Reduced folate carrier protein expression in osteosarcoma: implications for the prediction of tumor chemosensitivity. High-dose methotrexate (MTX) is an important component of current protocols for the treatment of osteosarcoma. Although MTX uptake proceeds primarily through the reduced folate carrier (RFC) protein and efflux occurs via multidrug resistance protein 1 (MRP1), RFC protein expression in osteosarcoma remains unexamined.RFC and MRP1 expression (normalized to beta-actin expression) was examined with Western blot analysis in 11 osteosarcoma specimens obtained at diagnosis and 9 osteosarcoma specimens obtained on recurrence.The average RFC level in specimens obtained on recurrence was significantly higher than the level in specimens obtained at diagnosis (P = 0.0005). Furthermore, in all three matched pairs of diagnosis and recurrence specimens, RFC levels were higher in recurrence specimens than in the corresponding diagnosis specimens. Potential correlations between RFC and MRP1 expression and histologic response to preoperative chemotherapy were examined. Tumors with poor histologic responses (i.e., </= 90% necrosis) had significantly lower RFC levels than did those with favorable responses to chemotherapy (P = 0.0016). In contrast, there was no correlation between MRP1 levels at diagnosis and histologic response to chemotherapy (P = 0.8764). The elevated MRP1 levels in specimens obtained on recurrence relative to MRP1 levels in specimens obtained at diagnosis were not statistically significant (P = 0.2056).The significant correlation between low RFC levels at diagnosis and poor histologic response to preoperative chemotherapy suggests that RFC levels at diagnosis may be a useful predictor of chemosensitivity and warrants large-scale studies. In addition, postchemotherapy progression to recurrence is associated with a significant increase in RFC expression. To our knowledge, the current study is the first to examine RFC protein levels in tumor specimens. Cancer 2003.Copyright 2003 American Cancer Society.