MK-8776

Resistance to olaparib inevitably develops in ovarian cancer (OC) patients, highlighting the necessity for effective strategies to improve its efficacy. Here, we established a novel olaparib-resistant patient-derived xenograft model of high-grade serous OC with BRCA1/2 mutations and examined the molecular characteristics of acquired resistance and resensitization to olaparib in treatment-naïve tumors in vivo. Olaparib-resistant xenografts were treated with olaparib, ATR inhibitor (ATRi, ceralasertib), CHK1 inhibitor (CHK1i, MK-8776) or their combinations. Proliferation, apoptosis, ATR/CHK1 activity, PARP signaling, DNA damage response (DDR), epithelial-to-mesenchymal transition (EMT), and MDR1 expression, were examined via RT-qPCR, western blot, and immunohistochemistry. Resistant tumors exhibited defects in PARP and ATR/CHK1 signaling, accompanied by altered expression of proteins involved in DDR and EMT. Olaparib rechallenge combined with ATR/CHK1 inhibitors showed promising synergistic effects on tumor growth inhibition. Mechanistically, combined treatments suppressed tumor proliferation without increasing apoptosis or necrosis, while inducing tumor cell vacuolization indicative of cell death. ATRi combined with olaparib induced or augmented downregulation of ATR, CHK1, PARP1, PARG, BRCA1, γH2AX, and PARylated protein expression, while reversing olaparib-induced upregulation of vimentin, BRCA2, and 53BP1. Our collective findings indicate that ATR/CHK1 pathway inhibition restores the olaparib efficacy in resistant BRCA1/2^MUT high-grade serous OC, highlighting promising approach for olaparib rechallenge of non-responsive patients. Uncovered mechanisms might improve our understanding of acquisition and overcoming resistance to olaparib in ovarian cancer.

The levels and activities of the DNA/RNA helicase schlafen11 (SLFN11) and the serine/threonine-protein kinase ataxia telangiectasia and Rad3-related protein (ATR) may determine cancer cell sensitivity to DNA damaging agents, including platinum drugs. Here, we studied the roles of SLFN11 and ATR in cisplatin resistance of ovarian cancer using cell lines displaying acquired or intrinsic cisplatin resistance. W1CR, the cisplatin-resistant subline of W1 ovarian cancer cells, displayed reduced SLFN11 levels. HDAC inhibition using entinostat returned an epigenetic downregulation of SLFN11 in W1CR cells, caused SLFN11 re-expression and re-sensitized these cells to cisplatin. Moreover, entinostat also sensitized intrinsically resistant EFO21 ovarian cancer cells to cisplatin by upregulating SLFN11. However, SLFN11 was not involved in cisplatin resistance in all other cell models. Thus, SLFN11 expression is not a general cisplatin resistance marker in ovarian cancer. In contrast, inhibition of the DNA damage repair master regulator ATR using sub-toxic concentrations of elimusertib sensitized parental cell lines as well as intrinsically resistant EFO21 cells to cisplatin, and fully reversed acquired cisplatin resistance in cisplatin-adapted sublines W1CR, A2780cis, and Kuramochi^rCDDP²⁰⁰⁰. Mechanisms underlying ATR-mediated cisplatin resistance differed between the cell lines and included CHK1/WEE1 signaling and induction of homologous recombination. In conclusion, SLFN11 and ATR are involved in ovarian cancer cisplatin resistance. Although our data identify ATR as key target for tackling cisplatin resistance in ovarian cancer, future studies are needed to identify biomarkers that indicate, which individual ovarian cancers benefit from SLFN11 re-activation and/or ATR inhibition.