Enhancement Of Decitabine Cytotoxicity By Methoxyamine Via Inhibition Of Base Excision Repair

AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO Decitabine (5aza-2\#8217;deoxycytidine) is a nucleoside analog used for the treatment of hematological malignancies. Previously, it was shown that the cytotoxic effect of low dose decitabine treatment is due to incorporation of its active metabolite, 5aza-2\#8217;deoxycytidine triphosphate, into DNA leading to inhibition of DNA methylation by binding irreversibly to DNA methyltransferases. We hypothesized that incorporated 5aza-2\#8217;deoxycytidine (or its deaminated analog, 5aza-2\#8217;deoxuridine) into DNA is also recognized and processed by the base excision repair (BER) pathway. In this case, inhibition of BER by methoxyamine (MX) would potentiate the cytotoxicity of decitabine. We evaluated role of BER in decitabine cytotoxicity in colon cancer, melanoma cells, and primary acute myelogenos leukemia (AML) cells. Decitabine-induced abasic sites (AP-sites) were increased proportionally with dose and duration of exposure. MX reduced the number of available AP-sites up to 80% indicating formation of stable MX-bound AP-sites that have the potential to interrupt BER pathway. A similar correlation between decitabine dose and the AP-sites formed was observed after in vitro exposure of primary AML cells to increase concentrations of decitabine. MX was able to bind a significant percentage (up to 60%) of these AP sites. Decitabine cytotoxicity was potentiated by MX. Cell survival assays demonstrate a 4-fold decrease in the IC50for decitabine when cells were co-treated with MX (IC50 dec = 4 \#956;M and IC50 Dec+ MX = 1 \#956;M). Apoptotic cell death measurements using Annexin V staining showed a 5-fold increase in cell death following decitabine and MX treatments. These events were accompanied by a concomitant increase in cleavage of PARP, and in \#947;H2AX. Moreover, MX enhanced decitabine-induced antitumor effect in mice bearing A375 human melanoma xenografts, as measured by tumor growth delay: 7 days (0.5 mg/kg decitabine) versus 14 days (0.5 mg/kg decitabine plus 2 mg/kg MX).These studies not only suggests for the first time the role of BER in the processing of incorporated decitabine, but they also provide insights into a new and promising cancer therapeutic strategy of combining decitabine with MX to block BER. Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr 5547.