Patient outcomes have been linked to the presence of tumor infiltrating lymphocytes (TIL) in solid tumors. In human breast cancer (BC), higher TIL infiltration is associated with a better prognosis and also predicts relevant responses to pre-operative chemotherapy. TIL are primarily composed of T cells, albeit around 20% of BC patients show significant B cell infiltration, and can organize in tertiary lymphoid structures (TLS) located in the peritumoral stroma 1, which are associated with survival in HER2+ and triple negative BC patients. Further, these studies revealed that CD4+ follicular helper T (Tfh) cells producing CXCL13 were specifically associated with peritumoral TLS. CXCL13 is an important B cell chemoattractant whose function is to recruit B cells to the germinal center (GC) in secondary lymphoid organs and TLS, where they can mature and differentiate into memory or antibody-producing B cells. Our recent efforts have focused on exploring the role of Tfh, B cells and CXCL13 play in the development and/or maintenance of GC-like structures in BC-associated TLS. We first derived a GC-B cell gene signature for integration in our published Tfh cell gene signature 1. The combined gene signature was tested for its ability to sensitively detect BC-associated TLS using a qRT-PCR-based assay and a retrospective series (n=54) of formalin-fixed paraffin-embedded (FFPE) BC tissues. These data revealed a correlation between gene signature expression and the extent of TIL and TLS scored by trained pathologists on dual-immunohistochemistry stained (CD3+CD20 for T and B cells, respectively) FFPE tissue sections. The detection of TLS using our combined GC-B cell/Tfh cell gene signature was subsequently confirmed using tissues from our prospective BC cohort (n=83). In addition, CXCL13 gene expression was well correlated with genes associated with GC-B cells and Tfh, indicating these parameters are closely related, as confirmed by immunofluorescence staining on FFPE tissues.
Further understanding the factors that promote TLS formation in vivo could provide important insight for treatment decisions in BC. CXCL13 expression was originally identified as an important signal associated with TLS that was predictive for patient outcomes 1. Factors capable of inducing CXCL13 expression in CD4+ T cells isolated from peripheral blood were investigated using flow cytometry. TGFβ1 alone or together with several cytokines (IL4, IL12, IL21, IL23 and in particular IL2 blockade) increased CXCL13 expression in activated CD4+ T cells. Similar to our characterization of Tfh TIL in fresh tumor tissues, these CXCL13-producing CD4+ T cells were CXCR5 negative and expressed the Tfh marker ICOS; however, they only had low levels of PD-1 expression compared to PD-1hi Tfh cells. CD8+ T cells were also found to produce CXCL13 albeit at low levels. The currently ongoing identification of critical genes involved in regulating CXCL13 production in treated CD4+T cells will help to elucidate the mechanism(s) underlying chemokine induction. The increased accuracy in TIL and TLS detection in BC together with a better understanding of the role Tfh and CXCL13 play in these structures (and GC) development should help to identify the critical immune components involved in BC TLS formation.
1Gu-Trantien et al. J Clin Invest. 2013
Citation Format: Edoardo Migliori, Chunyan Gu-Trantien, Soizic Garaud, Gert Van den Eynden, Alexandre De Wind, Pushpamali De Silva, Cinzia Solinas, Anais Boisson, Celine Naveaux, Denis Larsimont, Martine Piccart-Gebhart, Karen Willard-Gallo. Investigating the role of follicular helper T cells, B cells and CXCL13 in breast cancer-associated tertiary lymphoid structures. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A62.