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536 Divergent cancer etiologies drive distinct B cell signatures and tertiary lymphoid structures in head and neck cancer
  1. Ayana Ruffin,
  2. Anthony Cillo,
  3. Tracy Tabib,
  4. Angen Liu,
  5. Sayali Onkar,
  6. Sheryl Kunning,
  7. Caleb Lampenfeld,
  8. Irina Abecassis,
  9. Zengbiao Qi,
  10. Ryan Soose,
  11. Umamaheswar Duvvuri,
  12. Seungwon Kim,
  13. Steffi Oesterrich,
  14. Robert Lafyatis,
  15. Robert Ferris,
  16. Dario Vignali and
  17. Tullia Bruno
  1. University of Pittsburgh, Pittsburgh, PA, USA


Background Current FDA-approved immunotherapies aim to reinvigorate CD8+ T cells, but the contribution of the humoral arm of the immune response in human cancer remains poorly understood. B cells within tissues can mediate anti-tumor immunity and regulate immune responses by presenting antigen and producing tumor-specific antibodies and immunomodulatory cytokines. Head and neck squamous cell carcinoma (HNSCC) can be induced by human papillomavirus (HPV+) and carcinogens such as tobacco and alcohol (HPV-), and the immune infiltrate is quite distinct in the two etiologies, in particular, increased B cells in HPV+ HNSCC patients. Further, increased B cells in HNSCC patients correlate with improved patient survival. Our study seeks to differentiate B cell phenotype, function and location in HPV+ and HPV- HNSCC to identify putative B cell-centric immunotherapeutic targets.

Methods We utilized a multi-level approach to clearly categorize B cells in HNSCC patients. Single cell RNA sequencing (scRNAseq) was performed on CD45+ tumor infiltrating lymphocytes (TIL) from HPV+ and HPV- HNSCC patients. HNSCC TIL and PBL were stained via spectral cytometry (Cytek Aurora,25 parameters) for unbiased analysis of B cell subsets via computational spectral unmixing. Paraffin embedded slides from HNSCC primary tumors were utilized for multispectral immunofluorescence (mIF) to identify tertiary lymphoid structures (TLS) and identify differences in HPV+ and HPV- disease.

Results We demonstrated distinct trajectories for B cells in HPV+ and HPV- disease. HPV- HNSCC tumors mainly contained memory B cells and plasma cells, while the B cells in HPV+ HNSCC were naïve and germinal center (GC). Further, we quantified B cells and CD4+ T cells in TLS, and germinal center-like TLS were associated with improved outcome in HPV+ disease. We also observed that transcriptional and protein expression of Semaphorin A (SEMA4a) was restricted to GC B cells and increased on GC B cells in HNSCC patients compared to healthy tonsils. Additionally, we identified distinct waves of gene expression in GC B cells in HNSCC tumors, ultimately revealing a novel transitional state for GC B cells in the tumor microenvironment (TME).

Conclusions Understanding B cell function in human cancers and how different TMEs influence B cells and TLS are important for devising novel therapeutic options for cancer patients. Ultimately, development of therapeutics to enhance B cell responses in the TME should be prioritized as a compliment to T-cell mediated therapies.

This is an open access article distributed in accordance with the Creative Commons Attribution 4.0 Unported (CC BY 4.0) license, which permits others to copy, redistribute, remix, transform and build upon this work for any purpose, provided the original work is properly cited, a link to the licence is given, and indication of whether changes were made. See:

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