CD266+ myofibroblasts mediate resistance to immunochemotherapy through ECM stiffness in head and neck squamous cell...

The efficacy of immunotherapy in unresectable/recurrent/metastatic head and neck squamous cell carcinoma (U/R/M HNSCC) remains suboptimal compared to primary untreated resectable HNSCC. The immunological and stromal landscape during immunotherapy in U/R/M HNSCC is crucial for understanding drug resistance mechanisms. In this study, single-cell RNA sequencing was performed on 42 samples pre- and post-treatment from patients in our prospective trial (NCT05156970) who received immunochemotherapy. A subset of myofibroblasts expressing CD266 was identified in hampering the response to immunochemotherapy in patients with U/R/M HNSCC. This distinct subtype of cancer-associated fibroblasts (CAFs) enhanced extracellular matrix (ECM) stiffness and cellular tension through a Piezo1-mediated biomechanical mechanism, spatially segregated CD8+ T cell infiltration to establish immune-excluded niches, and ultimately facilitated immune evasion. Mechanistically, PIEZO1 inhibition suppressed YAP activation and reduced FAK phosphorylation, thereby mitigating matrix remodeling effects driven by myCAF CD266. We also observed that Piezo1 inhibition had synergistic effect with PD-1 blockade in HNSCC in vivo. In summary, our findings reveal an immunosuppressive mechanism in which stromal cells regulate ECM stiffness, presenting a potential therapeutic target for U/R/M HNSCC.