Abstract 3993: A different spatiotemporal distribution of TIL subtypes is associated with response to neoadjuvant chemotherapy in locally advanced breast cancer patients
Authors: Koenig Lisa , Mairinger Fabian Dominik , Hoffmann Oliver , Bittner Ann-Kathrin , Schmid Kurt Werner , Kimmig Rainer , Kasimir-Bauer Sabine , Bankfalvi Agnes 
Published: July 2017

Abstract
Background: Tumor infiltrating lymphocytes (TILs) are described as an important immune modulator in the tumor microenvironment in cancer and were shown to be associated with breast cancer (BC) outcome. Besides the analysis of total TILs, the discrimination into TIL subtypes is essential as they exhibit pro- or anti-tumorigenic potential. The spatiotemporal TIL distribution at the invasive tumor front (ITF) and the tumor center (TC) might provide insights into tumor progression. Here we analyzed TIL infiltration in core biopsies of BC patients (pts) prior to neoadjuvant chemotherapy (NACT) and correlated these findings with clinical parameters and tumor cell spread.
Material and Methods: Core biopsies from 87 pre-NACT BC pts were cut, deparaffinized and antigen retrieval and automated immunohistochemistry were performed using the following primary antibodies: CD3 (clone: SP7; 1:400, DCS Innovative Diagnostik Systeme, Hamburg, Germany), CD4 (clone: 1F6; 1:40, Zytomed Systems, Berlin, Germany), CD8 (clone: C8/144B; 1:150), CD20 (clone: L26; rtu), CD68 (clone: PG-M1; 1:500, all DAKOCytomation, Glostrup, Denmark). Binding of primary antibody was visualized using the OptiView DAB kit (Ventana Medical Systems, Tucson, USA). The densities of total TILs (H&E staining) and TIL subtypes were evaluated microscopically at the TC and ITF and classified into three categories: Low = 0-10%, Moderate = 11-30%, High ≥ 31% infiltration. TIL results were correlated with clinical parameters and disseminated tumor cells (DTCs) in the bone marrow, determined by immunocytochemistry applying the pan cytokeratin antibody A45-B/B3.
Results: TILs were differentially distributed at both tumor areas. The ITF was mainly infiltrated by CD3+ T- and CD20+ B cells, while low amounts of CD68+ macrophages, CD4+ and CD8+ T cell subtypes were present. Only CD3+ T cells were present in a higher level at the TC. A high total TIL amount and a high CD3+ infiltration at the ITF was associated with poorly differentiated tumors. Pre-NACT tumors &lt; T2 had a high CD4+ T cell infiltration at the TC, whereas in small post-NACT tumors a high infiltration of total TILs and all TIL subtypes at the ITF, except CD68+ cells, was observed. BC pts responding to NACT exhibited significantly more total TILs (p=0.02) and CD3+ T cells (p=0.02) at the TC. A high CD4+ T cell infiltration at the TC was significantly associated with the presence of DTCs post-NACT (p=0.029). CD68+ macrophages located at TC or ITF were not related to any clinical parameter.
Conclusion: The differential association of TIL subtypes with clinical parameters and NACT response underlines the necessity of detailed TIL analysis for gaining insights into immune modulatory processes in the tumor microenvironment. In this regard, BC patients not responding to standard NACT might be identified for application of additional immunotherapy.
Citation Format: Lisa Koenig, Fabian Dominik Mairinger, Oliver Hoffmann, Ann-Kathrin Bittner, Kurt Werner Schmid, Rainer Kimmig, Sabine Kasimir-Bauer, Agnes Bankfalvi. A different spatiotemporal distribution of TIL subtypes is associated with response to neoadjuvant chemotherapy in locally advanced breast cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3993. doi:10.1158/1538-7445.AM2017-3993