Abstract 2479: A functionally null RAD51D missense mutation is strongly associated with ovarian carcinoma
Authors: Rivera Barbara , Iorio Massimo R. Di , Frankum Jessica , Nadaf Javad , Fahiminiya Somayyeh , Arcand Suzanna L. , Burk David , Grapton Damien , Tomiak Eva , Hastings Valerie , Hamel Nancy , Wagener Rabea , Aleynikova Olga , Giroux Sylvie , Hamdan Fadi F. , Orthwein Alexandre , Zogopoulos George , Rousseau Francois , Berghuis Albert , Provencher Diane M. , Rouleau Guy A. , Michaud Jacques L. , Mes-Masson Anne-Marie , Majewski Jacek , Bens Susanne , Siebert Reiner , Narod Steven , Akbari Mohammad , Lord Chris J. , Tonin Patricia N. , Dionne-Laporte Alexandre , Foulkes William D. 
Published: July 2017

Abstract
Background and goal: RAD51D is a key player in DNA repair by homologous recombination (HR) and carriers of truncating RAD51D mutations have an increased risk for ovarian cancer (OC). However, the contribution of non-truncating RAD51D variants to cancer predisposition remains uncertain. We sought to fully characterize the previously described missense RAD51D variant c.620C&gt;T;p.S207L in order to elucidate its role in OC.
Methods: A clinical panel screening was used to identify the RAD51D variant c.620C&gt;T;p.S207L in two French Canadian (FC) kindred affected with familial High Grade Serous Cancer (HGSC) of the ovary or endometrium. High resolution melting, TaqMan genotyping and Sanger sequencing were used to genotype the p.S207L variant in a series of unselected cases of HGSC of the ovary and endometrium, breast, pancreas and colorectal cancer and healthy controls, all of a FC origin. Whole exome sequencing (WES) was performed to study the genetic signature characterizing RAD51D associated tumors. RAD51 foci formation and CRISPR-Cas9-stimulated and HR-mediated gene targeting assays were used to assess HR activity of RAD51D-S207L mutated CHO cells. HR activity in RAD51D-S207L mutated human cells was tested by a DR-GFP assay. The effect of RAD51D p.S207L on RAD51D-XRCC2 interactions was analyzed by co-immunoprecipitation and quantified in-vivo in a single cell colocalization assay. Sensitivity to PARP inhibitors (PARPi) was evaluated in a cell survival assay.
Results: Using deep sequencing and case-control genotyping studies, we showed that the missense RAD51D variant c.620C&gt;T;p.S207L is over-represented in the French Canadian population affected by HGSC of the ovary (3.8% cases vs 0.002% controls; p &lt; 0.0001).The frequency of the p.S207L variant did not differ from that of controls in breast, endometrial, pancreas and colorectal adenocarcinomas. A common haplotype shared by all the carriers suggested a founder origin for c.620C&gt;T;p.S207L mutation. WES analysis of RAD51D tumor profiles revealed the presence of signature 3 which is known to be associated with HR defects. Functionally, we show that this mutation impairs HR by disrupting the RAD51D-XRCC2 interaction and confers PARP-inhibitor sensitivity to CHO cells.
Conclusions: This work identifies RAD51D p.S207L as the first bona fide pathogenic missense susceptibility allele for HGSC of the ovary and supports the use of targeted PARPi therapies in OC patients carrying missense RAD51D mutations.
Citation Format: Barbara Rivera, Massimo R. Di Iorio, Jessica Frankum, Javad Nadaf, Somayyeh Fahiminiya, Suzanna L. Arcand, David Burk, Damien Grapton, Eva Tomiak, Valerie Hastings, Nancy Hamel, Rabea Wagener, Olga Aleynikova, Sylvie Giroux, Fadi F. Hamdan, Alexandre Orthwein, George Zogopoulos, Francois Rousseau, Albert Berghuis, Diane M. Provencher, Guy A. Rouleau, Jacques L. Michaud, Anne-Marie Mes-Masson, Jacek Majewski, Susanne Bens, Reiner Siebert, Steven Narod, Mohammad Akbari, Chris J. Lord, Patricia N. Tonin, Alexandre Dionne-Laporte, William D. Foulkes. A functionally null RAD51D missense mutation is strongly associated with ovarian carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2479. doi:10.1158/1538-7445.AM2017-2479