The majority of ovarian cancer patients are treated with platinum-based chemotherapy, but the emergence of resistance to such chemotherapy severely limits its overall effectiveness. We have shown that development of resistance to this treatment can modify cell signaling responses in a model system wherein cisplatin treatment has altered cell responsiveness to ligands of the erbB receptor family. A cisplatinresistant ovarian carcinoma cell line PE01 CDDP was derived from the parent PE01 line by exposure to increasing concentrations of cisplatin, eventually obtaining a 20-fold level of resistance. Whereas PE01 cells were growth stimulated by the erbB receptor-activating ligands, such as transforming growth factor-A (TGFA), NRG1A, and NRG1B, the PE01 CDDP line was growth inhibited by TGFA and NRG1B but unaffected by NRG1A. TGFA increased apoptosis in PE01 CDDP cells but decreased apoptosis in PE01 cells. Differences in extracellular signal-regulated kinase and phosphatidylinositol 3-kinase signaling were also found, which may be implicated in the altered cell response to ligands. Microarray analysis revealed 51 genes whose mRNA increased by at least 2-fold in PE01 CDDP cells relative to PE01 (including FRA1, ETV4, MCM2, AXL, MT3, TRAP1, and FANCG), whereas 36 genes (including IGFBP3, TRAM1, and KRT4 and KRT19) decreased by a similar amount. Differential display reverse transcriptase-PCR identified altered mRNA expression for TCP1, SLP1, proliferating cell nuclear antigen, and ZXDA. Small interfering RNA inhibition of FRA1, TCP1, and MCM2 expression was associated with reduced growth and FRA1 inhibition with enhanced cisplatin sensitivity. Altered expression of these genes by cytotoxic exposure may provide survival advantages to cells including deregulation of signaling pathways, which may be critical in the development of drug resistance. (Cancer Res 2005; 65(15): 6789-800)

There is currently much interest in the role of the Raf family in cancer, particularly since mutated B-Raf has been shown to be oncogenic in certain disease types. In this study we have explored the expression, signaling and function of the three known Raf isoforms (Raf-1, A-Raf and B-Raf) in patients with ovarian cancer. While increased expression of Raf-1 was associated with poor survival, increased expression of B-Raf was associated with improved survival. Using a panel of ovarian cancer cell lines, all three isoforms were shown to be involved in growth factor initiated signaling. Antisense inhibition of function in ovarian cancer cell lines indicated that both Raf-1 and A-Raf, but not B-Raf, were linked to cell proliferation. Raf-1 (but not A-Raf or B-Raf) was also associated with reduced apoptosis. While individual Raf reduction by isoform-targeted antisense oligonucleotides (ODNs) produced growth inhibition in some cell lines, similar use of the MEK inhibitor UO126 produced growth inhibition in all cell lines tested. These data suggest that Raf-1 is the predominant Raf isoform responsible for regulating cellular growth in ovarian cancer cells and may be particularly important in high grade serous ovarian cancers.

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