Abstract C74: Stabilization of mutant BRCA1 confers PARP inhibitor and platinum resistance.

Abstract

Abstract The BRCA1 gene is commonly mutated in hereditary breast and ovarian cancers. The BRCA1 protein has multiple domains that mediate protein interactions; BRCA1 gene mutations may produce truncated proteins that lose the ability to interact with associated proteins. Additionally, mutations in the BRCT domain of BRCA1 create protein folding defects that result in protease-mediated degradation. Cells that contain dysfunctional BRCA1 proteins are hypersensitive to DNA damaging agents. In particular, BRCA1-deficient cell lines are exquisitely sensitive to poly(ADP-ribose) polymerase (PARP) inhibitor treatment. Despite initial responses of BRCA1 mutant cancers to PARP inhibitor treatment, acquired resistance develops. To study PARP inhibitor resistance, specifically in the setting of BRCA1 C-terminal truncating mutations, we cultured the triple negative breast cancer cell line MDA-MB-436 (5396+1G>A) in the presence of the PARP inhibitor rucaparib. Drug resistant clones emerged approximately 2-4 months after initial exposure. Clones were highly resistant to rucaparib, and cross-resistant to olaparib, as well as cisplatin. No changes in the mutant BRCA1 DNA or mRNA sequences were detected and the PARP enzyme was efficiently inhibited by rucaparib. The mutant BRCA1 protein was undetectable in MDA-MB-436 parent cells, but was abundant in resistant clones by western blot. HSP90 co-immunoprecipitated with the stabilized mutant BRCA1 protein and treatment of resistant cells with the HSP90 inhibitor 17-DMAG reduced mutant BRCA1 protein levels and restored their sensitivity to PARP inhibition. The stabilized mutant BRCA1 protein also interacted with PALB2-BRCA2-RAD51, was essential for RAD51 focus formation and PARP inhibitor resistance. Resistant cells also acquired a TP53BP1 mutation that facilitated DNA end resection in the absence of a BRCA1 protein capable of binding CtIP. Finally, concomitant increased mutant BRCA1 and decreased 53BP1 protein expression occurs in clinical samples of BRCA1-mutated recurrent ovarian carcinomas that have developed resistance to platinum. These results provide evidence for a novel mechanism by which BRCA1-mutant tumors acquire anti-cancer therapy resistance. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C74. Citation Format: Neil Johnson, Shawn F. Johnson, Yifan Wang, Andrea Bernhardy, Wei Yao, Young-Eun Choi, Marzia Capelletti, Kristopher Sarosiek, Lisa Moreau, Dipanjan Chowdhury, Joyce Liu, Alan D'Andrea, Alexander Miron, Anneka Wickramanayake, Maria Harrell, Elizabeth M. Swisher, Geoffrey I. Shapiro. Stabilization of mutant BRCA1 confers PARP inhibitor and platinum resistance. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C74.