Abstract B35: A breast cancer susceptibility gene screening panel for variant discovery and gene exclusion in an African American cohort from Alabama
Authors: Chandler Madison R. , Shepp Kasey J. , Spina Stephanie M. , Merner Nancy D. 
Published: March 2017

African American (AA) women are diagnosed with breast cancer (BC) at a higher rate under the age of 40 compared to Caucasian women and are more often diagnosed with triple-negative (TN) BC, a less treatable and more aggressive BC sub-type. Since an early age of onset is a key characteristic of hereditary BC, genetic risk factors are likely contributing towards these BC disparities. Genes that harbor genetic risk variants are referred to as BC susceptibility genes. Prior to searching for novel BC susceptibility genes, previously identified BC susceptibility genes should initially be excluded. Since the introduction of next-generation sequencing, gene-screening panels have been implemented; this innovative approach involves designing probes for target-capture of genes of interest, amplification, and sequencing on a massively parallel array. AA BC genetics is understudied; however, recently, Churpek et al. published the first article to screen AA hereditary/early onset/TN BC cases for a panel of known BC susceptibility genes. This study identified 68 pathogenic germline mutations in 65 of 289 (22%) AA cases; 18% had a BRCA1 or BRCA2 mutation. The authors acknowledged the paucity of their study and that more gene panel screening studies are needed. Currently, it appears that the majority and spectrum of BC susceptibility genes/risk variants that contribute towards the disproportionate number of aggressive and young onset AA BCs remain to be identified.

Our group has custom-designed an Agilent Technologies (AT) HaloPlex probe kit, named BRAP (BReast And Prostate) that targets 87 known and candidate BC and prostate cancer (PC) susceptibility genes using the AT SureDesign program. PC genes were included on the panel since AA males are frequently diagnosed with PC in BC families, more susceptible to PC compared to Caucasians, and normally diagnosed at a younger age with larger tumors. Furthermore, men carrying BRCA1/2 mutations have a higher risk of PC, so further genetic overlap will likely be detected through BRAP screening. Ultimately, the probes were designed to generate 100 base pair (bp) reads on an Illumina platform. All coding exons as well as the 5' and 3' untranslated regions of the 87 genes were targeted, totaling 499.5 kbp. The designed probes covered 98.9% of the targeted region.

Forty-six seemingly unrelated individuals (24 AA and 22 Caucasians) from Alabama who were diagnosed with BC under the age of 45 and/or had a BC family history were selected for the initial BRAP capture; the HaloPlex HS Target Enrichment System For Illumina Sequencing Protocol (Version C0) was followed. Each individually prepared and indexed library was quantified using a Bioanalyzer 2100. The targeted DNA from 44 of the 46 individuals were successfully captured and amplified. The successful samples were pooled for sequencing; the sample was loaded onto one flow cell lane and run on an Illumina HiSeqTM 2500 at the HudsonAlpha Genomic Services Laboratory. The sequencing data was downloaded and is being processed using an in-house, bioinformatics pipeline. Two of the screened individuals served as positive controls since upon recruitment one reported a mutation in BRCA1 and the other a mutation in BRCA2. The pipeline has been optimized using the control with BRCA1 p.M1775R, which has been successfully identified. Variant calling files are being processed for the remaining screened individuals. This data provides great insight towards the array of mutations in known BC and PC susceptibility genes that contributes towards hereditary AA BC. Furthermore, non-mutation individuals/families identified through BRAP screening will be whole-genome sequenced in order to identify novel, BC susceptibility genes that may contribute towards disease disparity.

Citation Format: Madison R. Chandler, Kasey J. Shepp, Stephanie M. Spina, Nancy D. Merner. A breast cancer susceptibility gene screening panel for variant discovery and gene exclusion in an African American cohort from Alabama. [abstract]. In: Proceedings of the Ninth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2016 Sep 25-28; Fort Lauderdale, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2017;26(2 Suppl):Abstract nr B35.