Abstract LB-263: Synthetic lethal targeting of PTEN-associated homologous recombination (HR) deficient melanoma cells by human apurinic/apyrimidinic endonuclease (APE1) inhibitors

Abstract

Abstract Background Mutation of the tumour suppressor PTEN is well-characterised in many advanced malignancies, including 30-50% of metastatic melanomas. Emerging evidence suggests a new role for PTEN in DNA repair, through transcriptional downregulation of the critical HR protein RAD51. Tumour cells deficient in HR (such as BRCA-deficient breast and ovarian cancers) have been therapeutically targeted in a synthetic lethality (SL) strategy using PARP inhibitors that block single strand break repair (SSBR), a pathway related to base excision repair (BER). We have previously identified novel inhibitors of the critical BER enzyme APE1, and demonstrated these can induce SL in BRCA-deficient cell lines. In the current study, we have evaluated SL relationships in PTEN-proficient and -deficient melanoma systems for a potential novel treatment strategy. Method We screened a panel of melanoma cell lines (MeWo, MM418, SkMel28, SkMel30, HT144, UACC62) for PTEN, APE1 and RAD51 protein expression. We tested for SL using clonogenic survival assays in PTEN- and RAD51-deficient cell lines using our recently developed APE1 DNA repair domain inhibitors, methoxyamine (indirect APE1 inhibitor) and NU1025 (PARP inhibitor), either alone or in combination. Aldehyde reactive probe assay, neutral comet assay, γH2AX immunofluorescence and FACS analyses were performed to confirm biological activity in cells. Results Of the screened cell lines, SkMel28, HT144 and UACC62 were deficient in PTEN and RAD51 protein expression. APE1 inhibitors were more toxic to PTEN-deficient cells compared to PTEN-proficient cells (p<0.01). Significant sensitivity was also seen in PTEN-deficient cells treated with methoxyamine (p<0.05) and PARP inhibitors (p<0.001), confirming that BER modulation results in SL in PTEN deficiency. This increased cytotoxicity in PTEN deficiency is associated with accumulation of DNA double strand breaks (DSB), as assessed by comet assay (p<0.001) and γH2AX accumulation (p<0.05). DSB accumulation leads to G2/M arrest and apoptosis, as demonstrated by FACS analysis. Conclusion Our data provides the first pre-clinical evidence that targeting APE1 in PTEN-RAD51 deficient melanoma cells is a promising new SL strategy. We are complementing the current small molecule studies by developing PTEN knockout systems using shRNA technology. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-263. doi:1538-7445.AM2012-LB-263