Oral Presentation 29th Lorne Cancer Conference 2017

Secondary mutations in RAD51C and RAD51D as a mechanism of resistance to PARP inhibition in high-grade serous ovarian cancer. (#26)

Olga Kondrashova 1 , Kristy Shield-Artin 1 , Minh Nguyen 2 , Gwo Yaw Ho 1 3 4 , Elizabeth Lieschke 1 , Maria I Harrell 5 , Matthew Wakefield 1 , Elizabeth Swisher 5 , Kevin K Lin 2 , Clare L Scott 1 3
  1. Walter and Eliza Hall Institute, Melbourne
  2. Clovis Oncology, Inc., San Francisco, CA
  3. The Peter MacCallum Cancer Centre, Melbourne
  4. The Royal Women's Hospital, Melbourne
  5. University of Washington, Seattle, WA

Introduction:
PARP inhibition (PARPi) is an effective therapy that targets Homologous Recombination (HR)-deficient cancers, best proven for women with BRCA1/2 mutated high-grade serous ovarian cancer (HGSOC)1. Better understanding of HR defects responsible for PARPi susceptibility and resistance is required to improve clinical management. A well-described mechanism of PARPi resistance in BRCA1/2 mutated cases is that of secondary mutation, which restores BRCA1/2 function and thereby the HR pathway2,3. We have investigated secondary mutations in HR genes as a mechanism of PARPi resistance using tumour biopsies collected from the phase 2 ARIEL2 clinical trial of rucaparib, a potent PARPi, in patients with platinum-sensitive HGSOC.
Methods:
Pre-treatment and post-progression tumour biopsies were collected from seven patients, and sequenced with an NGS-based assay. Effects of the identified primary and secondary mutations on PARPi response were modelled in vitro in PEO4 and OVCAR8 HGSOC cell lines, using Cas9/CRISPR-based gene knockout, followed by expression of mutated transcripts. Tumour tissue was transplanted into mice to generate a patient-derived xenograft (PDX). Whole-genome sequencing, serial biopsy sectioning and deep mutational analysis were performed to study intra-tumour heterogeneity.
Results:
For three of the seven patients, all of whom responded clinically to rucaparib, primary and secondary mutations in BRCA1, RAD51C and RAD51D were identified. For the RAD51C case, four distinct secondary mutations were identified within a single post-progression groin lymph node biopsy, and were shown to be spatially heterogeneous. In vitro modelling of the detected RAD51C mutations in two independent cell lines supported the primary mutation sensitizing the cells to PARPi, while the secondary mutations caused resistance. A RAD51C PDX is being expanded for in vivo analysis.
Conclusion:
While important clinical benefit was achieved in these patients, analysis of post-progression biopsies identified secondary mutations in BRCA1, RAD51C and RAD51D that restored the open reading frame, supporting the role of primary mutations of these HR genes in conferring PARPi sensitivity, and the secondary mutations as a novel mechanism of acquired resistance.

  1. Lee JM, Ledermann JA, Kohn EC: PARP inhibitors for BRCA1/2 mutation-associated and BRCA-like malignancies. Ann Oncol 25:32-40, 2014
  2. Norquist B, Wurz KA, Pennil CC, et al: Secondary somatic mutations restoring BRCA1/2 predict chemotherapy resistance in hereditary ovarian carcinomas. J Clin Oncol 29:3008-3025, 2011
  3. Barber LJ, Sandhu S, Chen L, et al: Secondary mutations in BRCA2 associated with clinical resistance to a PARP inhibitor. J Pathol 229:422- 429, 2013