Abstract LB-019: BRCA1 protein expression and subcellular localization in primary breast cancer: automated digital microscopy analysis of tissue microarrays

Abstract

Abstract Mutations in BRCA1 are associated with familial as well as sporadic aggressive subtypes of breast cancer. However, it is not clear to what extent BRCA1 expression or its subcellular localization contributes to breast cancer progression. The goal of this analysis was to examine the differential expression and subcellular localization of BRCA1 among normal breast tissue and breast cancer cases, and whether it could serve as an additional prognostic marker in an ethnically diverse sample of 287 patients (86 Non-Hispanic White, 84 Hispanic and 116 African American). Tissue microarrays (TMAs) were constructed from invasive breast cancer samples obtained from the Breast Cancer Care in Chicago study in addition to 46 age and race-matched normal breast tissue samples. In these TMAs, BRCA1 was immunolabeled with Alexa647; epithelial cytoplasm with Alexa488; and nuclei with DAPI. Slides were visualized and quantified using “VECTRA Automated Multispectral Image Analysis System” and “InForm software”. BRCA1 expression was evaluated based on the percentage of positive cells and staining intensity using the H-score. The H score is a product of the percentage of cells (0-100%) in each intensity category (0, 1+, 2+ and 3+). The final score is on a continuous scale between 0 and 300. The average nuclear score for BRCA1 (Nuc) was higher than that of the cytoplasmic score (Cyto) in normal breast tissue (Nuc: 157.7 ±6.0; Cyto: 150.7±7.5, p = 0.02) and breast cancer cells (Nuc: 140.7 ±3.3; Cyto: 118.0±3.4, p<0.0001). Normal breast tissue had higher levels of BRCA1 protein than breast cancer cells for both the nuclear (p<0.01) and the cytoplasmic (p<0.0001) fractions. However, the nuclear to cytoplasmic ratio of BRCA1 protein was significantly higher in breast cancer cells (1.5±0.1) than in normal mammary epithelial cells (1.1±0.1, p = 0.01). This alteration in the nuclear to cytoplasmic ratio may suggest either a role of BRCA1 shuttling in the pathogenesis of breast cancer or the increased need for BRCA1 inside the nucleus to help repairing cancer-induced DNA damage. Cytoplasmic and nuclear BRCA1 expression was then classified as low or high using the mean of the H-score as a cutoff. BRCA1 cytoplasmic and nuclear scores correlated negatively with breast cancer nuclear grade (r2 = -0.2, p = 0.02); 64% of grade 1 breast cancer cases expressed high levels of BRCA1 while only 36% expressed low levels. In contrast, 60% of the cases in grade 2 and 54% of the cases in grade 3 had low levels of BRCA1. Similarly, an inverse correlation was found between high BRCA1 nuclear scores and stage of breast cancer; 53% in stage 1 and 42% of stage 2 (p = 0.05). In addition, high BRCA1 expression significantly correlated with progesterone receptor positivity (P < 0.01), bcl2 positivity (P = 0.01), high androgen receptor expression (p<0.0001) and low Ki67 index (P = 0.04). In conclusion, in this multi-ethnic sample of breast cancer patients, we found that BRCA1 was associated with less aggressive, lower grade disease. Accordingly, evaluation of BRCA1 expression could provide additional clinically relevant information in routine classification of breast cancer. Note: This abstract was not presented at the meeting. Citation Format: Abeer Mostafa Mahmoud, Umaima Al-alem, Virgilia Macias, Ryan J. Deaton, Andre Balla, Peter Gann, Garth Rauscher. BRCA1 protein expression and subcellular localization in primary breast cancer: automated digital microscopy analysis of tissue microarrays. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-019. doi:10.1158/1538-7445.AM2015-LB-019