Abstract P5-04-23: Characterization of the tumor microenvironment in a large series of ER/PR negative breast cancer
Authors: Vos Hanne , Lambein Kathleen , Floris Giuseppe , Mariën Bram , Berben Lieze , Neven Patrick , Wildiers Hans , Nevelsteen Ines , Smeets Ann 
Published: March 2020

Abstract
Introduction: Recent data show that a subgroup of patients with breast cancer might benefit from immune checkpoint blockade (ICB). However, adequate biomarkers to select patients for ICB are lacking. A better understanding of the immune microenvironment of breast cancer is paramount in the quest for appropriate biomarkers. We investigated several immune related parameters in hormone receptor negative breast cancer, attempting to characterize the tumor microenvironment. We observed the immune micro-environment from three angles. Firstly, we counted and further characterised stromal tumour infiltrating lymphocytes (sTILs). Secondly, we measured the expression of PD1 and PD-L1. Lastly, the expression of CD73 on tumor cells, a potential regulatory molecule of immune escape, was analysed.
Methods: Resection specimens of 198 patients, diagnosed with ER/PR negative invasive adenocarcinoma of &gt; 2 cm between 2005 and 2010, were analysed. TILs percentages were analysed according to the international guidelines. Immunohistochemical (IHC) analyses of CD3, CD4, CD8, CD68, CD73, FoxP3 and PD1 were assessed. PD-L1 IHC assay and interpretation was performed by QualTek and a modified proportion score (MPS) was used. The immune parameters were correlated with clinicopathological parameters.
Results: Clinicopathological characteristics are summarized in Table 1. The median expression of the immune parameters ranges from 0 to 10 and is presented in Table 2. Further, sTIL percentages were categorised into three groups, with 10% and 30% as cut-off, which matches with respectively 31%, 50% and 19% of the patients. Applying a cut-off of 1%, only 17% of the samples showed PD-L1 expression while 97% of the samples showed PD-1 expression on TILs. As could be expected a significant positive association was observed between sTILs and CD3, CD4 and CD8, but also with CD68, FoxP3 and PD-L1. No association could be noted between PD-1 expression and any of the other parameters. When taking patient age into account, older patients have significant less sTILs and expression of CD3, CD4, CD8 and FoxP3.
An increase in the fraction of sTILs with 10% was significantly associated with a lower risk of metastasis ([HR] = 0.529, p &lt; 0.01). This also reflects in the observation that an increase in CD3, CD8 and FoxP3 lowers the risk of metastasis ([HR] of respectively 0.940 (p &lt; 0.01), 0.876 (p = 0.02) and 0.658 (p = 0.01)). No significant association between CD73 and the risk of metastasis was observed (p = 0.1098).
Table 1: Clinicopathological informationAge (mean)56,36 N%Histological typeNon special type16382,32Special type3517,68 Molecular type Triple negative14271,72HER2 positive5628,28 Tumour grade 110,512115,56318693,94pT217487,8832311,62410,51pN09748,9917638,382136,573126,06
Table 2: Expression of the different immune parametersVariableMedian (%)IQR (%)Range (%)sTILs105 - 305 - 80CD3105 - 205 - 70CD455 - 55 - 20CD855 - 105 - 60CD68105 - 205 - 80CD7300 - 50 - 90FoxP321 - 20 - 10PD-155 - 50 - 30PD-L100 - 00 - 100
Conclusion: The expression of immune parameters is this cohort of patients was rather low. Only 19% have high number of sTILs (&gt;30%) and PD-L1 expression (&gt;1%) was present in 17% of patients. Moreover, the number of sTILs as well as subtypes of lymphocytes decreases with age. Further, our data show a significant positive association between sTILs, subtypes of lymphocytes (CD3, CD4, CD8, FoxP3), macrophages (CD68) and PD-L1. Lastly, high sTIL count, expression of CD3, CD8 and FoxP3 were associated with a decreased risk of metastasis.
Citation Format: Hanne Vos, Kathleen Lambein, Giuseppe Floris, Bram Mariën, Lieze Berben, Patrick Neven, Hans Wildiers, Ines Nevelsteen, Ann Smeets. Characterization of the tumor microenvironment in a large series of ER/PR negative breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P5-04-23.