Mechanism for the benefits of cetuximab in combination with oxaliplatin, in oxaliplatin sensitive and resistant colon cancer cell lines Free

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We previously reported that the chimeric IgG1 antibody against EGFR, cetuximab enhances the in vivo anti-tumor effects of oxaliplatin on subcutaneous colon cancer xenografts that are sensitive or resistant to oxaliplatin. This study was designed to evaluate the mechanism of action underlying the oxaliplatin resistance independent benefits of cetuximab. In vitro studies on oxaliplatin resistant cell lines showed increased cytotoxicity, observed as a cetuximab induced reduction in the GI₅₀ of oxaliplatin by 10 fold in HT-29 resistant (HT29-OXR) and 2 fold in KM12 resistant (KM12-OXR) cells compared to 5 fold in parental HT-29 and no change in parental KM12 cells. The apoptotic index was increased with combination compared to oxaliplatin alone in both the parental and resistant cell lines at 48 hours of drug exposure. DNA damage analysis based on apurinic/apyramidic site formation detected by aldehyde reactive probe, showed that cetuximab increased DNA damage caused by oxaliplatin (p<0.0001), independent of the cell line oxaliplatin sensitivity (p=0.2). DNA damage repair genes ERCC-1, XPF and XPD levels were evaluated by western blotting. Our preliminary analysis indicates that cetuximab lowers ERCC-1 and XPF, however XPD was not altered in both the cell types in line with the reports on resistance towards platinum based drugs. Clinical resistance towards platinum based drugs is frequently associated with increased activation of DNA damage repair genes, especially the nucleotide excision repair (NER) pathway. DNA damage repair gene expression and activation is dependent on growth factor signaling through Ras and MAPK. ERCC-1, XPD and XPF are the key determinant proteins involved in NER process. EGFR signaling through MAPK can increase the expression of NER genes and blocking this pathway may offer a means to overcome oxaliplatin resistance. Further detailed analysis of this process is in progress. Based on our results, we conclude that cetuximab can overcome oxaliplatin resistance in colon cancer models by increasing DNA damage and oxaliplatin cytotoxicity.