Abstract 3908: DHX9 inhibition as a novel therapeutic for cancer with loss-of-function mutations in DNA damage repair genes BRCA1 and BRCA2

Abstract

Abstract DHX9 is a DEAH-box RNA helicase which can unwind regions of double-stranded DNA and RNA but has a greater propensity for secondary structures such as DNA/RNA hybrids (R-loops), circular RNA and DNA/RNA G-quadruplexes. Given the delicate balance of R-loop formation and resolution in maintaining efficient transcription and replication, the ability of DHX9 to unwind R-loops is important in helping to maintain genomic stability. In addition, DHX9 can interact and regulate a large variety of proteins, including key proteins in DNA damage repair pathways such as BRCA1, ATR, Ku86, and WRN. Previously we demonstrated that DHX9 inhibition was efficacious in microsatellite high (MSI-H) CRC xenograft tumor models. Efficacy attributed to DHX9 loss in MSI-H correlates with defective DNA repair pathways such as mismatch repair (MMR). Here we report results of preclinical studies in ovarian and breast cancer models that indicate patients with Loss-of-Function (LOF) mutations in the DNA damage repair genes BRCA1 and/or BRCA2, may also benefit from DHX9 inhibitor treatment. DHX9 small molecule inhibitor ATX968 was tested for anti-proliferative activity in a large cell panel of 300 cell lines, and bioinformatic analyses was performed to identify molecular variants that co-associate with sensitivity or resistance cell proliferation outcomes. Notably, selective dependency on DHX9 was observed in both ovarian and breast cancer cell lines that exhibit BRCA LOF, as defined by somatic mutations including single-nucleotide variants and/or copy number loss in BRCA1 and/or BRCA2. DHX9 inhibition leads to increased RNA/DNA secondary structures such as R-loops and G-quadruplexes, resulting in subsequent DNA damage and increased replication stress. Cell lines that exhibit BRCA LOF appear unable to resolve this replication stress and show S-G2 phase cell cycle arrest prior to onset of apoptosis. Furthermore, a potent and selective DHX9 inhibitor tool compound was dosed orally in vivo to assess DHX9 dependency within multiple human xenografts representing triple negative breast cancer and high-grade serous ovarian cancer with BRCA LOF. In all models, the tool DHX9 inhibitor was dosed orally at 100 mg/kg BID and it was well tolerated for a period of up to 28 days. Robust and significant tumor growth inhibition and regression was observed in multiple BRCA LOF models with minimal tumor growth inhibition observed in BRCA1 and BRCA2 wild type xenograft models. These results extend the opportunity for DHX9 inhibition to provide therapeutic benefit for patients with solid tumors beyond what was previously reported for MSI-H CRC. Together, this preclinical data package validates DHX9 as a tractable new target with potential utility as a novel treatment for patients with defective DNA repair, such as MMR and BRCA1 and/or BRCA2 LOF, across multiple tumor types including colorectal, breast and ovarian cancer. Citation Format: Jennifer B. Castro, Matthew H. Daniels, Sunaina Nayak, Monique Laidlaw, David Brennan, Brian T. Johnston, Jie Wu, Anugraha Raman, Chuang Lu, Stephen J. Blakemore, Serena J. Silver, P. Ann Boriack-Sjodin, Kenneth W. Duncan, Jason A. Sager, Robert A. Copeland. DHX9 inhibition as a novel therapeutic for cancer with loss-of-function mutations in DNA damage repair genes BRCA1 and BRCA2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3908.