Abstract A053: Activated B cells in human primary tumors present antigen and increase antitumor function of CD4 T-cells in tertiary lymphoid structures
Authors: Bruno Tullia C. , Ruffin Ayana T. , Cillo Anthony R. , Ferris Robert L. , Vignali Dario A.A. 
Published: February 2019

Immunotherapy, specifically anti-PD1, has improved patient survival in a range of tumor types including head and neck squamous cell carcinoma (HNSCC) and non-small cell lung cancer (NSCLC). Despite the success of anti-PD1 therapy, only 20% of patients produce a durable response to this treatment. Thus, a need exists to develop additional therapeutic strategies to treat these patients, which includes evaluation of other tumor-infiltrating immune cells that could further augment the CD8+ and CD4+ T-cell response. Tumor-infiltrating B cells (TIL-B) represent a possible target for immunotherapy due to their predominance in the tumor microenvironment (TME) and crucial role in the immune response. However, TIL-B function in cancer and in the context of immunotherapy has been understudied. In fact, conclusions on an anti- or protumor role for TIL-Bs in the TME is dependent on the study. However, in HNSCC and NSCLC patients, current evidence suggests an antitumor role for TIL-Bs. Specifically, detection of TIL-Bs within tertiary lymphoid structures (TLS) correlates with better prognosis. While TIL-Bs have been identified in HNSCC and NSCLC patients, their complete phenotypic signature and function in the TME has been understudied with no focus on their role as antigen presenting cells (APCs) and their influence on CD8+ and CD4+ tumor infiltrating lymphocytes (TILs). We hypothesize that TIL-Bs help generate potent, long-term immune responses against cancer by presenting tumor antigens to CD4 TILs within TLS.Using unmanipulated, primary human B cells from fresh tumor, we quantified and further characterized TIL-Bs in HNSCC and NSCLC utilizing single-cell RNAseq and multiparameter flow cytometry. We observed increased numbers of activated TIL-Bs in these primary tumors compared to other immune subsets, specifically CD27+ TIL-Bs. We further assessed the TIL-Bs by correlating phenotype of the TIL-B with its location in the TME, predominantly separating out differences between TIL-Bs within TLS and outside TLS. In addition, we generated a specific antigen presentation assay in vitro, and we observed three types of CD4+ TIL responses when TIL-Bs presented autologous tumor antigens. There were activated responder CD4+ TILs that proliferated when combined with TIL-Bs alone, which indicates stimulation with endogenous tumor antigens. There were antigen-associated responders that required exogenous autologous tumor lysate to elicit a CD4+ TIL response, and there were patient CD4 TILs that did not respond to antigen presentation by TIL-Bs. Within the activated and antigen-associated responders, the TIL-B phenotype influenced the CD4+ TIL phenotype; if the TIL-Bs were activated (CD27+), the CD4+ TILs were T helper (antitumor) CD4+ T-cells and if the TIL-Bs were non-activated (CD27-), the CD4+ TILs were T regulatory cells (protumor). These data suggest that TIL-Bs influence the phenotype and function of CD4+ TILs in patient tumors. In conclusion, activated TIL-Bs are increased in human primary tumors, they can present antigen to CD4+ TILs and influence their overall phenotype. Determining the complete activation signature of TIL-Bs in HNSCC and NSCLC patients will determine the extent of their antitumor function in these cancers. Comparison of TIL-Bs in HNSCC and NSCLC is important as there are not many unified studies on TIL-B function across tumor types. Further, because HNSCC has two etiologies (viral vs. carcinogen induced), we are able to better study the differential function of activated and nonactivated TIL-Bs in solid tumors. Ultimately, results from this study will help predict how to target TIL-B functions in future TIL-B-specific immunotherapies or in combination with current immunotherapies for HNSCC and NSCLC patients like blockade of the inhibitory receptor, PD-1.

Citation Format: Tullia C. Bruno, Ayana T. Ruffin, Anthony R. Cillo, Robert L. Ferris, Dario A.A. Vignali. Activated B cells in human primary tumors present antigen and increase antitumor function of CD4 T-cells in tertiary lymphoid structures [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A053.