PD-L1 and HSP-70 molecules are part of immunosupressive environment in the deep layer of the lymphocyte predominant breast cancer (LPBC)

Background: Tumor infiltrating lymphocytes (TILs) are involved in host imunity against tumor cells. However, in later phases of the disease high TIL infiltration is related to disease progression. Tumor immunogenicity is strongly correlated with the higher tumor mutation burden. Triple negative (TN) and HER-2 enriched breast cancers have the highest immunogenic potential so the aim of our study was to investigate the TIL infiltration and expression of PD-L1, HSP-70 in such tumors. Material and Methods: TIL infiltration was investigated in the 112 tissue samples of TN and HER-2 enriched breast cancers of women diagnosed and treated in the Clinical Hospital Centre Rijeka, Croatia, in the period between 2008 and 2016. The invasive front of the tumor (host-tumor interface), the surface layer, as well as the deep layer of the tumor were analysed. Immunohistochemistry staining of PDL-1 (SP142), HSP70 (ab2787), CD4 (SP35 Cell Marque) and CD8 (144B DakoCytomation) was performed. The results were analysed using Statistica 13 software. Results: Overall, there is a statistically significant correlation of high (over 50%) TIL infiltration with longer 5-year survival (p = 0.035, Long rank test). In the surface layer of the tumor (invasive front) there is statistically significant correlation of the intermediate TIL infiltration with the higher survival (p = 0.051, Long rank test) whereas there is no significant difference in the deep layer of the tumor. There is significant association of TIL infiltration with CD8+ T lymphocyte expression in the surface and deep layers of the tumor (Mann Whitney U test, p = 0.004 and p < 0.001, respectively), CD4+ lymphocyte expression (p < 0.001, p < 0.001, respectively) and PDL-1 expression (p < 0.001, p < 0.001, respectively). Statistically significant correlation of TIL infiltration and HSP-70 protein was only detected in the deep tumor layer (Mann Whitney U test, p < 0.001). Furthermore, in the TIL infiltrated deep tumor layer there is statistically significant positive correlation of PD-L1 and HSP-70 expression (Mann Whitney U test, p = 0.029) as well as positive European Journal of Cancer 138, Suppl. 1 (2020) correlation of the HSP-70 expression and the stage of the disease (Anova, p = 0.08). Conclusion: Although TIL infiltration in the surface layer of the tumor is corellated with higher survival rate, there is no such correlation in the deep layer. We have shown that in both layers there is increased expression of CD4 and CD8 positive T lymphocytes. However, the increased expression of inhibitory molecules PD-L1, and in the deep layer HSP-70 protein is noted as well. It is possible that in this context HSP-70 is involved in activation of Tregs and thus inducing immunotolerance to oncoproteins and along with PD-L1 molecule stimulates the development of immunosuppressive environment in the deep tumor layer thus supporting tumor immune evasion.