Abstract P2-04-04: BRCA gene mutations do not shape the extent and organization of tumor infiltrating lymphocytes in triple negative breast cancer
Authors: Willard-Gallo K , Solinas C , Marcoux D , t'Kint de Roodenbeke D , Garaud S , Van den Eynden G , de Wind A , Boisson A , Larsimont D , Piccart M 
Published: March 2017

The remarkable responses observed in metastatic cancer patients treated with immunotherapies, including inhibitors directed to the PD-1 and PD-L1 checkpoint molecules, makes it a priority to identify critical variations in pro- and anti-tumor immune responses in breast cancer (BC). In patients with triple negative (TN) BC, an increased presence of tumor infiltrating lymphocytes (TIL) and tertiary lymphoid structures (TLS) have been associated with good clinical outcomes. However, the frequency of specific lymphocyte subpopulations, PD-1 and/or PD-L1 expression and their prognostic significance remains an open question. Our recent work found that PD-1 and PD-L1 expression are specifically associated with higher TIL densities and an increased number of TLS in BC. We further demonstrated that TIL density, TLS and PD-L1 expression were correlated with more aggressive breast tumor characteristics, including higher proliferation and hormone receptor negativity. In this project, we examined the prevalence of TIL, TLS, PD-1 and PD-L1 expression in TNBC and further compared these immune parameters between TNBC patients harboring BRCA1 or BRCA2 germline gene mutations with those carrying the wild-type (wt) genes.

A total of 1402 BC patients whose blood was genetically tested for germline BRCA1 and BRCA2 mutations were examined for inclusion in this study. Ninety-eight chemotherapy-naïve patients with primary invasive ER–, PR– and HER2– BC and demonstrated germline BRCA1 or BRCA2 wt or mutated-gene status were included in this study. Ninety-four tumors were determined to be suitable for evaluating immune cell infiltration (51 BRCA wt and 43 BRCA-mutated). FFPE tumor tissue from the surgical specimens was analyzed by immunohistochemistry (IHC) staining of full-face tissue sections. IHC was performed as a dual label using CD3 plus CD20 for T and B cells, CD4 plus CD8 for the major T cell subpopulations and PD-1 plus PD-L1 for individual or paired expression of these receptors. The stained slides were independently scored by two experienced pathologists for TIL, TIL subpopulations, TLS and checkpoint molecule expression.

These analyses revealed that 87% of our TNBC cohort was TIL-positive (≥10% TIL) with 35% classified as lymphocyte predominant BC (LPBC; ≥50% TIL). T cells were the principal component of the lymphocytic infiltrate with no significant differences between the BRCA wt and BRCA-mutated groups detected in total T cells (CD3+), helper T cells (CD4+), cytotoxic T cells (CD8+) or B cells (CD20+). TLS were identified in 73% of tumors with again no significant differences between the BRCA groups. Examination of checkpoint molecule expression identified 33% tumors as PD-1 positive and 40% as PD-L1 positive. PD-1 expression was correlated with PD-L1 expression and both with TIL positivity and the level of immune infiltration but not BRCA mutational status.

Overall, our analyses revealed that BRCA wt and BRCA-mutated TNBC are remarkably similar in terms of TIL heterogeneity, a TLS presence and checkpoint molecule expression. These data suggest that BRCA gene mutations are not immunogenic nor do they directly drive immune infiltration in TNBC.

Citation Format: Willard-Gallo K, Solinas C, Marcoux D, t'Kint de Roodenbeke D, Garaud S, Van den Eynden G, de Wind A, Boisson A, Larsimont D, Piccart M. BRCA gene mutations do not shape the extent and organization of tumor infiltrating lymphocytes in triple negative breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P2-04-04.