A51

Abstract

The purpose of the study was to assess parameters of local cellular immunity, expression of apoptosis-controlling proteins and some other receptors in various molecular subtypes of breast cancer (BC) with different clinical course and prognosis.

Materials and methods

100 BC patients aged 30–76 years (59 ± 3.4) were recruited. Tissues of tumor and peritumoral area were homogenized by MediMachine, subset contents of T, B, NK-lymphocytes were estimated by flow cytometer FACS CantoII. Percentage of lymphocytes expressing CD3, CD4, CD8, CD19, CD16/56, were counted from total amount of CD45+ lymphocytes. Sections of paraffin-embedded blocks were studied by immunohistochemical method with polyvalent HRP-DAB detection system. Staining was performed using Thermo Scientific autostainer. Tumor was considered p53-positive when >25% of tumor cell nuclei were positively stained, and bcl-2-positive when >25% of cells showed specific cytoplasmic staining. Expression of E-cadherin was assessed by the number of cells with positive membrane expression of this marker, taking into account intensity of the reaction. Expression of topoisomerase 2α and androgen receptors was assessed by the number of tumor cells with positive nuclear expression of these markers; number of stained nuclei per 100 nuclei in 3 fields of view was counted.

Results

Some differences characterizing tumor cells of molecular breast cancer subtypes were found. Tumor tissue contained higher amount of T-lymphocytes than blood (85.9 ± 2.36% vs 60.1 ± 4.5%, P < 0.05) predominantly CD3+CD8+ cells (34.0 ± 1.9% vs 20.3 ± 5.36%, P < 0.05) but lower percentage of B- and NK-cells. In tissue of peritumoral area amounts of CD3+ and CD3+CD8+ cells were also higher, while CD19+ level was lower than in blood. In the same samples content of CD3+CD4+ was lower and amount of NK-cells was higher than in tumor tissue. Triple- negative subtype was characterized by maximal content of CD3+ lymphocytes (92.2 ± 1.4%), luminal A contained the highest amount of CD3+CD16/56+ cells (10.6 ± 1.73%) vs luminal B (4.34 ± 1.27%) and triple-negative (4.13 ± 1.63%). Her2+ neu BC demonstrated high amount of NK-cells in peritumoral area but not in tumor.

p53 overexpression was equally often recorded in luminal B and Her2+ subtypes of BC and slightly less frequently in triple-negative subtype. Accumulation of p53 was 2 times less frequently detected in luminal A subtype in comparison with triple -negative cancer and 2.5 and 2.7 times less frequently in comparison with luminal B and Her2+ BC, respectively. Study of bcl-2 expression showed reducing the frequency of positive tumor cell staining in dependence on the receptor status of tumor: positive cases were several times less frequent in triple-negative and Her2+ subtypes in comparison with luminal A and B subtypes.

Luminal A subtype was characterized by the minimal p53, topoisomerase 2$\alpha$ and androgen receptor expression and maximal expression of bcl-2 and E-cadherin. Luminal B subtype showed a high expression of p53, bcl-2 and androgen receptors and moderate expression of topoisomerase 2$\alpha$ and E-cadherin. High expression of topoisomerase 2$\alpha$, low expression of androgen receptors and moderate expression of E- cadherin were registered in triple-negative BC. The maximal expression of topoisomerase 2$\alpha$ and p53, as well as low expression of bcl-2 and E-cadherin, was observed in Her2+ subtype.

Conclusions

CD8+ T-lymphocytes dominate in tumor tissue of BC. Besides, tumor tissue of luminal A BC contains more NK-cells than other subtypes. A number of differences in expression of apoptosis-controlling proteins was observed in various molecular subtypes of breast cancer. Luminal A BC was noted for the minimal p53 expression and maximal bcl-2expression. Luminal B subtype had a high p53 and bcl- 2 expression. In Her2+ BC p53 expression was maximal and bcl-2 expression was low. The described differences allow assessing molecular subtypes of breast cancer and predicting the disease course from previously unexplored perspectives.