cell, lung cancer cell, and lymphoma cell. However, the effect of proteasome inhibitors on osteocsarcoma cells and the mechanisms are seldom studied. In this study, we found proteasome inhibitor MG132 was an effective inducer of apoptosis in human osteosarcoma MG-63 cells. On normal human diploid fibroblast cells, MG132 did not show any apoptosis-inducing effects. Apoptotic changes such as DNA fragment and apoptotic body were observed in MG132-treated cells and MG132 mostly caused MG-63 cell arrest at G(2)-M-phase by cell cycle analysis. Increased activation of caspase-8, accumulation of p27(Kip1), and an increased ratio of Bax:Bcl-2 were detected by RT-PCR and Western blot analysis. Activation of caspase-3 and caspase-9 were not observed. This suggests that the apoptosis induced by MG132 in MG63 cells is caspase-8 dependent, p27 and bcl-2 family related.', 'http://www.ncbi.nlm.nih.gov/pubmed/17493842', 'no', 'no', 'MG132', 1),
(17505002, 'Kersting C, Gebert C, Agelopoulos K, Schmidt H, van Diest PJ, Juergens H, Winkelmann W, Kevric M, Gosheger G, Brandt B, Bielack S, Buerger H', 'Epidermal growth factor receptor expression in high-grade osteosarcomas is associated with a good clinical outcome.', 'Clin Cancer Res', '2007', 'Epidermal growth factor receptor expression in high-grade osteosarcomas is associated with a good clinical outcome. The expression of the epidermal growth factor receptor (EGFR) in osteosarcomas has repeatedly been described. With the introduction of anti-EGFR-targeted therapies in clinical practice, these findings regain increased attention. Experience with anti-EGFR-targeted therapies in other cancers has made clear that besides the expression status of EGFR, a detailed knowledge about gene mutations is of major predictive power. We therefore aimed to explore the EGFR expression and gene mutation status in high-grade osteosarcomas.We investigated tumor samples of osteosarcoma patients of all age groups by means of immunohistochemistry (n=111) and egfr fluorescence in situ hybridization (n=39). Sixty-three patients were treated according to the Cooperative Osteosarcoma Study Group protocols and complete clinical follow-up was available in these cases.Ninety-one of 111 (81%) of osteosarcomas revealed an expression of EGFR. EGFR expression showed a dose-response relation with improved event-free and overall survival. This was independent of the degree of tumor regression due to neoadjuvant chemotherapy. Nine of 39 (23%) osteosarcomas showed egfr amplifications by means of fluorescence in situ hybridization. All these cases expressed EGFR. When comparing EGFR expression between primary biopsy and resection specimen (n=19), viable residual tumor cells in resection specimens revealed a lower EGFR expression and a tendency toward membranous staining compared with the initial biopsy.In conclusion, expression and amplification of EGFR are frequently observed in high-grade osteosarcomas and are associated with improved prognosis in a dose-responsive way. This implies that low EGFR expression possibly predicts lack of response to conventional treatment in high-grade osteosarcomas and may warrant a more intensive therapeutic approach, although not based on EGFR targeting.', 'http://www.ncbi.nlm.nih.gov/pubmed/17505002', 'yes', 'yes', 'NA', 1),
(17506944, 'Guo QC, Shen JN, Jin S, Wang J, Huang G, Zhang LJ, Huang G, Yin JQ, Zou CY, Li MT', 'Comparative proteomic analysis of human osteosarcoma and SV40-immortalized normal osteoblastic cell lines.', 'Acta Pharmacol Sin', '2007', 'Comparative proteomic analysis of human osteosarcoma and SV40-immortalized normal osteoblastic cell lines. Comparative proteomics provide a powerful approach in screening for alterations in protein levels and post-translational modifications that are associated with tumors. In the present study, we aimed to identify candidate biomarkers to distinguish osteosarcoma (OS) cells from normal osteoblastic cells.We employed 3 OS cell lines (U2OS, IOR/OS9, and SaOS-2), and used the SV40-immortalized normal osteoblastic cell line (hFOB1.19) as the control. The differential protein levels in OS and osteoblastic cells were identified using 2-D gel electrophoresis followed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry analyses. Two proteins of interest, the levels of which were significantly increased in OS cells, were further characterized by Western blot analyses.Twenty-six proteins were identified, the expression level of which was either significantly increased or decreased in the OS cells as compared to the control cells. The expression level of the activator of 90 kDa shock protein ATPase homolog 1 (AHA1), was enhanced 12.4-, 24.1-, and 23.8-fold in SaOS-2, IOR/OS9, and U2OS cells, respectively, and the level of the stomatin-like protein 2 (SLP-2) was increased by 10.4- and 7.8-fold in IOR/OS9 and U2OS cells, respectively, as compared to normal osteoblastic cells. Those observations were confirmed by Western blot analyses.A differential proteomic analysis was successfully used to identify AHA1 and SLP-2 that were significantly overproduced in OS cells as compared to normal osteoblastic cells, suggesting that those proteins among others may be effective biomarker candidates for the identification of OS cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/17506944', 'no', 'no', 'NA', 2),
(17506984, 'Maehara H, Kaname T, Yanagi K, Hanzawa H, Owan I, Kinjou T, Kadomatsu K, Ikematsu S, Iwamasa T, Kanaya F, Naritomi K', 'Midkine as a novel target for antibody therapy in osteosarcoma.', 'Biochem Biophys Res Commun', '2007', 'Midkine as a novel target for antibody therapy in osteosarcoma. Osteosarcoma is a malignant tumor with poor prognosis, and lack of accurate prognostic factors is one of the reasons that make this tumor difficult to cure. The heparin-binding growth factor, midkine is involved in generation and progression of many types of tumors. However, the relationship between midkine and osteosarcoma has been unclear. We show here that midkine is overexpressed in osteosarcoma and the level of midkine expression is correlated with prognosis (P<0.05; log-rank test). Treatment with functional antibodies against midkine suppresses growth of osteosarcoma cell lines, 9N2, 3N1, Saos-2, and NOS-1, to 25-65% of untreated controls. Our results suggest that midkine is useful as a prognostic marker, and is a candidate therapeutic target for osetosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/17506984', 'yes', 'yes', 'NA', 1),
(17509570, 'Daubie V, De Decker R, Nicaise C, Pochet R', 'Osteosarcoma cell-calcium signaling through tissue factor-factor VIIa complex and factor Xa.', 'FEBS Lett', '2007', 'Osteosarcoma cell-calcium signaling through tissue factor-factor VIIa complex and factor Xa. The cells responsible for bone formation express protease-activated receptors. Although serine protease thrombin has been shown to elicit functional responses in bone cells that impact on cell survival and alkaline phosphatase activity, nothing is known about tissue factor, factor VIIa, and factor Xa, the serine proteases that act upstream of thrombin in the coagulation cascade. This paper demonstrates that tissue factor is expressed in the osteoblast-like cell line SaOS-2 and, that tissue factor in a factor VIIa-bound complex induces a transient intracellular Ca(2+) increase through protease-activated receptor-2. In SaOS-2 cells, factor Xa induced a sustained intracellular Ca(2+) response, as does SLIGRL, a PAR2-activating peptide, and PAR-1-dependent cell viability.', 'http://www.ncbi.nlm.nih.gov/pubmed/17509570', 'no', 'no', 'NA', 3),
(17509661, 'Wen YH, Koeppen H, Garcia R, Chiriboga L, Tarlow BD, Peters BA, Eigenbrot C, Yee H, Steiner G, Greco MA', 'Epidermal growth factor receptor in osteosarcoma: expression and mutational analysis.', 'Hum Pathol', '2007', 'Epidermal growth factor receptor in osteosarcoma: expression and mutational analysis. Activation of intracellular mitogenic signal transduction pathways driven by the ErbB family of receptor tyrosine kinases has been implicated in the development and/or progression of a variety of cancers. Studies on ErbB receptors in osteosarcoma have focused on HER-2 and have produced conflicting results with few studies evaluating the expression of the epidermal growth factor receptor (EGFR). In this study, we determined the level of expression of EGFR and the mutational status of the EGFR receptor in a subset of osteosarcoma tumor samples as well as in a series of established bone tumor-derived cell lines. EGFR protein expression was detected in the form of strong membranous staining by immunohistochemistry in 21 (57%) of 37 cases analyzed. Six of 12 (50%) osteosarcoma cell lines revealed moderate to high expression levels of EGFR. Two somatic alterations (E829E and R831C) were identified in the cytoplasmic domain of the EGFR gene in 1 of 10 tumor samples. The significance of these findings for the pathobiology of osteosarcomas will be investigated further.', 'http://www.ncbi.nlm.nih.gov/pubmed/17509661', 'no', 'no', 'NA', 1),
(17513610, 'Huang G, Mills L, Worth LL', 'Expression of human glutathione S-transferase P1 mediates the chemosensitivity of osteosarcoma cells.', 'Mol Cancer Ther', '2007', 'Expression of human glutathione S-transferase P1 mediates the chemosensitivity of osteosarcoma cells. Chemoresistance is a major reason that patients with osteosarcoma fail to achieve a lasting chemotherapy response, and it contributes to disease relapse, progression, and death. Human glutathione S-transferase P1 (GSTP1), a phase II detoxification enzyme, contributes to chemoresistance in many cancers. However, the role of GSTP1 in osteosarcoma chemoresistance is ill defined. We hypothesized that GSTP1 has cytoprotective effects in human osteosarcoma. To assess this possibility, we used GSTP1 cDNA transfection or RNA interference to overexpress or suppress GSTP1 in osteosarcoma cells, and assessed the cytotoxic effect of chemotherapeutic agents on these cells. Our results showed that GSTP1 expression was up-regulated in osteosarcoma cells when they were treated with doxorubicin or cisplatin. GSTP1 overexpression in SAOS-2 osteosarcoma cells caused the cells to be more resistant to doxorubicin and cisplatin. In contrast, GSTP1 suppression in HOS cells caused more apoptosis and extensive DNA damage in response to doxorubicin and cisplatin. The cytotoxicity assay also showed that GSTP1 suppression caused a 2.5-fold increase in cell growth inhibition resulting from doxorubicin and cisplatin treatments [the IC(50)s are approximately 0.16 micromol/L (doxorubicin) and 1.8 micromol/L (cisplatin) for parental HOS versus 0.06 micromol/L (doxorubicin) and 0.75 micromol/L (cisplatin) for GSTP1-silenced HOS]. Moreover, GSTP1 suppression decreased the activation of extracellular signal-regulated kinase 1/2, which is induced by cisplatin and doxorubicin. Taken together, these findings show that GSTP1 contributes to doxorubicin and cisplatin resistance in osteosarcoma, which may be mediated in part by the activation of extracellular signal-regulated kinase 1/2. Targeting of GSTP1 combined with chemotherapy may have synergistic therapeutic effects on osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/17513610', 'no', 'no', 'cisplatin|doxorubicin', 3),
(17520676, 'Li Y, Tanaka K, Li X, Okada T, Nakamura T, Takasaki M, Yamamoto S, Oda Y, Tsuneyoshi M, Iwamoto Y', 'Cyclin-dependent kinase inhibitor, flavopiridol, induces apoptosis and inhibits tumor growth in drug-resistant osteosarcoma and Ewing''s family tumor cells.', 'Int J Cancer', '2007', 'Cyclin-dependent kinase inhibitor, flavopiridol, induces apoptosis and inhibits tumor growth in drug-resistant osteosarcoma and Ewing''s family tumor cells. Multimodal therapies play important roles in the treatment of osteosarcoma (OS) and Ewing''s family of tumors (EFTs), two most frequent malignant bone tumors. Although the clinical outcome of primary OS and EFTs is greatly improved, the relapsed cases often are associated with multidrug resistance of the tumors and the prognosis of these patients is still poor. Flavopiridol, a pan cyclin-dependent kinase (CDK) inhibitor is a novel antitumor agent that can induce cell cycle arrest and apoptosis in many cancer cells. However, there have been no studies about the effects of flavopiridol on drug-resistant OS and EFTs. Here, we demonstrated that flavopiridol induced the cleavage of poly-ADP-ribose polymerase (PARP) in a time and dose dependent manner in adriamycin-resistant OS and EFTs cells expressing P-glycoprotein (P-gp) and multidrug resistance-associated protein 1 (MRP(1)) as effectively as in their parental cells. Our data also showed that flavopiridol caused the release of mitochondrial cytochrome c and the activation of caspase-9, caspase-8 and caspase-3, with an increase ratio of the proapoptotic protein level (Bax) to the antiapoptotic protein level (Bcl-2 and Bcl-X(L)), while apoptosis was inhibited by pan caspase inhibitor (Z-VAD-FMK) and caspase-3 inhibitor (Z-DEVD-FMK), not by caspase-8 inhibitor (Z-IETD-FMK). The treatment with flavopiridol further inhibited the tumor growth in mouse models of the drug-resistant OS and EFTs. These results suggest that flavopiridol might be promising in clinical therapy for the relapsed OS and EFTs.(c) 2007 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/17520676', 'no', 'no', 'flavopiridol', 9),
(17536311, 'Masi L, Recenti R, Silvestri S, Pinzani P, Pepi M, Paglierani M, Brandi ML, Franchi A', 'Expression of cyclooxygenase-2 in osteosarcoma of bone.', 'Appl Immunohistochem Mol Morphol', '2007', 'Expression of cyclooxygenase-2 in osteosarcoma of bone. Several studies indicate that cyclooxygenase-2 (COX-2) is overexpressed in human malignancies, where it produces high levels of prostaglandins and contributes to tumor growth. In this study we have analyzed the expression of COX-2 in a series of 48 skeletal osteosarcomas of different subtypes by immunohistochemistry. In addition, we examined the effects of the specific COX-2 inhibitor Celecoxib on the growth of the human osteosarcoma cell line SaOS-2. Immunoreactivity for COX-2 was observed in 39 out of 48 tumors (81.2%), 30 (76.9%) of which showed a moderate or diffuse immunostaining. Considering the group of 42 primary osteosarcomas, COX-2 immunoreactivity was significantly higher in high grade osteosarcomas, where moderate or diffuse expression was detected in 23 out of 32 cases (71.8%), than in low grade osteosarcomas, where moderate or diffuse expression was detected in 2 out of 10 cases (20%) (P = 0.008, Fisher exact test). In addition, low COX-2 expression was always associated with a good response to chemotherapy (5 out of 5 cases), whereas moderate or diffuse COX-2 expression was associated with a good response in 11 out of 20 cases (55%) (P = 0.12, Fisher exact test). In SaOS-2 osteosarcoma cells, which express COX-2, treatment with Celecoxib determined inhibition of cell proliferation and induction of apoptosis. These results indicate that COX-2 is expressed at high levels in high grade osteosarcomas and support the use of COX-2 inhibitors to improve both the tumor response to chemotherapy and the outcome of osteosarcoma patients.', 'http://www.ncbi.nlm.nih.gov/pubmed/17536311', 'yes', 'yes', 'celecoxib', 1),
(17549480, 'Sakayama K, Mashima N, Kidani T, Miyazaki T, Yamamoto H, Masuno H', 'Effect of cortisol on cell proliferation and the expression of lipoprotein lipase and vascular endothelial growth factor in a human osteosarcoma cell line.', 'Cancer Chemother Pharmacol', '2008', 'Effect of cortisol on cell proliferation and the expression of lipoprotein lipase and vascular endothelial growth factor in a human osteosarcoma cell line. The aim of this study is to investigate whether cortisol inhibited cell proliferation and the expressions of lipoprotein lipase (LPL), a key enzyme involved in the energy metabolism in tumor cells, and vascular endothelial growth factor (VEGF), a potent angiogenic factor in the tumor, in cultures of OST cells, a human osteosarcoma cell line.OST cells were treated for 48 h with or without cortisol. To examine the effect of cortisol on cell proliferation, the expression of proliferating cell nuclear antigen (PCNA) was examined by Western blotting, and the amount of (3)H-thymidine incorporated into DNA during the last 30 min of the 48-h treatment period was measured. To examine the effect of cortisol on the expression of LPL, the activity and mass of LPL were measured in the extract of acetone/ether powder of cells, and the amount of (35)S-methionine incorporated into LPL during the last 2 h of the 48-h treatment period was measured by immunoprecipitation. The expression of VEGF was examined by immunohistochemistry and Western blotting.The amount of (3)H-thymidine incorporated into DNA and the level of PCNA were lower in the cortisol-treated cultures than in the untreated cultures, thus indicating that cortisol inhibited the proliferation of OST cells. The synthetic rate and activity of LPL were lower in the cortisol-treated cultures than in the untreated cultures but no difference in the specific activity of LPL between the two cultures was observed, thus indicating that cortisol inhibited LPL synthesis, thereby resulting in a decreased LPL activity. The expression of VEGF was lower in the cortisol-treated cultures than in the untreated cultures.Cortisol not only has the ability to inhibit cell proliferation but also the ability to inhibit the expressions of LPL and VEGF in cultures of OST cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/17549480', 'no', 'no', 'cortisol', 2),
(17560052, 'Fuchs B, Mahlum E, Halder C, Maran A, Yaszemski M, Bode B, Bolander M, Sarkar G', 'High expression of tumor endothelial marker 7 is associated with metastasis and poor survival of patients with osteogenic sarcoma.', 'Gene', '2007', 'High expression of tumor endothelial marker 7 is associated with metastasis and poor survival of patients with osteogenic sarcoma. Our objective is to identify genes regulating metastasis of osteogenic sarcoma (OGS) since metastasis is the primary cause of mortality among patients with OGS. To identify such genes, we first created a database of differentially expressed genes between six low-grade and six high-grade OGS tumors, and between a normal immortalized osteoblast cell line (FOB) and four commercially available OGS-derived cell lines. We specifically searched for surface proteins over-expressed in high-grade OGS, since we hypothesize that tumor-cell specific surface markers are key to metastasis. A gene encoding Tumor Endothelial Marker7 (TEM7) was selected as a candidate for further study. TEM7 expression pattern was assessed by RT-PCR, Western blotting and immunostaining. TEM7 mRNA was abundantly expressed in SAOS cells (derived from high-grade OGS), but not in FOB or MG63 cells (derived from low-grade OGS). Virtually no expression of TEM7 protein was observed in FOB cells but abundant expression was noted in SAOS and TE85 cells. Employing immunostaining of 92 human OGS specimens (50 high-grade and 42 low-grade) collected before chemotherapy show 97% (37 of 38) of high-grade OGS specimens with metastasis have high TEM7 staining. Further, we found that elevated expression of TEM7 correlated with poor survival (p<0.04) of affected patients. Inhibiting TEM7 function by siRNA inhibited invasion and migration of OGS cells with metastatic potential. Our results suggest TEM7 expression level closely parallels histology-based prognostication of OGS metastasis and, therefore, it is a therapeutic target. This is the first demonstration of a link between TEM7 and cancer metastasis.', 'http://www.ncbi.nlm.nih.gov/pubmed/17560052', 'yes', 'yes', 'NA', 1),
(17568187, 'Oshima Y, Sasaki Y, Negishi H, Idogawa M, Toyota M, Yamashita T, Wada T, Nagoya S, Kawaguchi S, Yamashita T, Tokino T', 'Antitumor effect of adenovirus-mediated p53 family gene transfer on osteosarcoma cell lines.', 'Cancer Biol Ther', '2007', 'Antitumor effect of adenovirus-mediated p53 family gene transfer on osteosarcoma cell lines. Osteosarcoma (OS) is one of the most common malignancies of the bone. Although prognosis of OS has improved significantly during the past several years due to more intensive chemotherapy and radiotherapy regimens, new therapeutic approaches are needed for recurrent and inoperable cases, p73 and p63, like their homologue, the tumor suppressor p53, are able to induce apoptosis in several cell types. Here, we evaluated the antitumor effects of p73 and p63 on eleven different human OS cell lines. In vitro, adenovirus-mediated transduction of p63gamma induced apoptosis in OS cells that are resistant to p53-mediated apoptosis, while less effect was observed following transduction of p73alpha or p63alpha. Interestingly, the apoptotic effects of p63gamma were greater than those of wild-type p53 in OS cells carrying MDM2-amplification. We then determined the in vivo therapeutic effect of intratumoral injection of adenovirus-vector expressing p53 family members on xenografts derived from Saos-2 cells implanted in nude mice, and showed that infection with p63y significantly suppressed tumor growth compared with p53. In addition, exogenous p73beta and p63gamma significantly increased the chemosensitivity of OS cells to doxorubicin and cisplatin, chemotherapeutic agents commonly used in the treatment of OS. Our results suggest that adenovirus-mediated transduction of p53 family members may have utility in gene therapy for OS, particularly in combination with chemotherapeutic agents.', 'http://www.ncbi.nlm.nih.gov/pubmed/17568187', 'no', 'no', 'doxorubicin|cisplatin', 4),
(17569615, 'Zhang Y, Howell RD, Alfonso DT, Yu J, Kong L, Wittig JC, Liu CJ', 'IFI16 inhibits tumorigenicity and cell proliferation of bone and cartilage tumor cells.', 'Front Biosci', '2007', 'IFI16 inhibits tumorigenicity and cell proliferation of bone and cartilage tumor cells. IFI16 is a member of the interferon-inducible p200-protein family, capable of modulating cell proliferation, and cellular senescence. In this study, these effects of IFI16 were studied in tumor cells derived from bone and cartilage. The level of IFI16 was markedly lower in human osteosarcomas as compared with its level in normal bone. Overexpression of functional IFI16 in human osteosarcoma and chondrosarcoma cell lines markedly inhibited colony formation, and significantly inhibited cell growth, an effect that could be reversed by introduction of gene specific siRNA into tumor cells. These inhibitory effects of IFI16 were associated with upregulation of p21 and inhibition of cyclin E, cyclin D1, c-Myc and Ras. In addition, ectopic expression of IFI16 in tumor cells increased senescence-associated beta-galactosidase and induced a senescence-like phenotype. In view of such effects, IFI16 might be a suitable target for therapeutic intervention in osteosarcoma and chondrosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/17569615', 'no', 'no', 'NA', 6),
(17598902, 'Chen CL, Loy A, Cen L, Chan C, Hsieh FC, Cheng G, Wu B, Qualman SJ, Kunisada K, Yamauchi-Takihara K, Lin J', 'Signal transducer and activator of transcription 3 is involved in cell growth and survival of human rhabdomyosarcoma and osteosarcoma cells.', 'BMC Cancer', '2007', 'Signal transducer and activator of transcription 3 is involved in cell growth and survival of human rhabdomyosarcoma and osteosarcoma cells. Stat3 has been classified as a proto-oncogene and constitutive Stat3 signaling appears to be involved in oncogenesis of human cancers. However, whether constitutive Stat3 signaling plays a role in the survival and growth of osteosarcomas, rhabdomyosarcomas, and soft-tissue sarcomas is still unclear.To examine whether Stat3 is activated in osteosarcomas, rhabdomyosarcomas and other soft-tissue sarcomas we analyzed sarcoma tissue microarray slides and sarcoma cell lines using immunohistochemistry and Western blot analysis, respectively, with a phospho-specific Stat3 antibody. To examine whether the activated Stat3 pathway is important for sarcoma cell growth and survival, adenovirus-mediated expression of a dominant-negative Stat3 (Y705F) and a small molecule inhibitor (termed STA-21) were used to inhibit constitutive Stat3 signaling in human sarcoma cell lines expressing elevated levels of Stat3 phosphorylation. Cell viability was determined by MTT assays and induction of apoptosis was analyzed by western blotting using antibodies that specifically recognize cleaved caspases-3, 8, and 9.Stat3 phosphorylation is elevated in 19% (21/113) of osteosarcoma, 27% (17/64) of rhabdomyosarcoma, and 15% (22/151) of other soft-tissue sarcoma tissues as well as in sarcoma cell lines. Expression of the dominant-negative Stat3 and treatment of STA-21 inhibited cell viability and growth and induced apoptosis through caspases 3, 8 and 9 pathways in human sarcoma cell lines expressing elevated levels of phosphorylated Stat3.This study demonstrates that Stat3 phosphorylation is elevated in human rhabdomyosarcoma, osteosarcomas and soft-tissue sarcomas. Furthermore, the activated Stat3 pathway is important for cell growth and survival of human sarcoma cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/17598902', 'yes', 'no', 'NA', 1),
(17609619, 'Ogino W, Takeshima Y, Mori T, Yanai T, Hayakawa A, Akisue T, Kurosaka M, Matsuo M', 'High level of ezrin mRNA expression in an osteosarcoma biopsy sample with lung metastasis.', 'J Pediatr Hematol Oncol', '2007', 'High level of ezrin mRNA expression in an osteosarcoma biopsy sample with lung metastasis. Osteosarcoma (OS) remains a life-threatening malignancy and its molecular character is not fully understood. Ezrin is a cytoskeleton linker protein involved in regulating the growth and metastatic capacity of cancer cells. However, the correlation between ezrin mRNA expression and clinical severity has not yet been examined in OS biopsy samples. Furthermore, recent evidence has demonstrated that the level of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression is increased in human cancers of various origins, but this has not yet been examined in OS cells. To clarify the correlation between the clinical severity and the levels of ezrin and GAPDH mRNA expression, we quantified these mRNA levels in 4 pediatric OS biopsy samples using real-time quantitative polymerase chain reaction. Among these 4 samples, ezrin mRNA expression was approximately 5-fold higher in a case with lung metastasis compared with the other cases without metastasis, suggesting an association between the ezrin mRNA expression level and metastasis. On the other hand, the GAPDH mRNA expression level was not related to the clinical severity. This is the first report to demonstrate a high level of ezrin mRNA expression in an OS biopsy sample with lung metastasis.', 'http://www.ncbi.nlm.nih.gov/pubmed/17609619', 'yes', 'yes', 'NA', 1),
(17611411, 'Yuan XW, Zhu XF, Huang XF, Sheng PY, He AS, Yang ZB, Deng R, Feng GK, Liao WM', 'P14ARF sensitizes human osteosarcoma cells to cisplatin-induced apoptosis in a p53-independent manner.', 'Cancer Biol Ther', '2007', 'P14ARF sensitizes human osteosarcoma cells to cisplatin-induced apoptosis in a p53-independent manner. The tumor suppressor p14ARF, encoded by the INK4a/ARF locus, is often disrupted in human cancers, p14ARF triggers cell cycle arrest and sensitizes cells to apoptosis in the presence of collateral signals. To investigate the role of p14ARF in chemotherapeutic drugs-induced apoptosis, p14ARF was overexpressed by stable transfection in human osteosarcoma cell lines, U2OS (p53-wt/p14ARF-null) and MG63 (p53-mt/p14ARF-null). The results showed that ectopic p14ARF sensitized both cell lines to cisplatin-induced cytotoxicity and apoptosis. This sensitization of cisplatin-induced apoptosis was associated with upregulation of p53, Bax and p21 in U2OS cells. Conversely, such a result was not observed in MG63 cells. Moreover, the sensitization of cisplatin-induced cytotoxicity in U2OS cells was unaltered by p53 siRNA. Together, we show here p14ARF sensitizes human osteosarcoma cells to cisplatin-induced apoptosis in a p53-independent manner. Proper combinations of p14ARF gene transfer and conventional chemotherapy may be a valuable strategy in human osteosarcoma treatment.', 'http://www.ncbi.nlm.nih.gov/pubmed/17611411', 'no', 'no', 'cisplatin', 4),
(17611684, 'Iguchi T, Miyakawa Y, Saito K, Nakabayashi C, Nakanishi M, Saya H, Ikeda Y, Kizaki M', 'Zoledronate-induced S phase arrest and apoptosis accompanied by DNA damage and activation of the ATM/Chk1/cdc25 pathway in human osteosarcoma cells.', 'Int J Oncol', '2007', 'Zoledronate-induced S phase arrest and apoptosis accompanied by DNA damage and activation of the ATM/Chk1/cdc25 pathway in human osteosarcoma cells. Osteosarcoma is one of the most common primary malignant tumors of the bone in children and adolescents. Some patients continue to have a poor prognosis, as they have metastatic disease and frequent occurrence of drug resistance. Zoledronate is a nitrogen-containing bisphosphonate that has been used for the treatment of hypercalcemia and bone metastasis, because it induces apoptosis in osteoclasts and tumor cells by inhibiting the isoprenylation of intracellular small G proteins. Besides inhibiting isoprenylation, little is known about the manner by which bisphosphonates inhibit cellular proliferation and induce apoptosis. This prompted us to investigate the inhibitory effects of zoledronate in human osteosarcoma cell lines, HOS and MG63. HOS cells accumulated in S phase around 6 h after treatment with 10 microM zoledronate, followed by apoptosis. When HOS cells were treated with zoledronate, ATM kinase and its substrate, check-point kinase (Chk)1, were phosphorylated. Zoledronate also induced phosphorylation of cdc25a (Thr506) in HOS cells, which is a substrate of Chk1, and its phosphorylation is known to be critical for S phase arrest. Following treatment with zoledronate, phosphorylated histone H2AX (gamma-H2AX) displayed patterns of nuclear foci in HOS cells. As gamma-H2AX accumulates at dsDNA breaks, these results demonstrate that zoledronate induced DNA damage and S phase arrest, accompanied by activation of the ATM/Chk1/cdc25 pathway in a human osteosarcoma cell line.', 'http://www.ncbi.nlm.nih.gov/pubmed/17611684', 'no', 'no', 'Zoledronate', 4),
(17611696, 'Tanaka H, Yoshida K, Okamura H, Morimoto H, Nagata T, Haneji T', 'Calyculin A induces apoptosis and stimulates phosphorylation of p65NF-kappaB in human osteoblastic osteosarcoma MG63 cells.', 'Int J Oncol', '2007', 'Calyculin A induces apoptosis and stimulates phosphorylation of p65NF-kappaB in human osteoblastic osteosarcoma MG63 cells. Previously, we reported that okadaic acid, a specific inhibitor of serine/threonine protein phosphatases, induced apoptosis in human osteoblastic cells. However, it is not clear whether calyculin A, another inhibitor of protein phosphatases, would induce apoptosis in human osteoblastic cells and if so, which mechanisms are involved and whether the phosphorylation status of NF-kappaB could be affected by the treatment with calyculin A. In this report, we demonstrate that calyculin A induced apoptosis in MG63 cells, as judged by WST-8 assay, nuclear fragmentation, and DNA ladder formation. Expression of PTEN, FasL, and FasR mRNA was stimulated by calyculin A treatment in MG63 cells. Calyculin A also enhanced the phosphorylation level of NF-kappaB, as judged from the results of Western blot analysis and an in vitro dephosphorylation assay. Western blot analysis with anti-phospho-p65NF-kappaB antibody disclosed that the NF-kappaB was phosphorylated on serine 536 in cytosol and translocated into nucleus with calyculin A-treatment. The phosphorylation status of p65NF-kappaB was further confirmed by using the phosphorylation site-mutated p65NF-kappaB gene transfected into HEK293 cells. Unlike TNF-alpha, calyculin A treatment did not degraded IkappaBalpha within 10 min, while it degraded IkappaBalpha at 2-h treatment. Our findings indicate that calyculin A elicit phosphorylation of NF-kappaB on serine 536 in MG63 cells, resulting in the translocation of phospho-NF-kappaB to the nucleus, thereby promoting transcriptional activity of NF-kappaB-related genes.', 'http://www.ncbi.nlm.nih.gov/pubmed/17611696', 'no', 'no', 'calyculin A', 6),
(17627080, 'Kalariti N, Lembessis P, Papageorgiou E, Pissimissis N, Koutsilieris M', 'Regulation of the mGluR5, EAAT1 and GS expression by glucocorticoids in MG-63 osteoblast-like osteosarcoma cells.', 'J Musculoskelet Neuronal Interact', 'NA', 'Regulation of the mGluR5, EAAT1 and GS expression by glucocorticoids in MG-63 osteoblast-like osteosarcoma cells. Growth factors, cytokines, sex steroid hormones and glucocorticoids have differential and complex effects on skeletal metabolism. Recently, the presence of the glutamatergic (Glu) system in bone cells has provided new evidence for its possible role in bone physiology. Consequently, we have investigated the regulation of certain components of the Glu system by glucocorticoids in MG-63 osteoblast-like osteosarcoma cells, in vitro.We characterized the effects of dexamethasone on the expression of the mGluR5, EAAT1 and GS, at mRNA and protein level, using relative quantitative RTPCR and Western blot analysis, respectively.We confirmed the induction of GS expression by dexamethasone published previously. In addition, we documented for the first time the expression of the mGluR5 and EAAT1 in MG-63 cells, as well as the ability of dexamethasone to upregulate the expression of the mGluR5 and EAAT1 in the MG-63 cells.Components of the glutamatergic system may play a role in bone pathophysiology.', 'http://www.ncbi.nlm.nih.gov/pubmed/17627080', 'no', 'no', 'dexamethasone', 2),
(17632732, 'Nedelcu T, Kubista B, Koller A, Sulzbacher I, Mosberger I, Arrich F, Trieb K, Kotz R, Toma CD', 'Livin and Bcl-2 expression in high-grade osteosarcoma.', 'J Cancer Res Clin Oncol', '2008', 'Livin and Bcl-2 expression in high-grade osteosarcoma. The evaluation of prognosis in patients with osteosarcoma is limited to clinical parameters. Although numerous molecular markers have been studied, none are currently in routine clinical use. The aim of this study was to determine if Livin and Bcl-2, acting as antiapoptotic proteins through different mechanisms, are expressed in osteosarcoma, and whether they can be used as prognostic markers in human osteosarcoma.Tumor specimens of 29 patients with high-grade central osteosarcoma, with complete clinical follow-up for a minimum of 5 years, were studied. The localization and distribution of Livin and Bcl-2 were investigated using immunohistochemistry. Results were correlated with the histological response to chemotherapy, 5-year disease-free and 5-year overall survival.Bcl-2 was expressed only in the cytoplasm of 16/29 cases and there was no statistically significant correlation between expression and any of the studied parameters. Livin was detected in 17/29 cases, in the cytoplasm of all 17 and in the nucleus of only 3 cases. Nuclear expression was significantly correlated with a decreased overall survival (P < 0.0002) compared with those patients without nuclear expression.The results of this study indicate that Bc1-2 and Livin are highly expressed in osteosarcoma cells and that possibly, the evaluation of nuclear Livin expression might be a useful prognostic marker in osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/17632732', 'yes', 'yes', 'NA', 2),
(17645699, 'Toma CD, Svoboda M, Arrich F, Ekmekcioglu C, Assadian O, Thalhammer T', 'Expression of the melatonin receptor (MT) 1 in benign and malignant human bone tumors.', 'J Pineal Res', '2007', 'Expression of the melatonin receptor (MT) 1 in benign and malignant human bone tumors. The beneficial effects of melatonin on bone homeostasis have been shown in various diseases. As this indoleamine causes dose-dependent modulation of bone-forming osteoblast and bone-resorbing osteoclast activities by receptor-independent and -dependent pathways, we investigated the expression of G-protein-coupled melatonin receptors (MTs) in malignant and non-malignant human bone lesions. By TaqMan polymerase chain reaction (PCR), we analyzed 30 specimens from osteosarcoma and 11 from benign bone tumors for MT1-mRNA expression. Furthermore, we determined mRNA expression levels of the osteoclast activity-stimulating receptor activator of nuclear factor-kappa B ligand (RANKL) and its counterpart osteoprotegerin (OPG). Although mean MT1-mRNA levels were similar (P = 0.596) in malignant (4.39 +/- 4.98-fold) and benign samples (4.64 +/- 6.81-fold), the highest MT1-mRNA levels (up to 27-fold) were observed in individual osteosarcomas, particularly, in two specimens of patients with local recurrence of the tumor. Moreover, mean RANKL- and OPG-mRNA levels were similar in malignant and benign specimens (RANKL: 7.38 +/- 9.61-fold versus 3.57 +/- 3.11-fold, P = 0.207; OPG: 23.45 +/- 32.76 versus 8.07 +/- 7.23-fold, P = 0.133). Again, highest RANKL- and OPG-mRNA levels (up to 41- and 160-fold, respectively) were observed in individual osteosarcomas. Expression of MT1-mRNA was confirmed in two human osteosarcoma cell lines (HOS, MG63). High expression levels of MT1-mRNA together with low OPG-mRNA were found in both osteosarcoma cell lines, while in normal human osteoblasts and bone marrow stromal cells, high OPG-mRNA levels were associated with low MT1-mRNA levels. These data on the abundant expression of MT1-mRNA in human bone tumors and osteosarcoma cells lines suggest an important role for MT1 in bone pathology.', 'http://www.ncbi.nlm.nih.gov/pubmed/17645699', 'yes', 'no', 'NA', 3),
(17660802, 'Fellenberg J, Bernd L, Delling G, Witte D, Zahlten-Hinguranage A', 'Prognostic significance of drug-regulated genes in high-grade osteosarcoma.', 'Mod Pathol', '2007', 'Prognostic significance of drug-regulated genes in high-grade osteosarcoma. About 25-45% of patients with high-grade osteosarcoma poorly respond to chemotherapy with an increased risk of relapse and the development of metastasis. Therefore, the aim of this study was the evaluation of the prognostic value of eight previously identified drug-regulated candidate genes on osteosarcoma therapy outcome. Gene expression of 8 candidate genes was analyzed in 35 formalin-fixed, paraffin-embedded, laser-microdissected osteosarcoma biopsies. The prognostic value of these genes was evaluated by the correlation of gene expression with therapy outcome, overall survival and event-free survival in univariate and multivariate analysis. Upon univariate analysis, the expression of MALAT-1, IMPDH2, FTL and RHOA significantly correlated with response to chemotherapy. Expression of all four genes was increased in the poor responder group. Upon multivariate analysis, IMPDH2 maintained its independent prognostic value (P=0.025). Concerning the overall survival of the patients, we observed a significant association with the expression of FTL, PHB, ATAD2, ACTN1 and RRM2 as well as lactate dehydrogenase serum levels. In the subgroups of patients with high expression of these genes and those with elevated lactate dehydrogenase levels, the mean overall survival was decreased 1.7-, 1.9-, 2.2-, 2.4-, 1.5- and 4.5-fold, respectively. Except RRM2, all genes and lactate dehydrogenase serum levels remained significant in the multivariate analysis. In addition, the event-free survival was significantly decreased in the subgroups of patients with high FTL, ATAD2 and IMPDH2 expression (1.8-, 6.3- and 2.4-fold, respectively). These data demonstrate that among the identified genes are valuable markers for the prediction of osteosarcoma therapy outcome. Especially IMPDH2 and FTL are promising candidates for the stratification of osteosarcoma patients into low- and high-risk groups. Owing to their involvement in drug action these genes may further be potential targets for the modulation of drug sensitivity.', 'http://www.ncbi.nlm.nih.gov/pubmed/17660802', 'yes', 'yes', 'NA', 9),
(17682054, 'Lu NZ, Collins JB, Grissom SF, Cidlowski JA', 'Selective regulation of bone cell apoptosis by translational isoforms of the glucocorticoid receptor.', 'Mol Cell Biol', '2007', 'Selective regulation of bone cell apoptosis by translational isoforms of the glucocorticoid receptor. Glucocorticoids are widely used in the treatment of inflammatory and other diseases. However, high-dose or chronic administration often triggers troublesome side effects such as metabolic syndrome and osteoporosis. We recently described that one glucocorticoid receptor gene produces eight translational glucocorticoid receptor isoforms that have distinct gene-regulatory abilities. We show here that specific, but not all, glucocorticoid receptor isoforms induced apoptosis in human osteosarcoma U-2 OS bone cells. Whole human genome microarray analysis revealed that the majority of the glucocorticoid target genes were selectively regulated by specific glucocorticoid receptor isoforms. Real-time PCR experiments confirmed that proapoptotic enzymes necessary for cell death, granzyme A and caspase-6, were induced by specific glucocorticoid receptor isoforms. Chromatin immunoprecipitation assays further suggested that glucocorticoid receptor isoform-dependent induction of proapoptotic genes was likely due to selective coregulator recruitment and chromatin modification. Interestingly, the capabilities to transrepress proinflammatory genes were similar among glucocorticoid receptor isoforms. Together, these findings provide new evidence that translational glucocorticoid receptor isoforms can elicit distinct glucocorticoid responses and may be useful for the development of safe glucocorticoids with reduced side effects.', 'http://www.ncbi.nlm.nih.gov/pubmed/17682054', 'no', 'no', 'NA', 3),
(17683065, 'Husmann K, Muff R, Bolander ME, Sarkar G, Born W, Fuchs B', 'Cathepsins and osteosarcoma: Expression analysis identifies cathepsin K as an indicator of metastasis.', 'Mol Carcinog', '2008', 'Cathepsins and osteosarcoma: Expression analysis identifies cathepsin K as an indicator of metastasis. Osteosarcoma is the most frequent malignant bone tumor with a poor survival rate for patients with metastasis. Previous studies have shown that beside other proteases, distinct sets of cathepsins are involved in the process of metastasis of different tumors. In this study we investigated the expression of cathepsin proteases in human osteosarcoma metastasis. First, the mRNA expression of 14 human cathepsins was studied in SAOS-2 osteosarcoma cells and the highly metastatic LM5 and LM7 sublines by reverse transcriptase (RT)-polymerase chain reaction (PCR). The expression of cathepsin D, K, and L mRNA was found upregulated and that of cathepsin F, H, and V downregulated in the highly metastatic LM5 and LM7 cells. A subgroup of the cathepsin proteases was further studied at the protein level by Western blot analysis of cell extracts. The expression of cathepsin B and H was decreased and that of cathepsin D, K, and L was increased in the highly metastatic cell lines as compared to the SAOS-2 cell line. Diagnostic relevance of cathepsin K expression in osteosarcoma was revealed upon correlation of survival and metastasis with immunohistochemical cathepsin K staining of biopsies collected from 92 patients prior to chemotherapy. Patients with metastatic high-grade osteosarcoma and low cathepsin K expression at diagnosis had a better prognosis than those with high expression. Thus, it appears that cathepsin K expression is of predictive prognostic value for patients with high-grade tumors and metastasis at diagnosis.(c) 2007 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/17683065', 'yes', 'yes', 'NA', 3),
(17684144, 'Savage SA, Woodson K, Walk E, Modi W, Liao J, Douglass C, Hoover RN, Chanock SJ', 'Analysis of genes critical for growth regulation identifies Insulin-like Growth Factor 2 Receptor variations with possible functional significance as risk factors for osteosarcoma.', 'Cancer Epidemiol Biomarkers Prev', '2007', 'Analysis of genes critical for growth regulation identifies Insulin-like Growth Factor 2 Receptor variations with possible functional significance as risk factors for osteosarcoma. Osteosarcoma, the most common malignant primary bone tumor, typically occurs during the adolescent growth spurt. Germ-line genetic variation in genes critical in growth regulation could confer altered risk of osteosarcoma.Fifty-two common single nucleotide polymorphisms (SNP) in 13 genes were genotyped in a prospective case-control study of osteosarcoma (104 osteosarcoma cases and 74 orthopedic controls). Genotype data analyzed with contingency tables suggested the strongest association with insulin-like growth factor 2 receptor (IGF2R) SNPs. Additional SNPs were genotyped to capture IGF2R common haplotypes and resequencing was done across the IGF2R block associated with osteosarcoma risk. Percentage methylation was determined by pyrosequencing of the IGF2R variant allele located in a CpG island.IGF2R Ex16+88G>A (rs998075) and IVS16+15C>T (rs998074) SNPs were associated with increased risk for osteosarcoma compared with orthopedic controls (haplotype odds ratio, 2.04; 95% confidence interval, 1.29-3.24). Follow-up genotyping showed that IGF2R IVS15+213C>T was also associated with increased osteosarcoma risk. Resequence analysis identified two additional SNPs linked to the risk-associated SNPs; linkage disequilibrium was strongest in a 1-kb pair region around them. The Ex16+88G>A SNP is located within a CpG island and alters methylation at that site.This pilot study of germ-line genetic variation in growth pathway genes and osteosarcoma identified a haplotype block in IGF2R associated with increased risk of osteosarcoma. The presence of a SNP in this block results in loss of methylation at a CpG island, providing corroborative evidence of a possible functional variant. Our analysis of the IGF2R haplotype structure will be applicable to future studies of IGF2R and disease risk.', 'http://www.ncbi.nlm.nih.gov/pubmed/17684144', 'yes', 'yes', 'NA', 1),
(17690908, 'Honorati MC, Cattini L, Facchini A', 'Possible prognostic role of IL-17R in osteosarcoma.', 'J Cancer Res Clin Oncol', '2007', 'Possible prognostic role of IL-17R in osteosarcoma. An intense vascularization of primary tumor mass is associated with a fatal outcome in various types of invasive solid tumors. Interleukin 17 (IL-17), a CD4+ T-cell-derived cytokine, stimulates some tumor cells to secrete angiogenic factors, among which venous endothelial growth factor (VEGF). We assessed whether the expression of IL-17 receptor (IL-17R) represents a marker for the metastasizing ability of osteosarcoma (OS), a very malignant bone tumor.We immunoassayed the amount of VEGF secreted by three OS cell lines expressing IL-17R in differing amounts: HOS, MG63 and U-2 OS, and their sensitivity to IL-17 stimulation to secrete VEGF.U-2 OS, which best expresses IL-17R, secreted the highest amounts of VEGF and was the most sensitive to IL-17, whereas MG63 expressed the lowest level of IL-17R, secreted the lowest amount of VEGF and was not sensitive to IL-17. IL-17R expression correlated with VEGF secretion and IL-17 sensitivity. U-2 OS expressed the most dedifferentiated phenotype, which is associated with tumor malignancy.These results suggest that IL-17R in OS might represent a marker of tumor metastasis potential.', 'http://www.ncbi.nlm.nih.gov/pubmed/17690908', 'no', 'yes', 'NA', 2);
INSERT INTO `pm` (`pmid`, `author`, `title`, `journal`, `year`, `abstract`, `url`, `tumor`, `value`, `addon`, `frequency`) VALUES
(17699771, 'Cantiani L, Manara MC, Zucchini C, De Sanctis P, Zuntini M, Valvassori L, Serra M, Olivero M, Di Renzo MF, Colombo MP, Picci P, Scotlandi K', 'Caveolin-1 reduces osteosarcoma metastases by inhibiting c-Src activity and met signaling.', 'Cancer Res', '2007', 'Caveolin-1 reduces osteosarcoma metastases by inhibiting c-Src activity and met signaling. Caveolin-1 (Cav-1) is highly expressed in normal osteoblasts. This article reports that Cav-1 down-regulation is part of osteoblast transformation and osteosarcoma progression and validates its role as oncosuppressor in human osteosarcoma. A survey of 6-year follow-up indicates a better overall survival for osteosarcoma expressing a level of Cav-1 similar to osteoblasts. However, the majority of primary osteosarcoma shows significantly lower levels of Cav-1 than normal osteoblasts. Accordingly, Met-induced osteoblast transformation is associated with Cav-1 down-regulation. In vitro, osteosarcoma cell lines forced to overexpress Cav-1 show reduced malignancy with inhibited anchorage-independent growth, migration, and invasion. In vivo, Cav-1 overexpression abrogates the metastatic ability of osteosarcoma cells. c-Src and c-Met tyrosine kinases, which are activated in osteosarcoma, colocalize with Cav-1 and are inhibited on Cav-1 overexpression. Thus, Cav-1 behaves as an oncosuppressor in osteosarcoma. Altogether, data suggest that Cav-1 down-modulation might function as a permissive mechanism, which, by unleashing c-Src and Met signaling, enables osteosarcoma cells to invade neighboring tissues. These data strengthen the rationale to target c-Src family kinases and/or Met receptor to improve the extremely poor prognosis of metastatic osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/17699771', 'no', 'yes', 'NA', 3),
(17786187, 'Entz-Werle N, Lavaux T, Metzger N, Stoetzel C, Lasthaus C, Marec P, Kalifa C, Brugieres L, Pacquement H, Schmitt C, Tabone MD, Gentet JC, Lutz P, Babin A, Oudet P, Gaub MP, Perrin-Schmitt F', 'Involvement of MET/TWIST/APC combination or the potential role of ossification factors in pediatric high-grade osteosarcoma oncogenesis.', 'Neoplasia', '2007', 'Involvement of MET/TWIST/APC combination or the potential role of ossification factors in pediatric high-grade osteosarcoma oncogenesis. Dysregulated cell growth or differentiation due to misexpression of developmental critical factors seems to be a decisive event in oncogenesis. As osteosarcomas are histologically defined by malignant osteoblasts producing an osteoid component, we prospected in pediatric osteosarcomas treated with OS94 protocol the genomic status of several genes implied in ossification processes. In 91 osteosarcoma cases, we focused on the analysis of the fibroblast growth factor receptors (FGFRs) TWIST, APC, and MET by allelotyping, real-time quantitative polymerase chain reaction, gene sequencing, and protein polymorphism study. Our study supports the frequent role of TWIST, APC, and MET as osteosarcoma markers (50%, 62%, and 50%, respectively). TWIST and MET were mainly found to be deleted, and no additional APC mutation was identified. Surprisingly, FGFRs are abnormal in only < 30%. Most of these factors and their abnormalities seem to be linked more or less to one clinical subgroup, but the most significant correlation is the link of MET, TWIST, and APC abnormalities to a worse outcome and their combination within abnormal tumors. A wider cohort is mandatory to define more robust molecular conclusions, but these results are to be considered as the beginning of a more accurate basis for diagnosis, in search of targeted therapies, and to further characterize prognostic markers.', 'http://www.ncbi.nlm.nih.gov/pubmed/17786187', 'yes', 'yes', 'NA', 3),
(17786325, 'Matsui TA, Sowa Y, Murata H, Takagi K, Nakanishi R, Aoki S, Yoshikawa M, Kobayashi M, Sakabe T, Kubo T, Sakai T', 'The plant alkaloid cryptolepine induces p21WAF1/CIP1 and cell cycle arrest in a human osteosarcoma cell line.', 'Int J Oncol', '2007', 'The plant alkaloid cryptolepine induces p21WAF1/CIP1 and cell cycle arrest in a human osteosarcoma cell line. We previously established a bioassay method to screen for compounds that activate the promoter activity of p21(WAF1/CIP1), a potent inhibitor of cyclin-dependent kinases, in a p53-independent manner. As an activator of p21(WAF1/CIP1) promoter activity, we isolated cryptolepine (CLP: 5-methyl indolo (2,3b)-quiniine), an indoloquinoline alkaloid, from the traditional Ayurvedic medicinal plant Sida cordifolia. We show here that CLP induces the expression of p21(WAF1/CIP1) with growth arrest in p53-mutated human osteosarcoma MG63 cells. Four micromolar of CLP completely inhibited the growth of MG63 cells and caused G2/M-phase arrest. CLP up-regulated the expression of p21(WAF1/CIP1) at both mRNA and protein levels in a dose-dependent manner. Using several mutant p21(WAF1/CIP1) promoter constructs, we found that the CLP-responsive element is an Sp1 site at -82 relative to the transcription start site of the p21(WAF1/CIP1) promoter. These findings suggest that CLP arrests the growth of MG63 cells by activating the p21(WAF1/CIP1) promoter through the specific Sp1 site in a p53-independent manner. In addition, CLP-mediated cell cycle arrest was reduced by the knockout of the p21(WAF1/CIP1) gene in human colon cancer HCT116 cells, suggesting that the cell cycle arrest by CLP was at least partially mediated through the induction of p21(WAF1/CIP1) expression. Although we need further study of chemotherapeutic effect in vivo, these results raise the possibility that CLP might be a suitable chemotherapeutic agent for treatment of osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/17786325', 'no', 'no', 'cryptolepine', 2),
(17786474, 'Salas S, Bartoli C, Deville JL, Gaudart J, Fina F, Calisti A, Bollini G, Curvale G, Gentet JC, Duffaud F, Figarella-Branger D, Bouvier C', 'Ezrin and alpha-smooth muscle actin are immunohistochemical prognostic markers in conventional osteosarcomas.', 'Virchows Arch', '2007', 'Ezrin and alpha-smooth muscle actin are immunohistochemical prognostic markers in conventional osteosarcomas. Ezrin is a cytoskeleton linker protein that is actively involved in the metastatic process of cancer cells. We have searched for a prognostic value of ezrin and some of its partners: alpha-smooth muscle actin and CD44H in 37 patients with an osteosarcoma. Automate immunohistochemistry (IHC) with anti-ezrin, alpha-smooth muscle actin and CD44H antibodies was performed in 66 specimens: 37 biopsies before chemotherapy, 16 resected tumours of \\poor\\ responders and 13 metastases. The messenger RNA (mRNA) levels of ezrin of 13 frozen biopsies and 4 metastases were evaluated by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). All results were correlated to the following clinical data. Ezrin expression by IHC was found in 62% of 37 biopsies in the different histological subtypes. A good correlation was found between positive or negative samples by IHC and mRNA levels. Ezrin expression was recorded in 84.5% of metastastic samples. The mean expression of ezrin was higher in metastases than biopsies (p = 0.024). In multivariate analysis, ezrin was an independent prognostic marker for event-free survival and overall survival (OS) with p < 0.001 and p = 0.003, respectively, and alpha-smooth muscle actin for OS only (p = 0.024). Our findings suggest that ezrin and alpha-smooth muscle actin are predictive IHC prognostic markers for patients with an osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/17786474', 'yes', 'yes', 'NA', 3),
(17849429, 'Kovacevic A, Hammer A, Stadelmeyer E, Windischhofer W, Sundl M, Ray A, Schweighofer N, Friedl G, Windhager R, Sattler W, Malle E', 'Expression of serum amyloid A transcripts in human bone tissues, differentiated osteoblast-like stem cells and human osteosarcoma cell lines.', 'J Cell Biochem', '2008', 'Expression of serum amyloid A transcripts in human bone tissues, differentiated osteoblast-like stem cells and human osteosarcoma cell lines. Although the liver is the primary site of cytokine-mediated expression of acute-phase serum amyloid A (SAA) protein, extrahepatic production has also been reported. Besides its role in amyloidosis and lipid homeostasis during the acute-phase, SAA has recently been assumed to contribute to bone and cartilage destruction. However, expression of SAA in human osteogenic tissue has not been studied. Therefore, we first show that SAA1 (coding for the major SAA isoform) but not SAA2 transcripts are expressed in human trabecular and cortical bone fractions and bone marrow. Next, we show expression of (i) IL-1, IL-6, and TNF receptor transcripts; (ii) the human homolog of SAA-activating factor-1 (SAF-1, a transcription factor involved in cytokine-mediated induction of SAA genes); and (iii) SAA1/2 transcripts in non-differentiated and, to a higher extent, in osteoblast-like differentiated human mesenchymal stem cells. Third, we provide evidence that human osteoblast-like cells of tumor origin (MG-63 and SAOS-2) express SAF-1 under basal conditions. SAA1/2 transcripts are expressed under basal conditions (SAOS-2) and cytokine-mediated conditions (MG-63 and SAOS-2). RT-PCR, Western blot analysis, and immunofluorescence technique confirmed cytokine-mediated expression of SAA on RNA and protein level in osteosarcoma cell lines while SAA4, a protein of unknown function, is constitutively expressed in all osteogenic tissues investigated.Copyright 2007 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/17849429', 'no', 'no', 'NA', 5),
(17873971, 'Wang R, Dong K, Lin F, Wang X, Gao P, Wei SH, Cheng SY, Zhang HZ', 'Inhibiting proliferation and enhancing chemosensitivity to taxanes in osteosarcoma cells by RNA interference-mediated downregulation of stathmin expression.', 'Mol Med', 'NA', 'Inhibiting proliferation and enhancing chemosensitivity to taxanes in osteosarcoma cells by RNA interference-mediated downregulation of stathmin expression. Stathmin (Oncoprotein18), a signal transduction regulatory factor, plays an important role in cell division and malignant tumor development. Stathmin is a ubiquitous intracellular phosphoprotein that is overexpressed in a variety of human malignancies, including osteosarcoma. To investigate the potential use of stathmin as a therapeutic target for human osteosarcomas, we employed RNA interference [small interfering RNA (siRNA)] to reduce stathmin expression in human osteosarcoma cell lines and analyzed their phenotypic changes. Results showed that the downregulation of stathmin expression in human osteosarcoma cells significantly inhibited cell proliferation in vitro and tumorigenicity in vivo. The specific downregulation induced cell arrest in the G(2)/M phase of cell cycle and eventually apoptotic cell death. Taxanes are a group of effective chemotherapeutic agents whose activity is mediated through stabilization of the microtubules of the mitotic spindle. In the present study, we also observed a synergistic enhancement of the cytotoxicity effect by combination use of taxanes and RNA interference-mediated stathmin downregulation. All these experimental data indicate that stathmin downregulation can lead to potent antitumor activity and chemosensitizing activity to taxanes in human osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/17873971', 'no', 'yes', 'taxanes', 1),
(17886294, 'Ferrari S, Zanella L, Alberghini M, Palmerini E, Staals E, Bacchini P', 'Prognostic significance of immunohistochemical expression of ezrin in non-metastatic high-grade osteosarcoma.', 'Pediatr Blood Cancer', '2008', 'Prognostic significance of immunohistochemical expression of ezrin in non-metastatic high-grade osteosarcoma. Ezrin is a membrane-cytoskeleton linker protein involved in regulating growth and metastatic behaviour of cancer cells. The study evaluated ezrin expression and its prognostic value in patients with non-metastatic osteosarcoma.Ezrin expression and pattern of staining (cytoplasmic or membraneous and cytoplasmic) were assessed using immunohistochemistry on slides from tumour biopsy. We studied 95 patients (median 16 years, range: 4-39 years) with primary non-metastatic osteosarcoma of the extremity treated by neoadjuvant chemotherapy based on methotrexate, cisplatin, doxorubicin and ifosfamide.Seventy-six patients (80%) showed ezrin immunoreactivity: in cytoplasm (37, 47%) and in cytoplasm and membrane (42, 53%) of tumour cells. Immunohistochemical staining score was: 1+ (16, 24%), 2+ (10, 13%), 3+ (17, 21.5%) and 4+ (36, 45.5%). Ezrin and score expression were not related to gender, site, alkaline phosphatase (AP), LDH serum levels, chemotherapy-induced tumour necrosis or patient outcome. A significant association was seen between expression pattern and prognosis. The 3-year probability of disease-free survival was 80% for patients with only cytoplasmic immunostaining and 54% for patients with cytoplasmic and membranous immunostaining (P < 0.02).Ezrin immunoreactivity can be detected in the majority of patients with non-metastatic osteosarcoma of the extremity. The pattern of ezrin staining can identify patients with different risks of relapse. In patients who only have ezrin cytoplasmic expression, a probability of EFS >80% at 5 years can be expected. These results suggest further investigations to define the relation between expression pattern, ezrin functional status and outcome in patients with non-metastatic osteosarcoma.(c) 2008 Wiley-Liss, Inc.', 'http://www.ncbi.nlm.nih.gov/pubmed/17886294', 'yes', 'yes', 'NA', 1),
(17892509, 'Wang D, Zhong ZY, Li MX, Xiang DB, Li ZP', 'Vector-based Ape1 small interfering RNA enhances the sensitivity of human osteosarcoma cells to endostatin in vivo.', 'Cancer Sci', '2007', 'Vector-based Ape1 small interfering RNA enhances the sensitivity of human osteosarcoma cells to endostatin in vivo. Osteosarcoma is a highly vascular and extremely destructive malignancy, and the survival of patients with osteosarcoma has not improved significantly in recent years. Antiangiogenic therapy currently holds great potential in conjunction with conventional treatment modalities for osteosarcoma. However, there are examples of gradual loss of response, and perhaps acquired resistance to antiangiogenic drugs. The acquired resistance of antiangiogenesis may be associated with a lot of hypoxia-response genes. The human apurinic/apyrimidinic endonuclease (Ape1) protein, a bifunctional redox factor and apurinic/apyrimidinic (AP) endonuclease, plays a crucial role in protecting against cell death due to hypoxia. We therefore hypothesized that Ape1 may contribute to the resistance of antiangiogenic therapy. To investigate the effect of Ape1 on the sensitivity of human osteosarcoma cells to endostatin, we constructed an Ape1 small interfering RNA expression vector, pSilenceApe1. Transfection of human osteosarcoma 9901 and HOS cells with pSilenceApe1 resulted in a dose-dependent loss of Ape1 protein. pSilenceApe1 also significantly suppressed the expression of vascular endothelial growth factor (VEGF) protein in the 9901 cells. Combined treatment with pSilenceApe1 and recombinant human endostatin (rhES) showed potent antiangiogenic effects in the transwell chamber invasion assay. Then, 20 nude mice bearing 9901 xenografts were divided into four groups: the phosphate-buffered saline treatment control group; the rhES treatment group (1.5 mg/kg, daily); the pSilenceApe1 treatment group (20 microg, once every 3 days); and the combination of rhES and pSilenceApe1 treatment group. pSilenceApe1 significantly suppressed the expression of Ape1 and VEGF protein in the 9901 xenografts. The tumor-inhibition rate of the pSilenceApe1, rhES, and combination of rhES and pSilenceApe1 treatment groups was 38.23, 35.29, and 62.18%, respectively. Furthermore, a significant decrease in microvessel density with an increase in apoptosis was observed following combined treatment with pSilenceApe1 and rhES, compared with control and either agent alone in 9901 xenografts. These results indicate that Ape1 small interfering RNA could enhance the sensitivity of osteosarcoma cells to endostatin.', 'http://www.ncbi.nlm.nih.gov/pubmed/17892509', 'no', 'no', 'NA', 2),
(17934334, 'Lee EJ, Choi EM, Kim SR, Park JH, Kim H, Ha KS, Kim YM, Kim SS, Choe M, Kim JI, Han JA', 'Cyclooxygenase-2 promotes cell proliferation, migration and invasion in U2OS human osteosarcoma cells.', 'Exp Mol Med', '2007', 'Cyclooxygenase-2 promotes cell proliferation, migration and invasion in U2OS human osteosarcoma cells. Osteosarcoma is the most common primary bone tumor, but the pathogenesis is not well understood. While cyclooxygeanse-2 (COX-2) is known to be closely associated with tumor growth and metastasis in several kinds of human tumors, the function of COX-2 in osteosarcoma is unclear. Therefore, to investigate the function of COX-2 in osteosarcoma, we established stable cell lines overexpressing COX-2 in U2OS human osteosarcoma cells. COX-2 overexpression as well as prostaglandin E2 treatment promoted proliferation of U2OS cells. In addition, COX-2 overexpression enhanced mobility and invasiveness of U2OS cells, which was accompanied by increases of matrix metalloproteinase-2 and -9 (MMP-2 and -9) activities. Selective COX-2 inhibitors, NS-398 and celecoxib, inhibited cell proliferation and abrogated the enhanced mobility, invasiveness and MMP activities induced by COX-2 overexpression. These results suggest that COX-2 is directly associated with cell proliferation, migration and invasion in human osteosarcoma cells, and the therapeutic value of COX-2 inhibitors should be evaluated continuously.', 'http://www.ncbi.nlm.nih.gov/pubmed/17934334', 'no', 'no', 'NS-398|celecoxib', 3),
(17952379, 'Liu SJ, Zhang DQ, Sui XM, Zhang L, Cai ZW, Sun LQ, Liu YJ, Xue Y, Hu GF', 'The inhibition of in vivo tumorigenesis of osteosarcoma (OS)-732 cells by antisense human osteopontin RNA.', 'Cell Mol Biol Lett', '2008', 'The inhibition of in vivo tumorigenesis of osteosarcoma (OS)-732 cells by antisense human osteopontin RNA. Osteopontin (OPN) is a secreted, non-collagenous, sialic acid-rich protein which functions by mediating cell-matrix interactions and cellular signaling via binding with integrins and CD44 receptors. An increasing number of studies have shown that OPN plays an important role in controlling cancer progression and metastasis. OPN was found to be expressed in many human cancer types, and in some cases, its over-expression was shown to be directly associated with poor patient prognosis. In vitro cancer cell line and animal model studies have clearly indicated that OPN can function in regulating the cell signaling that ultimately controls the oncogenic potential of various cancers. Previous studies in our laboratory demonstrated that OPN is highly expressed in human osteosarcoma (OS) cell line OS-732. In this study, we successfully reduced the tumorigenecity of OS-732 cells xenotransplanted into nude mice, using the antisense human OPN (hOPN) RNA expression vector.', 'http://www.ncbi.nlm.nih.gov/pubmed/17952379', 'no', 'yes', 'NA', 1),
(17956871, 'Lamour V, Detry C, Sanchez C, Henrotin Y, Castronovo V, Bellahcène A', 'Runx2- and histone deacetylase 3-mediated repression is relieved in differentiating human osteoblast cells to allow high bone sialoprotein expression.', 'J Biol Chem', '2007', 'Runx2- and histone deacetylase 3-mediated repression is relieved in differentiating human osteoblast cells to allow high bone sialoprotein expression. Bone sialoprotein (BSP) is a bone matrix glycoprotein whose expression coincides with terminal osteoblastic differentiation and the onset of mineralization. In this study we show that BSP expression is considerably increased in confluent Saos-2 human osteosarcoma cells and in differentiating normal human osteoblasts, concomitantly with the decrease of Runx2, a key transcription factor controlling bone formation. Therefore, we investigated the role of Runx2 in the regulation of BSP expression in Saos-2 cells. Using a mobility shift assay, we demonstrated that Runx2 binds to the BSP promoter only in preconfluent cells. Histone deacetylase 3 (HDAC3) has been recently shown to act as a Runx2 co-repressor. Chromatin immunoprecipitation assays demonstrated that both Runx2 and HDAC3 are detectable at the BSP promoter in preconfluent Saos-2 cells but not when they are confluent and overexpress BSP. Consistently, nuclear Runx2 protein level is down-regulated, whereas Saos-2 cells became increasingly confluent. Finally, the suppression of HDAC3, Runx2, or both by RNA interference induced the expression of BSP at both mRNA and protein levels in Saos-2 cells. Our data demonstrate that Runx2 and HDAC3 repress BSP gene expression and that this repression is suspended upon osteoblastic cell differentiation. Both the nuclear disappearance of Runx2 and the non-recruitment of HDAC3 represent new means to relieve Runx2-mediated suppression of BSP expression, thus allowing the acquisition of a fully differentiated and mineralization-competent phenotype by osteoblast cells.', 'http://www.ncbi.nlm.nih.gov/pubmed/17956871', 'no', 'no', 'NA', 3),
(17959036, 'Yuan XW, Zhu XF, Huang XF, Sheng PY, He AS, Yang ZB, Deng R, Feng GK, Liao WM', 'Interferon-alpha enhances sensitivity of human osteosarcoma U2OS cells to doxorubicin by p53-dependent apoptosis.', 'Acta Pharmacol Sin', '2007', 'Interferon-alpha enhances sensitivity of human osteosarcoma U2OS cells to doxorubicin by p53-dependent apoptosis. To determine whether interferon-alpha(IFNalpha) can enhance doxorubicin sensitivity in osteosarcoma cells and its molecular mechanism.Cell viability was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptosis was studied using Flow cytometry analysis, Hoechst33258 staining, DNA fragmentation assay, as well as the activation of caspase-3 and poly (ADP-ribose) polymerase. Protein expression was detected by Western blotting. The dependence of p53 was determined using p53-siRNA transfection.IFNalpha increased doxorubicin-induced cytotoxicity to a much greater degree through apoptosis in human osteosarcoma p53-wild U2OS cells, but not p53-mutant MG63 cells. IFNalpha markedly upregulated p53, Bax, Mdm2, and p21, downregulated Bcl-2, and activated caspase-3 and PARP cleavage in response to doxorubicin in U2OS cells. Moreover, the siRNA-mediated silencing of p53 significantly reduced the IFNalpha/doxorubicin combination-induced cytotoxicity and PARP cleavage.IFNalpha enhances the sensitivity of human osteosarcoma U2OS cells to doxorubicin by p53-dependent apoptosis. The proper combination with IFNalpha and conventional chemotherapeutic agents may be a rational strategy for improving the treatment of osteosarcoma with functional p53.', 'http://www.ncbi.nlm.nih.gov/pubmed/17959036', 'no', 'no', 'doxorubicin', 7),
(17980151, 'Aoyama T, Okamoto T, Kohno Y, Fukiage K, Otsuka S, Furu M, Ito K, Jin Y, Nagayama S, Nakayama T, Nakamura T, Toguchida J', 'Cell-specific epigenetic regulation of ChM-I gene expression: crosstalk between DNA methylation and histone acetylation.', 'Biochem Biophys Res Commun', '2008', 'Cell-specific epigenetic regulation of ChM-I gene expression: crosstalk between DNA methylation and histone acetylation. The expression of the chondromodulin-I (ChM-I) gene, a cartilage-specific gene, is regulated by the binding of Sp3 to the core promoter region, which is inhibited by the methylation of CpG in the target genome in the osteogenic lineage, osteosarcoma (OS) cells. The histone tails associated with the hypermethylated promoter region of the ChM-I gene were deacetylated by histone deacetylase 2 (HDAC2) in three ChM-I-negative OS cell lines. Treatment with an HDAC inhibitor induced the binding of Sp3 in one cell line, which became ChM-I-positive. This process was associated with acetylation instead of the dimethylation of histone H3 at lysine 9 (H3-K9) and, surprisingly, the demethylation of the core promoter region. The demethylation was transient, and gradually replaced by methylation after a rapid recovery of histone deacetylaion. These results represent an example of the plasticity of differentiation being regulated by the cell-specific plasticity of epigenetic regulation.', 'http://www.ncbi.nlm.nih.gov/pubmed/17980151', 'no', 'no', 'NA', 4),
(17980359, 'Choi HJ, Eun JS, Kim DK, Li RH, Shin TY, Park H, Cho NP, Soh Y', 'Icariside II from Epimedium koreanum inhibits hypoxia-inducible factor-1alpha in human osteosarcoma cells.', 'Eur J Pharmacol', '2008', 'Icariside II from Epimedium koreanum inhibits hypoxia-inducible factor-1alpha in human osteosarcoma cells. Hypoxia-inducible factor-1 (HIF-1) is an important tumor-selective therapeutic target for solid tumors. Icariside II was isolated from Epimedium koreanum through successive fractionation with ethyl acetate, n-butanol, chloroform and hexane, followed by gel column chromatography. Icariside II attenuated the protein level of HIF-1alpha induced by hypoxia in human osteosarcoma (HOS) cells in a concentration-dependent manner, probably by enhancing the interaction rate between von Hippel-Lindau (VHL) and HIF-1alpha. Furthermore, Icariside II down-regulated the levels of HIF-inducible genes involved in angiogenesis, metastasis, and glucose metabolism, such as vascular endothelial growth factor (VEGF), urokinase plasminogen activator receptor (uPAR), adrenomedullin (ADM), matrix metalloproteinase 2 (MMP2), aldolase A, and enolase 1 in HOS cells. Icariside II also inhibited the migration rate in HOS cells and tube formation rate in human umbilical vein endothelium cells (HUVECs). Overall, these results suggest the potential use of Icariside II as a therapeutic candidate against various diseases that involve overexpression of HIF-1alpha.', 'http://www.ncbi.nlm.nih.gov/pubmed/17980359', 'no', 'no', 'Icariside II', 8),
(17981215, 'Selvarajah S, Yoshimoto M, Maire G, Paderova J, Bayani J, Squire JA, Zielenska M', 'Identification of cryptic microaberrations in osteosarcoma by high-definition oligonucleotide array comparative genomic hybridization.', 'Cancer Genet Cytogenet', '2007', 'Identification of cryptic microaberrations in osteosarcoma by high-definition oligonucleotide array comparative genomic hybridization. Osteosarcoma (OS) is an aggressive bone tumor characterized by complex abnormal karyotypes and a high level of genomic instability. Using high-resolution array comparative genomic hybridization (aCGH), a novel class of localized copy number variations called microaberrations has been detected. These genomic anomalies typically involve DNA imbalances affecting 700 kb to 1 Mb DNA, and are often associated with some type of genetic syndromes. Because the origin of instability in OS is poorly understood, we used aCGH to determine whether microaberrations were a characteristic of four OS cell lines: U-2 OS, HOS, MG-63, and SAOS-2. TP53 is mutated in SAOS-2, a line in which 17 microaberrations were found. In contrast, U-2 OS, which has a wild-type TP53, had only six such anomalies, the lowest incidence. A 500-kb microaberration within a region of gain at 5p15.33 in SAOS-2 was confirmed by fluorescence in situ hybridization. Significantly, this genomic location is close to the TERT gene, a region of gain in all four cell lines. To our knowledge, this is the first systematic analysis of the incidence of microaberrations in OS. The high levels of these anomalies detected suggest that the instability processes in OS that lead to a highly abnormal karyotypes may also be associated with acquisition of genomic microaberrations.', 'http://www.ncbi.nlm.nih.gov/pubmed/17981215', 'no', 'no', 'NA', 2),
(17982618, 'Mori K, Berreur M, Blanchard F, Chevalier C, Guisle-Marsollier I, Masson M, Rédini F, Heymann D', 'Receptor activator of nuclear factor-kappaB ligand (RANKL) directly modulates the gene expression profile of RANK-positive Saos-2 human osteosarcoma cells.', 'Oncol Rep', '2007', 'Receptor activator of nuclear factor-kappaB ligand (RANKL) directly modulates the gene expression profile of RANK-positive Saos-2 human osteosarcoma cells. Receptor activator of nuclear factor kappaB (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) are the key regulators of bone metabolism. Recent findings demonstrated a crucial role of RANK in several bone-associated tumors. Indeed, we have recently demonstrated functional RANK expression both in a mouse and several human osteosarcoma cell lines. However, RANKL effects on osteosarcoma cells remain to be determined. In this study, we determined RANKL effects on RANK-positive Saos-2 human osteosarcoma cells. cDNA microarray and quantitative RT-PCR analyses clearly demonstrated that RANK-positive osteosarcoma cells were the target of RANKL as well as osteoclasts/osteoclast precursors. Thus, we present for the first time that RANKL can directly and significantly modulate gene expression of RANK-expressing Saos-2 cells. RANKL-modulated genes included genes that were implicated in protein metabolism, nucleic acid metabolism, intracellular transport, cytoskeleton organization and biogenesis, apoptosis and signaling cascade. Our results strengthen the involvement of the RANK/RANKL/OPG axis in osteosarcoma biology and capability to identify novel therapeutic approaches targeting RANK-positive osteosarcomas.', 'http://www.ncbi.nlm.nih.gov/pubmed/17982618', 'no', 'no', 'NA', 3),
(17987039, 'Lee N, Smolarz AJ, Olson S, David O, Reiser J, Kutner R, Daw NC, Prockop DJ, Horwitz EM, Gregory CA', 'A potential role for Dkk-1 in the pathogenesis of osteosarcoma predicts novel diagnostic and treatment strategies.', 'Br J Cancer', '2007', 'A potential role for Dkk-1 in the pathogenesis of osteosarcoma predicts novel diagnostic and treatment strategies. Canonical Wnt signalling is an osteoinductive signal that promotes bone repair through acceleration of osteogenic differentiation by progenitors. Dkk-1 is a secreted inhibitor of canonical Wnt signalling and thus inhibits osteogenesis. To examine a potential osteoinhibitory role of Dkk-1 in osteosarcoma (OS), we measured serum Dkk-1 in paediatric patients with OS (median age, 13.4 years) and found it to be significantly elevated. We also found that Dkk-1 was maximally expressed by the OS cells at the tumour periphery and in vitro, Dkk-1 and RANKL are coexpressed by rapidly proliferating OS cells. Both Dkk-1 and conditioned media from OS cells reduce osteogenesis by human mesenchymal cells and by immunodepletion of Dkk-1, or by adding a GSK3beta inhibitor, the effects of Dkk-1 were attenuated. In mice, we found that the expression of Dkk-1 from implanted tumours was similar to the human tumour biopsies in that human Dkk-1 was present in the serum of recipient animals. These data demonstrate that systemic levels of Dkk-1 are elevated in OS. Furthermore, the expression of Dkk-1 by the OS cells at the periphery of the tumour probably contributes to its expansion by inhibiting repair of the surrounding bone. These data demonstrate that Dkk-1 may serve as a prognostic or diagnostic marker for evaluation of OS and furthermore, immunodepletion of Dkk-1 or administration of GSK3beta inhibitors could represent an adjunct therapy for this disease.', 'http://www.ncbi.nlm.nih.gov/pubmed/17987039', 'yes', 'yes', 'NA', 2),
(17998668, 'Rakesh Kumar V, Gupta N, Kakkar N, Sharma SC', 'Prognostic and predictive value of c-erbB2 overexpression in osteogenic sarcoma.', 'J Cancer Res Ther', 'NA', 'Prognostic and predictive value of c-erbB2 overexpression in osteogenic sarcoma. Osteogenic sarcoma (OS) is a highly malignant tumor of the bone. There are few reports in the literature regarding determination of c-erb B-2 (HER-2/neu) expression in cases of OS and also the results are quite variable.The present study was undertaken to correlate the expression of c-erbB2 with the survival of patients of OS.A retrospective study of cases of OS, in which the expression of c-erbB-2 by immunohistochemistry (IHC) was studied and correlated with the survival rate of the patients.Out of a total of 49 cases, proper follow up data in the form of total and disease free survival was available in only 20 cases. IHC for c-erb-B2 was carried out in these 20 cases.The staining pattern was as follows: no staining in 3 cases, 1 + in 5 cases, 2 + in 3 cases, 3 + in 3 cases and 4 + cytoplasmic positivity in 6 cases. Statistical analysis: Spearman rank correlation test was used to correlate the intensity of cytoplasmic staining of c-erbB-2 with survival rate of the patients, response to chemotherapy and metastasis.Previous studies have shown that c-erbB-2/HER-2 is an independent prognostic factor in osteogenic sarcoma, but in the present study, its expression was not seen to be correlating with survival rate of the patients. Therefore, further studies are needed to reach a consensus regarding the reliability of c-erbB-2 as an independent prognostic factor in OS.', 'http://www.ncbi.nlm.nih.gov/pubmed/17998668', 'no', 'no', 'arctiin', 2),
(18019405, 'Bui MM, Bagui TK, Boulware DC, Letson DG, Nasir A, Kaiser HE, Pledger WJ, Coppola D', 'Altered expression of cell cycle regulatory proteins in benign and malignant bone and soft tissue neoplasms.', 'In Vivo', 'NA', 'Altered expression of cell cycle regulatory proteins in benign and malignant bone and soft tissue neoplasms. The binding of cyclins to cyclin-dependent kinases regulates cell proliferation. Overexpression of cyclins is believed to deregulate the cell cycle in human tumors. Here the expression of G1 cyclins D1 and D3, and of Ki-67 in a variety of bone and soft tissue sarcomas was assessed as compared to adjacent normal tissue and to a subset of leiomyomas.Twenty-nine human bone and soft tissue sarcomas were evaluated. Tissue sections from each case were subjected to immunostaining for cyclin D1, cyclin D3 and Ki-67 using the avidin-biotin complex method.Cyclin D1 nuclear positivity was detected in 28% of sarcomas and in none of the leiomyomas. Cyclin D3 nuclear positivity was present in 62% of sarcomas and in none of the leiomyomas. Ki-67 nuclear staining was positive in 86% of sarcomas but in only 16% of leiomyomas. In addition, upregulation of cyclin D1 was observed in leiomyosarcomas, pleomorphic sarcomas and gastrointestinal stromal tumors, but not in liposarcomas or osteosarcomas. Cyclin D3, however, was expressed in all of the sarcoma types including 2 out of 5 liposarcomas and 1 out of 4 osteosarcomas. The normal soft tissue adjacent to the tumors when present (10 cases) was negative for cyclin D1 and D3, and expressed Ki-67 in 5% of the cell nuclei. The expression of cyclin D3 was also noted in human sarcoma cell lines (SKLMS, MG63, SaOS-2 and HT1080) by Western blot.The higher expression of cyclin D1 and D3 and of Ki-67 in bone and soft tissue sarcomas, as compared to leiomyomas and peritumoral normal soft tissue, suggests that high cyclin expression may contribute to deregulation of the cell cycle in bone and soft tissue tumors. These data suggest a role of cyclins in the process of human sarcomagenesis.', 'http://www.ncbi.nlm.nih.gov/pubmed/18019405', 'yes', 'no', 'NA', 3),
(18047805, 'Cho HJ, Lee TS, Park JB, Park KK, Choe JY, Sin DI, Park YY, Moon YS, Lee KG, Yeo JH, Han SM, Cho YS, Choi MR, Park NG, Lee YS, Chang YC', 'Disulfiram suppresses invasive ability of osteosarcoma cells via the inhibition of MMP-2 and MMP-9 expression.', 'J Biochem Mol Biol', '2007', 'Disulfiram suppresses invasive ability of osteosarcoma cells via the inhibition of MMP-2 and MMP-9 expression. Cancer cells, characterized by local invasion and distant metastasis, are very much dependent on the extracellular matrix. The expression of matrix metalloproteinases (MMPs) has been implicated in the invasion and metastasis of cancer cells. In this study, we reported the effects of disulfiram, a clinically used anti-alcoholism drug, on tumor invasion suppression, as well as its effects on the activity of MMP-2 and MMP-9 in human osteosarcoma cells (U2OS). Disulfiram has been used for alcohol aversion therapy. However, recent reports have shown that disulfiram may have potential in the treatment of human cancers. Herewith, we showed that the anti-tumor effects of disulfiram, in an invasion assay using U2OS cells and that disulfiram has a type IV collagenase inhibitory activity that inhibits expression of genes and proteins responsible for both cell and non-cell mediated invasion on pathways. In conclusion, disulfiram inhibited expression of MMP-2 and MMP-9 and it regulated the invasion of human osteosarcoma cells. These observations raise the possibility of disulfiram being used clinical for the inhibition of cancer invasion.', 'http://www.ncbi.nlm.nih.gov/pubmed/18047805', 'no', 'no', 'disulfiram', 2),
(18048360, 'Radjabi AR, Sawada K, Jagadeeswaran S, Eichbichler A, Kenny HA, Montag A, Bruno K, Lengyel E', 'Thrombin induces tumor invasion through the induction and association of matrix metalloproteinase-9 and beta1-integrin on the cell surface.', 'J Biol Chem', '2008', 'Thrombin induces tumor invasion through the induction and association of matrix metalloproteinase-9 and beta1-integrin on the cell surface. The procoagulatory serine protease, thrombin, is known to induce invasion and metastasis in various cancers, but the mechanisms by which it promotes tumorigenesis are poorly understood. Because the 92-kDa gelatinase (MMP-9) is a known mediator of tumor cell invasion, we sought to determine whether and how thrombin regulates MMP-9. The thrombin receptor, PAR-1, and MMP-9 are expressed in osteosarcomas, as determined by immunohistochemistry. Stimulation of U2-OS osteosarcoma cells with thrombin and a thrombin receptor-activating peptide induced pro-MMP-9 secretion as well as cell surface-associated pro-MMP-9 expression and proteolytic activity. This was paralleled by an increase in MMP-9 mRNA and MMP-9 promoter activity. Thrombin-induced invasion of U2-OS cells through Matrigel was mediated by the phosphatidylinositol 3-kinase signaling pathway and could be inhibited with an MMP-9 antibody. The stimulation of MMP-9 by thrombin was paralleled by an increase in beta1-integrin mRNA and beta1-integrin expression on the cell surface, which was also mediated by phosphatidylinositol 3-kinase and was required for invasion. Thrombin activation induced and co-localized both beta1-integrin and pro-MMP-9 on the cell membrane, as evidenced by co-immunoprecipitation, confocal microscopy, and a protein binding assay. The thrombin-mediated association of these two proteins, as well as thrombin-mediated invasion of U2-OS cells, could be blocked with a cyclic peptide and with an antibody preventing binding of the MMP-9 hemopexin domain to beta1-integrin. These results suggest that thrombin induces expression and association of beta1-integrin with MMP-9 and that the cell surface localization of the protease by the integrin promotes tumor cell invasion.', 'http://www.ncbi.nlm.nih.gov/pubmed/18048360', 'no', 'no', 'NA', 3),
(18081879, 'Dohi O, Hatori M, Suzuki T, Ono K, Hosaka M, Akahira J, Miki Y, Nagasaki S, Itoi E, Sasano H', 'Sex steroid receptors expression and hormone-induced cell proliferation in human osteosarcoma.', 'Cancer Sci', '2008', 'Sex steroid receptors expression and hormone-induced cell proliferation in human osteosarcoma. Sex steroid receptors including estrogen receptors (ER), progesterone receptors (PR), and androgen receptors (AR) have been sporadically reported in human osteosarcoma or its cell lines. Therefore, sex steroids have been considered to play some roles in human osteosarcoma, but no systematic and detailed studies regarding the correlation between the status of these receptors in sarcoma cells and clinicopathological parameters have been reported. We examined the existence of ER, PR and AR in 28 cases of osteosarcoma using immunohistochemistry. We then characterized the potential influence of sex steroids on cell proliferation of osteosarcoma cells using MG-63 human osteosarcoma cell line, which expressed all of these receptors. ER-beta and PR were detected in the great majority of the cases (23 and 24 cases, respectively) but ER-alpha and aromatase were not detected in all the cases, and AR was detected only in eight cases. There was a significant positive correlation between ER-beta and Ki-67 (MIB1) labeling indexes. The absence of aromatase in tumors also suggests the relative importance of concentrations of circulating sex steroids. Proliferation of MG-63 cells was significantly stimulated by estradiol, progesterone, and 5 alpha-dihydrotestosterone (DHT), and was significantly suppressed by the addition of fulvestrant (ICI), mifepristone (RU), and hydroxiflutamide, blockers for ER, PR and AR, respectively. Sex steroids, particularly estrogen and progesterone, are considered to play important roles in the regulation of cell proliferation in human osteosarcoma. In addition, these data suggest the potential for a novel endocrine therapy in osteosarcoma using clinically available inhibitors of progesterone and estrogen actions.', 'http://www.ncbi.nlm.nih.gov/pubmed/18081879', 'yes', 'no', 'estrogen|progesterone', 4),
(18090928, 'Koshkina NV, Kleinerman ES, Li G, Zhao CC, Wei Q, Sturgis EM', 'Exploratory analysis of Fas gene polymorphisms in pediatric osteosarcoma patients.', 'J Pediatr Hematol Oncol', '2007', 'Exploratory analysis of Fas gene polymorphisms in pediatric osteosarcoma patients. Defective apoptosis signaling by the Fas pathway has carcinogenic implications. We analyzed 123 pediatric patients with osteosarcoma for Fas single nucleotide polymorphisms: 2 of the promoter region (-1377 G>A and -670 A>G) and 2 of the coding region (exon 3 18272 A>G and exon 7 22628 C>T). As a comparison group, we used 510 adults without a history of cancer. We found an increased risk of osteosarcoma associated with the heterozygous genotype Fas exon 3 AG (genotype frequency 19.5% in cases vs. 12.0% in controls, P=0.028; adjusted odds ratio=1.6, 95% confidence interval=0.9-2.7], and this association was more pronounced in non-Hispanic whites (20.6% in cases vs. 10.1% in controls, P=0.014; adjusted odds ratio=2.3, 95% confidence interval=1.2-4.6). Additionally, the frequency of the variant allele (exon 3 G) was significantly higher in cases than in controls for both the entire group and non-Hispanic whites (P=0.046 and P=0.030, respectively). We found no significant association between osteosarcoma risk and the other Fas polymorphisms. This study suggests an association between the Fas exon 3 A>G polymorphism and osteosarcoma risk; however, further study is needed with pediatric controls and a larger sample size.', 'http://www.ncbi.nlm.nih.gov/pubmed/18090928', 'yes', 'no', 'NA', 1),
(18093185, 'Nikitovic D, Berdiaki A, Zafiropoulos A, Katonis P, Tsatsakis A, Karamanos NK, Tzanakakis GN', 'Lumican expression is positively correlated with the differentiation and negatively with the growth of human osteosarcoma cells.', 'FEBS J', '2008', 'Lumican expression is positively correlated with the differentiation and negatively with the growth of human osteosarcoma cells. Osteosarcoma is the most common primary bone tumour associated with childhood and adolescence. The possible role of the small leucine-rich proteoglycan, lumican, in the growth and metastasis of various cancer types has recently been investigated. In this study, the expression of lumican was examined in moderately differentiated (MG-63) and well-differentiated (Saos 2) human osteosarcoma cell lines of high and low metastatic capability, respectively. Real-time PCR, western blotting with antibodies against the protein core and keratan sulfate, and specific enzymatic digestions were the methods employed. The two human osteosarcoma cell lines were found to express and secrete lumican partly substituted with keratan sulfate glycosaminoglycans. Importantly, the non-metastatic, well-differentiated Saos 2 cells produced lumican at rates that were up to sevenfold higher than those of highly metastatic MG-63 cells. The utilization of short interfering RNA specific for the lumican gene resulted in efficient down-regulation of its mRNA levels in both cell lines. The growth of Saos 2 cells was inhibited by lumican, whereas their migration and chemotactic response to fibronectin were found to be promoted. Lumican expression was negatively correlated with the basal level of Smad 2 activation in these cells, suggesting that lumican may affect the bioavailability of Smad 2 activators. By contrast, these cellular functions of highly aggressive MG-63 cells were demonstrated not to be sensitive to a decrease in their low endogenous lumican levels. These results suggest that lumican expression may be positively correlated with the differentiation and negatively correlated with the progression of osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/18093185', 'no', 'no', 'NA', 3),
(18097539, 'Zucchini C, Rocchi A, Manara MC, De Sanctis P, Capanni C, Bianchini M, Carinci P, Scotlandi K, Valvassori L', 'Apoptotic genes as potential markers of metastatic phenotype in human osteosarcoma cell lines.', 'Int J Oncol', '2008', 'Apoptotic genes as potential markers of metastatic phenotype in human osteosarcoma cell lines. Metastasis is the most frequent cause of death among patients with osteosarcoma. We have previously demonstrated in independent experiments that the forced expression of L/B/K ALP and CD99 in U-2 OS osteosarcoma cell lines markedly reduces the metastatic ability of these cancer cells. This behavior makes these cell lines a useful model to assess the intersection of multiple and independent gene expression signatures concerning the biological problem of dissemination. With the aim to characterize a common transcriptional profile reflecting the essential features of metastatic behavior, we employed cDNA microarrays to compare the gene expression profiles of L/B/K ALP- and CD99-transfected osteosarcoma clones showing low metastatic ability with those of osteosarcoma cell lines showing contrasting behavior. Changes in gene expression were validated by real-time PCR and immunohistochemistry in independent samples. In our study we identified several differentially expressed genes (GADD45alpha, VCP, DHX9, survivin, alpha-catulin, ARPC1B) related to growth arrest and apoptosis. Most of these genes are functionally related with the nuclear factor (NF)-kappaB cell survival pathway that appeared to be inhibited in the less malignant osteosarcoma cells. Hence, we propose the inhibition of the NF-kappaB pathway as a rational strategy for effective management of human osteosarcoma.', 'http://www.ncbi.nlm.nih.gov/pubmed/18097539', 'no', 'no', 'NA', 6),
(18158779, 'Kaseta MK, Gomatos IP, Khaldi L, Tzagarakis GP, Alevizos L, Themistocleous GS, Leandros E, Soucacos PN', 'Prognostic value of bax, cytochrome C, and caspase-8 protein expression in primary osteosarcoma.', 'Hybridoma (Larchmt)', '2007', 'Prognostic value of bax, cytochrome C, and caspase-8 protein expression in primary osteosarcoma. The prognostic significance of bax, cytochrome c, and caspase-8 in patients with primary osteosarcoma is unknown. We examined the immunohistochemical expression of these genes in 35 surgically treated patients with primary osteosarcoma. Clinicopathological and survival data were correlated with the staining result. Eighteen tissue specimens from non-malignant osseous lesions were used as controls. Bax, cytochrome c, and caspase-8 positive staining was observed in 29 (82.9%), 16 (45.7%), and 0 (0%) patients, respectively, but did not stain any of the 18 benign osseous lesions used as controls. None of the genes studied predicted overall or disease-free survival. Patients, however, bearing bax(+)/cytochrome c(+) or bax(+)/cytochrome c(+ +) tumors had a decreased 4-year disease-free survival rate compared to the rest of the group (p = 0.0489 and p = 0.0208, respectively), identifying two groups of patients where more intensive adjuvant treatment could possibly be applied to prevent high postoperative recurrence rates.', 'http://www.ncbi.nlm.nih.gov/pubmed/18158779', 'yes', 'yes', 'NA', 3);
