Abstract Germline mutations in BRCA2 predispose to breast, ovarian and other cancers. Although most mutations in the gene are known to be pathogenic, there is an increasing number of missense variants for which the contribution to cancer risk is unclear. These missense substitutions are referred to as VUS (variants of uncertain significance). Because loss of BRCA2 HR function leads to defective DNA double-strand break repair and an increased risk of cancer, we evaluated the utility of a cell-based Homologous Recombination (HR) assay for determination of the cancer relevance of BRCA2 VUS. Specifically, we studied VUS localized in the BRCA2 DNA binding domain (DBD) for effects on HR. We transfected full length wild-type and mutant BRCA2 constructs into V-C8 BRCA2 deficient cells along with an I-Sce1-expressing plasmid which induces an unique double strand break in an integrated DR-GFP reporter plasmid. We evaluated the ability of each BRCA2 mutant to repair the induced break by quantifying the GFP positive cells, generated by HR repair of the DNA break, with fluorescent activated cell sorting (FACS). To establish the sensitivity and specificity of the HR assay we evaluated a series of known deleterious and known neutral with the HR assay. These mutations of known effect were identified using a multifactorial likelihood approach, based on available family data, with a prior probability approach based on sequence conservation (A-GVGD) algorithm (http://agvgd.iarc.fr/agvgd_input.php). We calculated the total odds of pathogenicity for co-segregation of variants with the disease in affected carrier families, co-occurrence of VUS (in trans) with deleterious mutations and for family history of cancer of VUS and combined this odds with the A-GVGD based prior probabilities to generate the posterior probability of pathogenicity for each variant. Using this approach we successfully classified 31 VUS as either neutral or deleterious. By comparing these data with results from the HR assay we established that the assay is 100% sensitive (95% CI: 75.3% – 100%) and 100% specific (95% CI: 82.4% – 100%) for deleterious mutations in the BRCA2 DBD domain. In summary we evaluated the sensitivity and specificity of the HR assay for VUS in the DBD relative to a probability model based on family data and A-GVGD algorithm and we showed that the assay was highly sensitive and specific. Based on these results we propose that the HR assay can be used as a classifier of the clinical relevance of the VUS in the absence of direct genetic and family data. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5591. doi:10.1158/1538-7445.AM2011-5591
Novel late-onset Alzheimer disease loci variants associate with brain gene expression
Abstract B39 Background The breast glandular epithelium is composed of lobules that undergo age-related involution that increases as women age and has been associated with lowered breast cancer risk. Mammographic density (MD) is an established risk factor for breast cancer that has been shown to decrease with age. This study examined the association of age-related lobular involution with mammographic breast density. Materials and Methods The study sample was selected from approximately 20,000 healthy women who were enrolled in a screening cohort at the Mayo Clinic (Mayo Mammography Health Study) from 2003 to 2006 and had a benign breast biopsy (BBD) within 2 years of the enrollment mammogram. A quantitative measure of mammographic density, percent density (PD) was estimated on the enrollment mammogram, in addition to its components dense and nondense area (NDA). All biopsies were reviewed by a breast pathologist for the type of benign breast disease (BBD) as well as lobular involution status. Involution was defined as the proportion of terminal ductal lobular units (TDLUs) in the background tissue on biopsy that showed at least 50% decrease in size and number of acini from normal lobules and was subjectively assessed as: No- 0% involution, Partial- 1 to 74%, or Complete- >75% involution. Mean density was estimated for the 3 categories of involution using generalized linear models and square root transformation. Confounders were assessed including age, parity, body mass index (BMI), type of BBD, menopausal status, age at first birth and family history Results A total of 317 women from the MMHS had BBD within 2 years of enrollment. Mean age at BBD diagnosis in this sample was 55 years. Age- and BMI- adjusted analyses suggested an inverse association of involution and percent density with adjusted mean PD values of 23.3 (95%CI [17.2, 30.3]) for no involution, 21.9 (95%CI [20.0, 23.9]) for partial involution, and 17.8 (95% CI [14.7, 21.3]) for complete involution (p for trend 0.05). This trend was driven primarily by the positive association of involution with nondense area with adjusted mean NDA values of 103.1cm2 (95% CI [86.1, 121.7]) for no involution, 104.9 cm2 (95% CI [99.4, 110.7]) for partial involution, and 126.2 cm2 (95% CI [115.0, 137.9]) for complete involution (p for trend 0.004). Discussion Our findings suggest that MD is inversely associated with involution of breast tissues, providing a plausible mechanism through which MD influences breast cancer risk. Citation Information: Cancer Prev Res 2008;1(7 Suppl):B39.