Abstract 418: Modelling PARP inhibitor resistance using CRISPR/Cas9 genome engineering in a PARP inhibitor-sensitive organoid model identified by organoid panel screening

Abstract Background: Inhibition of poly(ADP-ribose)polymerase (PARP) is synthetically lethal in cells with impaired homologous recombination (HR) DNA repair Hence, PARP-inhibitors (PARPi) are used to treat tumors with disruptive mutations in HR factors. However, both pre-existing and acquired resistance mechanisms limit the clinical success of PARPi, highlighting the need to develop new drugs and/or treatment strategies. Methods: PARPi efficacy was tested in a CellTiterGlo (CTG)-based OrganoidXplore™ organoid screening panel of 50 organoid models, spanning 7 cancer indications, including models with mutations in HR genes. CRISPR/Cas9-based genome editing was used to knock-out SLFN11 in an originally PARPi-sensitive organoid model, to mimic clinical PARPi resistance. Lastly, isogenic pairs of PARPi sensitive and resistant organoids were used in drug efficacy studies to assess treatment efficacy of multiple therapies. Results: The OrganoidXplore panel screening captured the heterogeneous response to talazoparib that is also observed in patients. 19 models demonstrated sensitivity to talazoparib (IC50s <1 μM), including models carrying inactivating mutations in HR genes (BRCA1, BRCA2 and RAD51C). However, not all models that carry mutations in HR genes displayed PARPi sensitivity and not all sensitive organoid models carry mutations in known HR genes, stressing the importance of in vitro efficacy testing. Inactivating mutations into SLFN11, a well-known clinical biomarker for PARPi and chemotherapy response, was introduced in one of the most PARPi-sensitive models, using CRISPR/Cas9. The isogenic SLFN11 wild-type and knockout pair were used to assess single agent sensitivity to talazoparib, and the chemotherapeutic drugs etoposide, doxorubicin and camptothecin (the parent compound to irinotecan and topotecan), and found that SLFN11 loss causes resistance to all tested agents. Conclusion: In vitro organoid screening successfully identified PARPi-sensitive organoid models containing known and unknown mechanisms of PARPi sensitivity. Further investigation is required to determine the sensitivity of certain models, as they may bear novel biomarkers of PARPi sensitivity, or possess unknown intrinsic resistance mechanisms. Furthermore, CRISPR/Cas-mediated genome editing can be used to establish isogenic organoid pairs that can subsequently be used for single agent as well as combination therapy efficacy testing in vitro. Citation Format: Lenno Krenning, Dorrith Verstegen, Irene Drosou, Marten Hornsveld, Mariusz Madej, Hester Bange, Peter van Schaik, Ludovic Bourre, 2Marrit Putker1. Modelling PARP inhibitor resistance using CRISPR/Cas9 genome engineering in a PARP inhibitor-sensitive organoid model identified by organoid panel screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 418.