Design and synthesis of RAD51-BRCA2 disruptors to inhibit homologous recombination and achieve synthetic lethality in pancreatic cancer cells

Synthetic lethality (SL) is a phenotypic condition in which the cell does not survive due to the combination of two specific genetic perturbations, which taken individually do not induce cell death. One straightforward application of SL in drug development is the use of the PARP inhibitor Olaparib in oncology patients with BRCA mutations. PARP is involved in the repair of single-strand breaks (SSBs), whereas BRCA2 is essential for repairing DNA double-strand breaks (DSBs) by homologous recombination (HR) as it recruits RAD51 from the cytosol. Due to cancer cells genomic instability, the simultaneous impairment of both DNA repair pathways triggers the SL. In this context, as a new anticancer drug discovery concept, we propose to trigger a fully small molecule induced synthetic lethality combining Olaparib with a BRCA2-RAD51 protein-protein (PP) interaction disruptor that mimics BRCA2 mutation.1 In-depth studies of the target structures identified two PP critical “hotspots” on RAD51 surface, zone I and II, as suitable targets for the design of small molecule PP interaction inhibitors.2 Following a structure-based approach focused on zone II,3 a virtual screening (VS) campaign allows us to identify a potential quinolinic-based hit compound. Compound ARN22142 (Figure A) showed to inhibit BRCA2-RAD51 PP interaction in biochemical and biophysical assays. Moreover, it was able to inhibit HR in BRCA2-proficient pancreatic cancer cell lines (BxPC-3). To discover more effective compounds and depict general structure-activity relationship (SAR) studies, a library of new quinolinic analogues was synthesized. To promote sustainable chemistry, we privileged protocols based on green chemistry principles, which include microwave-assisted and solvent-free reactions and in parallel synthesis. Among the different synthetized compounds, derivative ARN26912 showed a high ability to disrupt RAD51-BRC4 interaction by ELISA assay and an increasing dose-response HR inhibition. Moreover, the compound was synergistic on the antiproliferative efficacy in combination with Olaparib in BRCA2-proficient pancreatic cancer cell lines (BxPC-3 and HPAC) (Figure B). This supports the idea that small organic molecules can mimic genetic mutations. Further in vitro biological studies will be done with this promising compound.