Immunostimulatory Cancer-Associated Fibroblast Subpopulations Can Predict Immunotherapy Response in Head and Neck Cancer

Purpose: Cancer-associated fibroblasts (CAF) have been impli-cated as potential mediators of checkpoint immunotherapy response. However, the extensive heterogeneity of these cells has precluded rigorous understanding of their immunoregulatory role in the tumor microenvironment. Experimental Design: We performed high-dimensional single-cell RNA sequencing (scRNA-seq) on four patient tumors pretreat-ment and posttreatment from a neoadjuvant trial of patients with advanced-stage head and neck squamous cell carcinoma that were treated with the aPD-1 therapy, nivolumab. The head and neck CAF (HNCAF) protein activity profiles, derived from this cohort of paired scRNA-seq, were used to perform protein activity enrich-ment analysis on the 28-patient parental cohort of clinically anno-tated bulk transcriptomic profiles. Ex vivo coculture assays were used to test functional relevance of HNCAF subtypes. Results: Fourteen distinct cell types were identified with the fibroblast population showing significant changes in abundance following nivolumab treatment. Among the fibroblast subtypes, HNCAF-0/3 emerged as predictive of nivolumab response, while HNCAF-1 was associated with immunosuppression. Functionally, HNCAF-0/3 were found to reduce TGF(3-dependent PD-1 thorn TIM-3 thorn exhaustion of CD8 T cells, increase CD103 thorn NKG2A thorn resident memory phenotypes, and enhance the overall cytolytic profile of T cells. Conclusions: Our findings demonstrate the functional impor-tance of distinct HNCAF subsets in modulating the immuno-regulatory milieu of human HNSCC. In addition, we have identified clinically actionable HNCAF subtypes that can be used as a biomarker of response and resistance in future clinical trials.

Automated summary

This study uses high-dimensional single-cell RNA sequencing to reveal the heterogeneity of cancer-associated fibroblasts in the tumor microenvironment and identifies different subtypes that have an impact on the response and resistance to immunotherapy, which is of great significance for future clinical trials.