Role of Tertiary Lymphoid Structures and Immunotherapy in Head & Neck Squamous Cell Carcinoma

Purpose/Objective(s) The development of novel immunotherapies has provided an avenue to treat many malignancies, including Head and Neck Squamous Cell Carcinoma (HNSCC). Although much research has been done on identifying T-cell mediated immune changes after nivolumab treatment in HNSCC, characterization of humoral immune system changes remains to be fully understood. Furthermore, tertiary lymphoid structures (TLSs) with tumor-inhibiting functions such as antibody generation and clonal expansion have been shown to play a role in predicting positive patient outcomes in several cancers. The relationship that TLSs and immunotherapy might have in patients with HNSCC remains to be elucidated. Materials/Methods 11 HNSCC patients with tumors in the non-oropharynx were treated with neoadjuvant nivolumab 240mg on days 1 and 15 with or without tadalafil. Surgery was completed on day 31. Samples of tumors were taken before the commencement of immunotherapy and after the completion of immunotherapy. Samples after the completion of immunotherapy were stained with Hematoxylin & Eosin (H&E) staining, imaged with a light microscope, and uploaded to Leica's Aperio Imaging software. An analysis of H&E stained images was performed by classifying lymphocyte populations using a previously published organization scale of TLSs into no organization (grade 1), some organization (grade 2), more organization (grade 3), and most organization (grade 4). Results H&E staining showed three patients to not have any lymphocyte organization in their tumor samples after immunotherapy (grade 1), four patients having some lymphocyte organization (grade 2), two patients having more organization (grade 3), and one patient having organization of lymphocytes in a way that TLSs develop (grade 4). Conclusion These results warrant further characterization of TLSs in immunotherapy-treated HNSCC patients. Pre-immunotherapy and post-immunotherapy samples will be sent for CODEX multiplexed tissue imaging analysis for TLS markers: CD20 for B-cells, CD4 for helper T-cells, CD8 for cytotoxic T-cells, PNAd for high endothelial venues (HEVs), CD21 for follicular dendritic cells, and CD208 for mature dendritic cells. The development of novel immunotherapies has provided an avenue to treat many malignancies, including Head and Neck Squamous Cell Carcinoma (HNSCC). Although much research has been done on identifying T-cell mediated immune changes after nivolumab treatment in HNSCC, characterization of humoral immune system changes remains to be fully understood. Furthermore, tertiary lymphoid structures (TLSs) with tumor-inhibiting functions such as antibody generation and clonal expansion have been shown to play a role in predicting positive patient outcomes in several cancers. The relationship that TLSs and immunotherapy might have in patients with HNSCC remains to be elucidated. 11 HNSCC patients with tumors in the non-oropharynx were treated with neoadjuvant nivolumab 240mg on days 1 and 15 with or without tadalafil. Surgery was completed on day 31. Samples of tumors were taken before the commencement of immunotherapy and after the completion of immunotherapy. Samples after the completion of immunotherapy were stained with Hematoxylin & Eosin (H&E) staining, imaged with a light microscope, and uploaded to Leica's Aperio Imaging software. An analysis of H&E stained images was performed by classifying lymphocyte populations using a previously published organization scale of TLSs into no organization (grade 1), some organization (grade 2), more organization (grade 3), and most organization (grade 4). H&E staining showed three patients to not have any lymphocyte organization in their tumor samples after immunotherapy (grade 1), four patients having some lymphocyte organization (grade 2), two patients having more organization (grade 3), and one patient having organization of lymphocytes in a way that TLSs develop (grade 4). These results warrant further characterization of TLSs in immunotherapy-treated HNSCC patients. Pre-immunotherapy and post-immunotherapy samples will be sent for CODEX multiplexed tissue imaging analysis for TLS markers: CD20 for B-cells, CD4 for helper T-cells, CD8 for cytotoxic T-cells, PNAd for high endothelial venues (HEVs), CD21 for follicular dendritic cells, and CD208 for mature dendritic cells.