About 50% of patients with locally advanced head and neck squamous cell carcinoma (HNSCC) experience recurrences after definitive therapy. The presurgical administration of anti-programmed cell death protein 1 (PD-1) immunotherapy results in substantial pathologic tumor responses (pTR) within the tumor microenvironment (TME). However, the mechanisms underlying the dynamics of antitumor T cells upon neoadjuvant PD-1 blockade remain unresolved, and approaches to increase pathologic responses are lacking. In a phase 2 trial (NCT02296684), we observed that 45% of patients treated with two doses of neoadjuvant pembrolizumab experienced marked pTRs (≥50%). Single-cell analysis of 17,158 CD8⁺ T cells from 14 tumor biopsies, including 6 matched pre-post neoadjuvant treatment, revealed that responding tumors had clonally expanded putative tumor-specific exhausted CD8⁺ tumor-infiltrating lymphocytes (TILs) with a tissue-resident memory program, characterized by high cytotoxic potential (CTX⁺) and ZNF683 expression, within the baseline TME. Pathologic responses after 5 weeks of PD-1 blockade were consistent with activation of preexisting CTX⁺ZNF683⁺CD8⁺ TILs, paralleling loss of viable tumor and associated tumor antigens. Response was associated with high numbers of CD103⁺PD-1⁺CD8⁺ T cells infiltrating pretreatment lesions, whereas revival of nonexhausted persisting clones and clonal replacement were modest. By contrast, nonresponder baseline TME exhibited a relative absence of ZNF683⁺CTX⁺ TILs and subsequent accumulation of highly exhausted clones. In HNSCC, revival of preexisting ZNF683⁺CTX⁺ TILs is a major mechanism of response in the immediate postneoadjuvant setting.

	Year	Citations