The chemotherapy agent carboplatin induces DNA damage in the form of crosslinks resulting in apoptosis. Global genome repair (GGR) recognises this DNA damage and triggers apoptosis. In melanoma GGR expression is not induced in response to carboplatin resulting in resistance to this treatment. It is not known what causes the lack of induction of the GGR genes XPC, DDB1 and DDB2, but there is evidence methylation plays a role. 5-aza-2’-deoxycytidine (decitabine), is a DNA methyltransferases inhibitor used as a chemotherapy agent that results in global loss of methylation and re-expression of silenced genes. We hypothesised that restoring expression of XPC, DDB1 and DDB2 in melanoma using decitabine could overcome resistance to carboplatin-induced apoptosis.
Melanoma cell lines were treated with decitabine (0.26µM, 72 hours) or carboplatin (8µg/mL, 48 hours) alone, and in sequential combination for 5 days. qPCR was used to quantify GGR transcript expression before and after treatments and apoptosis was quantified using Annexin V apoptosis kit and flow cytometry. Global methylation was quantified using an ELISA assay.
Treatment with carboplatin alone did not significantly induce XPC, DDB1 or DDB2 expression or increase apoptosis in the majority of melanoma cell lines. Decitabine alone decreased global methylation (average 38.22±4.98% decrease) and increased XPC expression (range = 1.2-2.4 fold). When carboplatin was administered following decitabine, a greater XPC induction (range = 1.1-3.2 fold) occurred and significantly increased levels of apoptosis (range = 1.6-2.2 fold) and decreased proliferation in the majority of cell lines.
Silencing of XPC by methylation is a possible mechanism of carboplatin resistance in melanoma and sequential combination treatment of decitabine followed by carboplatin may be used restore GGR function and overcome resistance