Tumour-draining lymph nodes in head and neck cancer : immunological signatures and clinical implications

The lymphatic system is responsible for orchestrating immune responses, particularly in cancer. Oral squamous cell carcinoma (OSCC) represents a significant clinical challenge, with lymphatic metastasis being a crucial determinant of patient outcomes. Tumour-draining lymph nodes (TDLNs) serve as key sites for understanding the complex interplay between cancer and the immune system. This thesis investigates the immunological landscape of TDLNs in OSCC patients, aiming to elucidate their role in tumour progression and immune regulation.

In Paper I, we utilized flow cytometry to characterize T cell populations in TDLNs of OSCC patients. Our findings revealed high expression of PD-1 and other activation markers on tumour-infiltrating lymphocytes (TILs), indicating a potential state of exhaustion. Furthermore, we observed a significant association between PD-1 positivity on T cells and lymph node metastasis, suggesting a link between immune exhaustion and disease progression.

In Paper II, we investigated T regulatory cells (Tregs) in TDLNs and their potential role in immunosuppression. Our results demonstrated a high accumulation of Tregs expressing immune checkpoint molecules in TDLNs of OSCC patients, suggesting their involvement in dampening anti-cancer immunity. The elevated levels of Tregs in TDLNs correlated with disease progression, highlighting their potential as prognostic markers.

Paper III focused on the expression of immune checkpoint molecules on T cells within TDLNs. We observed expression of PD-1, TIGIT, CTLA-4, TIM-3, and LAG-3 on T cells in TDLNs, with significant associations between their expression level and the risk of recurrence. These findings underscore the potential of immune checkpoint molecules as therapeutic targets in OSCC.

In Paper IV, we investigated the role of B cells in TDLNs of OSCC patients. Our analysis revealed a higher proportion of naïve B cells in TDLNs compared to nTDLNs, suggesting a potential role of B cells in immune modulation. Furthermore, we identified elevated levels of B regulatory cells producing IL-10 in TDLNs, implicating their potential participation in immunosuppression and disease progression.

Paper V focused on dendritic cells (DCs) within TDLNs and their contribution to anti-tumour immunity. Our results showed increased levels of cDC1 and pDCs in TDLNs, along with higher expression of co-stimulatory molecules and immune checkpoint molecules on cDC1s. These findings highlight the potential role of DCs in regulating the immune microenvironment within TDLNs.

Finally, Paper VI investigated the presence and phenotypes of neutrophils in TDLNs of OSCC patients. Our analysis revealed notable phenotypic variations of neutrophils across different types of lymph nodes, with TDLNs exhibiting higher levels of activated neutrophils. Additionally, we observed a correlation between activated neutrophils in TDLNs and tumour size, suggesting their potential involvement in disease progression.

Conclusion: In conclusion, this thesis provides insights into the immunological landscape of TDLNs in OSCC. The findings highlight the complex interplay between immune cell subsets and their impact on tumour progression and patient outcomes. By elucidating the immune dynamics within TDLNs, this thesis lays the groundwork for developing novel prognostic markers and therapeutic strategies tailored to individual immune profiles in OSCC patients.

List of scientific papers

I. Piersiala K, Farrajota Neves da Silva P, Hjalmarsson E, Kolev A, Kågedal Å, Starkhammar M, Elliot A, Marklund L, Margolin G, Munck- Wikland E, Kumlien Georén S, Cardell LO. CD4+ and CD8+ T cells in sentinel nodes exhibit distinct pattern of PD-1, CD69, and HLA-DR expression compared to tumor tissue in oral squamous cell carcinoma. Cancer Sci. 2021 Mar;112(3):1048-1059.
https://doi.org/10.1111/cas.14816

II. Piersiala K, da Silva PFN, Lagebro V, Kolev A, Starkhammar M, Elliot A, Marklund L, Munck-Wikland E, Margolin G, Kumlien Georén S, Cardell LO. Tumour-draining lymph nodes in head and neck cancer are characterized by accumulation of CTLA-4 and PD-1 expressing Treg cells. Transl Oncol. 2022 Sep;23:101469.
https://doi.org/10.1016/j.tranon.2022.101469

III. Piersiala K, Hjalmarsson E, Vilma Lagebro, Farrajota Neves da Silva P, Bark R, Elliot A, Marklund L, Margolin G, Kumlien Georén S, Cardell LO. Prognostic Value of T Regulatory Cells and Immune Checkpoints Expression in Tumor-Draining Lymph Nodes for Oral Squamous Cell Carcinoma. [Submitted]

IV. Piersiala K, Hjalmarsson E, da Silva PFN, Lagebro V, Kolev A, Starkhammar M, Elliot A, Marklund L, Munck-Wikland E, Margolin G, Kumlien Georén S, Cardell LO. Regulatory B cells producing IL-10 are increased in human tumor draining lymph nodes. Int J Cancer. 2023 Aug 15;153(4):854-866.
https://doi.org/10.1002/ijc.34555

V. Hjalmarsson E, Piersiala K, Lagebro V, Farrajota Neves da Silva P, Petro M, Starkhammar M, Elliot A, Bark R, Margolin G, Kumlien Georén S, Cardell LO. Enhanced PD-L1 Expression on Conventional Dendritic Cells in Tumor-Draining Lymph Nodes Associates with Metastasis Risk and Poor Prognosis in Oral Squamous Cell Carcinoma. [Submitted]

VI. Ekstedt S, Piersiala K, Farrajota Neves da Silva P, Kolev A, Margolin G, Kumlien Georén S, Cardell LO. Phenotypical differences of neutrophils patrolling tumour-draining lymph nodes in head and neck cancer. [Submitted]