A RAD51/BRCA2 small molecule inhibitor enhances the antineoplastic effect of the PARPi talazoparib in pancreatic cancer

Introduction: PARP inhibitors (PARPi) are drugs showing efficacy in HR-defective cancers. The RAD51/BRCA2 interaction is a key step in HR, which is the main DNA double strand break (DSB) repair pathway. As this mechanism is activated following single strand break (SSB) repair inhibition by PARPi, the simultaneous impairment of HR and PARP should lead to increased antineoplastic power of PARPi. We characterized the mechanism of action of ARN24922 (1), which is a new promising RAD51/BRCA2 inhibitor, developed in house. Furthermore, we evaluated its potential to increase the effect of talazoparib (TLZ, a potent PARPi) on 2D and 3D models of pancreatic cancer (PAC). Materials and methods: We evaluated the potential of 1 in inhibiting HR activity through the mClover HR assay, which is based on the insertion of a mClover-containing sequence into a Cas9-generated DSB in the LaminA gene; the subsequent DSB repair leads to cell fluorescence. To confirm HR inhibition, we examined RAD51 nuclear foci after the induction of DNA damage by cisplatin (CPL) and RAD51/BRCA2 inhibition by 1. Finally, we studied the antineoplastic effect of the 1-TLZ combination in different PAC cell lines (BxPC-3, HPAC, AsPC-1) and in 3D models of PAC (PT291 and PDM41 organoids), with each of the models being characterized by functional BRCA genes and HR. Results and discussion: Preliminarily, by applying the mClover HR assay, we found that 1 produced an 80% HR inhibition. Moreover, 30 µM 1 caused a significant reduction of RAD51 nuclear foci in CPL exposed cells. This finding is in agreement with the results obtained in the HR assay, supporting the expected mechanism of action of 1. In 2D PAC cultures, we found that 50 µM 1 significantly increased the anti-proliferative effect of 2 µM TLZ. The cell death mechanism involved in this effect was studied in BxPC-3 cells and was characterized as apoptosis. When 1 was co-administered with TLZ, in 3D PAC models, a significant increase in the antineoplastic activity of TLZ was also observed. In addition, in PT291 organoid, 1 was found to increase the DNA damage signatures caused by TLZ, as shown by immunofluorescence detection of γ-H2AX in cell nuclei. Conclusions: The obtained data support the idea that the inhibition of RAD51/BRCA2 interaction can extend the use TLZ to HR proficient tumor forms, highlighting a promising anticancer strategy for PAC.