Abstract 2823: DNA methyltransferase inhibitors sensitize NSCLC cells to PARP inhibitors by induction of a double strand break repair defect
Authors: Rachel Abbotts, Michael Topper, Daniel Fontaine, Christopher Biondi, Eun Yong Choi, Rena Lapidus, Stephen Baylin, Feyruz Rassool
Journal: Cancer Research (2018)

Abstract Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death in the US. Poly (ADP-ribose) polymerase inhibitors (PARPi) show efficacy as radiosensitizers in NSCLC, but arguably have gained greater importance in cancer therapeutics by exhibiting synthetic lethality (SL) in tumors with defects in the homologous recombination (HR) pathway of double strand break repair (DSBR). Mechanistically, PARPi cytotoxicity is partially mediated through trapping of PARP1 at sites of DNA damage, with next generation PARPis such as talazoparib (Tal) being potent PARP trappers. Our group has previously demonstrated that combining Tal with a DNA methyltransferase inhibitor (DNMTi) enhances PARPi cytotoxicity in acute myeloid leukaemia and breast cancer models. DNMTis are cytosine analogs that become incorporated into replicating DNA, where they covalently bind their target enzyme DNMT1. When combined with PARPi, DNMTis enhance PARP trapping at damage sites, which collapse into cytotoxic double strand breaks (DSB) when encountered by a replication fork. In the current study, we demonstrate synergistic cytotoxicity of Tal in combination with the DNMTi 5-azacytidine (Aza) in NSCLC cell lines by combination index analyses and colony forming assays. Further potentiation of Tal+Aza cytotoxicity is observed following induction of DNA damage via IR. These results were substantiated in an A549 xenograft model (n=8 per group) where Tal+Aza+IR significantly reduced tumor volume and increased survival compared to vehicle (p&amp;lt;0.001) or Tal+Aza without IR (p&amp;lt;0.05). We and others have previously reported that low dose DNMTi results in stable transcriptional changes in the cancer epigenome, including DNA damage response pathways. Here, we show by microarray analysis that NSCLC cell lines (A549, H23, H1299, H460) treated with Aza exhibit up- and downregulation of multiple genes in the DNA repair reactome, including decreased expression of the key HR-related gene FANCD2, and the non-homologous end joining (NHEJ) gene Ku80. These results were validated by mRNA and protein quantification in Aza-treated NSCLC cell lines and xenograft tumor samples. In keeping with a FANCD2-associated HR defect, Aza exposure is associated with reduced accumulation of RAD51 foci following IR, and reduced HR activity in an extrachromosomal HR assay. Additionally, increased sensitivity to IR post Aza treatment can be explained by decreased expression of NHEJ factor Ku80. These results indicate that Aza treatment in NSCLC induces a DSBR deficiency that can be targeted for SL by PARPi and can further sensitize to IR. Given these agents are currently in use in the clinic, this combination offers a promising therapeutic strategy that could be rapidly translated into clinical practice. Citation Format: Rachel Abbotts, Michael Topper, Daniel Fontaine, Christopher Biondi, Eun Yong Choi, Rena Lapidus, Stephen Baylin, Feyruz Rassool. DNA methyltransferase inhibitors sensitize NSCLC cells to PARP inhibitors by induction of a double strand break repair defect [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2823.