Purpose: Cisplatin resistance is a major obstacle in successful treatment of late-stage ovarian cancer. The availability of reliable molecular markers to predict intrinsic and drug-induced cisplatin resistance in these cancer cells presents an enormous opportunity to individualized treatment for resistant patient sub-population. We have applied mass-spectrometry (MS) based proteomics approach to identify molecular markers that are over-expressed on cell surface of cisplatin resistant ovarian cell lines as compared to cisplatin-sensitive lines. This analysis has led to the identification of several candidate proteins that serve as potential diagnostic markers for ovarian cancer. Specifically, we have identified CD70 and B7-H2, co-stimulatory molecules involved in adaptive immunity, as candidate predictive and response markers respectively, for ovarian cancer. Experimental Design and Results: Mass-spectrometry analysis was performed on the cisplatin sensitive ovarian cell line A2780 and its cisplatin resistant derivative A2780cis. To reduce the complexity of the analysis, focus was placed on capturing proteins associated with the cell membrane including secreted/shed proteins which could serve as potential circulating biomarkers. Differentially expressed peptide ions corresponding to cell surface proteins between A2780 and its cisplatin derivative were identified by liquid chromatography-MS analysis and identified by searching MS/MS spectra against protein databases. 12 cell surface proteins, including Erb2 were expressed greater than 3-fold in the resistant line as compared to the sensitive line. Over-expression of the cell surface proteins on the A2780cis cell line was confirmed by flow cytometry and/or by Taqman mRNA analysis. The expression level of the candidate resistance markers was then evaluated in an independent set of 6 ovarian cancer lines exhibiting varying degrees of cisplatin sensitivity as measured by EC50 valuesusing Alamar blue assay. This analysis revealed expression levels of CD70, a member of TNF family of ligands, inversely correlated with the sensitivity to cisplatin for all the cell lines tested. Immunohistochemical analysis confirmed expression of CD70 in primary ovarian tumor specimens. Interestingly, the expression level of another immune modulator, B7-H2, a B7 family member, was increased 2-10 fold in sensitive lines upon cisplatin treatment but not in resistant lines, implying B7-H2 is a candidate response biomarker for cisplatin treatment. Conclusion: Using a MS based proteomics approach we have shown for the first time that expression of the immune modulators CD70 and B7-H2, is associated with cisplatin resistance in ovarian cancer. Follow-up examination of cell surface as well as secreted CD70 and B7-H2 levels in clinical tumor specimen with available cisplatin sensitivity data is warranted.
99th AACR Annual Meeting-- Apr 12-16, 2008; San Diego, CA