Abstract 3710: Synergy between PARP and Wee1 inhibitors suggests homologous recombination repair defect in NSCLC as a mechanistic target for combination therapy
Authors: Yang Daniel X. 
Published: July 2016

Abstract
Advanced non-small cell lung cancer (NSCLC) is the leading cause of cancer mortality. Despite progress in targeted molecular therapeutics and precision medicine, outcomes in this disease remain poor. Recent evidence suggests that impaired homologous recombination (HR) occurs in a significant subset of NSCLCs and may serve as a predictive biomarker for sensitivity to DNA damaging agents. Poly-ADP ribose polymerase (PARP) and Wee1 inhibition represent two mechanistically distinct approaches to augment the effects of DNA damage. Specifically, the PARP inhibitor olaparib impairs repair of DNA single strand breaks, which during replication lead to the formation of DNA double strand breaks (DSBs), resulting in synthetic lethality in HR deficient tumors. AZD1775 is a Wee1 inhibitor that abrogates the G2 checkpoint and thus removes a safeguard against cell cycle progression with unrepaired DNA damage. Moreover, AZD1775 has been recently reported to exhibit single-agent activity in patients harboring BRCA1/2 mutations. Therefore, we hypothesize that olaparib and AZD1775 would have synergistic effects in a subset of NSCLCs and that HR deficiency could be predictive of tumor response to combination therapy.
Utilizing Rad51 focus formation as a marker of HR deficiency, we prospectively selected representative NSCLC cell lines that either did (e.g. Calu6) or did not (e.g. A549) harbor putative defects in HR repair. We treated Calu6 and A549 and other NSCLC cells with AZD1775 and olaparib with varying drug dosing and sequencing to determine the optimal regimen for synergistic effect. Cytotoxicity was determined by CellTiter-Glo cell viability assays and synergy was quantified by calculating the combination index. Additionally, we investigated mechanistic protein markers by Western blot.
In response to combined olaparib and AZD1775 treatment, Calu6 cancer cells demonstrated markedly more pronounced synergistic sensitivity (median CI = 0.19) compared to A549 cancer cells (median CI = 0.90). Moreover, a similar trend toward a selective synergistic effect was demonstrated in a panel of 10 additional NSCLC lines. On biochemical analysis, we observed inhibition of p-Cdk1, upregulation of p-Chk1, and upregulation of p-KAP1, suggesting abrogation of the G2/M checkpoint and activation of ATM/ATR repair pathways, all consistent with the mechanistic underpinnings of our hypothesis.
Taken together, these results provide early pre-clinical evidence for the rational combination of Wee1 and PARP inhibition in the treatment of advanced NSCLC, and suggest HR deficiency as a predictive marker applicable to NSCLC. Continued mechanistic investigation and further confirmatory studies are warranted to inform the selection of patients who may maximally benefit from such combination treatment.
Citation Format: Daniel X. Yang. Synergy between PARP and Wee1 inhibitors suggests homologous recombination repair defect in NSCLC as a mechanistic target for combination therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3710.