QUANTITATIVE ESTIMATION OF BRCA1 PROTEIN EXPRESSION IN BREAST CANCER TISSUE USING THE METHOD OF FLOW CYTOMETRY

T. A. Bogush; E. A. Dudko; E. A. Shestakova; A. N. Grishanina; E. A. Bogush; V. Ju. Kirsanov; O. M. Ryabinina; N. O. Vichljantzeva

doi:10.17650/1726-9784-2016-15-4-49-52

QUANTITATIVE ESTIMATION OF BRCA1 PROTEIN EXPRESSION IN BREAST CANCER TISSUE USING THE METHOD OF FLOW CYTOMETRY

T. A. Bogush, E. A. Dudko, E. A. Shestakova, A. N. Grishanina, E. A. Bogush, V. Ju. Kirsanov, O. M. Ryabinina, N. O. Vichljantzeva

https://doi.org/10.17650/1726-9784-2016-15-4-49-52

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Abstract

Introduction. The disruption of BRCA1 function leads to the development of several cancers including breast cancer, ovarian cancer, non-small cell lung cancer, gastric cancer, pancreatic cancer. Recently it was demonstrated that BRCA1 protein as well as another DNA repair protein, ERCC1, could be prognostic markers of the course of the disease and predictive markers of platinum-based chemotherapy efficiency. Nevertheless, the obtained data are controversial. Aim. In current study the comparison of BRCA1 expression in breast cancer with favorable and unfavorable prognosis based on estrogen receptor status (ER+ and ER^- ) was carried out to evaluate its potential prognostic significance for subsets of breast cancer with different platinum chemotherapy sensitivity. Materials and methods. The estimation of BRCA1 expression was made in 50 samples of breast cancer tissue (40 ER+ and 10 ER^-) using the quantitative immunofluorescent method coupled with flow cytometry. Tumor cell suspensions were incubated with primary anti-BRCA1 antibodies (Abs) (SD118, Calbiochem) and secondary fluorescent Abs (DyLight650, ab98729). Cell fluorescence was measured at the Flow Cytometer FACSCanto II (Becton Dickinson, USA) using the program FACSDiva 6.0. The level of BRCA1 expression was analyzed using the program FlowJo 10.0 with statistical approach of Kolmogorov-Smirnov. The level of BRCA1 expression was determined as the ratio of specifically fluorescent cells to the total number of cells. Results. 1. The expression of BRCA1 was revealed in all breast cancer samples with mean expression level 36.8 ± 7.6 % and median 37 %. 2. Using the method of Student t-test statistically significant difference in mean value of BRCA1 expression level (p = 0.02) was revealed between ER+ and ER^- breast cancer groups (38.0 ± 7.5 % and 32.0 ± 6.3 %, respectively) 3. Comparison of BRCA1 expression level in ER+ and ER^- breast cancer groups using Mann - Whitney test also demonstrated statistically significant difference (p = 0.016). 4. These differences in mean value of BRCA1 expression level were especially prominent (p = 0.007) upon the comparison of ER+ and ER- breast cancer groups with BRCA1 expression levels below median (32.4 ± 5.0 % and 26.8 ± 2.9 %, respectively). 5. Comparison of BRCA1 expression levels in ER- breast tumors and the median for all studied breast cancer samples (37 %) demonstrated the correlation of decreased level of BRCA1 expression and negative ER status of the tumor. BRCA1 expression levels below the median were shown for 80 % of breast cancer patients with negative ER status in tumor, and over the median - in 2 0 % of cases, respectively. Conclusion. The results of current studies demonstrated that BRCA1 protein is expressed at different levels in the tumor tissue of all breast cancer patients. Lower level of BRCA1 expression in ER-negative as compared to ER-positive tumors, and established unfavorable prognostic value of ER-negative breast cancer status point to poor prognostic value of lower level of BRCA1 expression.