Abstract LB273: Pharmacological synthetic lethality by co-inhibition of PARP and HDAC enzymes

Abstract Introduction: Inhibition of poly(ADP-ribose) polymerase (PARP) induces synthetic lethality in cells exhibiting defects in homologous repair (HR) pathways, such as BRCA1 and BRCA2-mutated breast or ovarian cancers, and 4 different PARP inhibitors (PARPi) are currently approved by the FDA. While HR-deficient cancers usually respond well to single agent PARPi, HR-proficient cancers are generally less sensitive to the treatment. Acquired resistance to PARPi due to activating mutations in BRCA genes are also not uncommon. In this work, we have examined strategies for enhancing PARPi activity in a HR-proficient context and identified histone deacetylases (HDACs) as targets for pharmacological synthetic lethality in the context of PARPi. Materials and methods: PARP1 and PARP2 activity were measured using Trevigen Universal Colorimetric PARP Assay Kit, BPS Bioscience PARP2 Colorimetric PARP2 Assay Kit, and PARylation assay. HDAC activity was measured using HeLa cell nuclear extracts and a fluorogenic peptide-based biochemical assay. Cell survival EC50 were determined using CellTiter Glo viability assay. Cell cycle analysis was performed by flow cytometry with propidium iodide staining. DNA damage was investigated by immunoblotting and immunofluorescence using gamma-H2AX antibodies, and comet assays for confirmation of double-strand breaks. Spheroid assays were performed using the Incucyte® S3 spheroid analysis module. In vivo metastasis formations were examined using the pulmonary metastasis assay (PuMA). Results and discussion: Combinations of PARP and HDAC inhibitors in HR-proficient tumor cells demonstrated that HDACi could enhance the efficacy of PARPi in vitro. This was associated with increased acute and sustained DNA-damage distinct cell cycle profiles showing activity in both S- and G2/M-phases. Importantly, dual single-molecule PARP-HDAC inhibitors showed potent inhibition of PARP1 and PARP2 activity and PAR synthesis, comparable to olaparib, as well as inhibition of HDAC with IC50 values in the low µM range. Dual PARP-HDAC inhibition decreased survival of HR-proficient cells in both 2D cultures and 3D spheroid assays with lower EC50 values than olaparib or vorinostat alone. Importantly, lung metastasis formation was significantly impaired by dual PARP-HDAC inhibition. Conclusion: Combination of PARP inhibition and HDAC inhibition showed increased efficacy compared to single agent PARP inhibition in HR-proficient cells. Development of bifunctional inhibitors may provide a novel therapeutic opportunity for tumors with limited response rates to single agent PARPi. Citation Format: Sarah Truong, Louise Ramos, Beibei Zhai, Fariba Ghaidi, Mona Marzban, Hans Adomat, Xiaoqi Chen, John Langlands, Dennis Brown, Jeffrey Bacha, Colin Collins, Poul H. Sorensen, Wang Shen, Mads Daugaard. Pharmacological synthetic lethality by co-inhibition of PARP and HDAC enzymes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB273.