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Background Head and neck squamous cell carcinoma (HNSCC) are generally thought to be poorly responsive to immunotherapy, yet drivers of immune therapy resistance are not well understood. Single cell RNA sequencing (SCRNAseq) approaches allow for interrogation of the tumor and immune microenvironment, which may serve to elucidate resistance mechanisms. In this study, we present a meta-analysis of existing data to explore whether head and neck cancers harbor significant immune populations and whether immune related differences exist between different sites and subsites within HNSCC. We then use these reference samples as controls to compare to induction chemo/immunotherapy resistant HNSCCs.Methods Studies with single-cell data in HNSCC were included. Graph-based unsupervised clustering and UMAPs were used to reveal cell clusters, which were then annotated based on known cell-type specific marker genes. Wilcoxon signed-rank tests were used to detect differences between samples.Induction chemo/immunotherapy resistant HNSCCs were collected through the Mount Sinai Institutional Biorepository. Whole tumor was dissociated using GentleMACS and submitted for 5’ v3 GEX scRNAseq with TCRseq.Results A total of 383,843 cells were included in the analysis representing >70 unique samples. Compared to HPV+ oropharynx tumors, oral cavity HNSCC had significantly higher proportions of NK cells (p = 0.02), and CD4 T cells (p = 0.008), CD8 T cells (p = 0.001). We then compared oral cavity subsites to evaluate for differences in immune infiltration. After combining TIL samples with the CD45+ fraction of bulk tumor samples, there were no significant differences among the oral cavity subsites of NK cells (p = 0.73), CD4 T cells (p = 0.11), CD8 T cells (p = 0.56), or B cells (p = 0.98) between subsites. Finally we compared reference samples to several tumors resistant to chemo/immunotherapy induction. We used the relevant comparison reference sample (e.g., lateral tongue reference to lateral tongue resistant tumor). These exploratory analyses revealed differences in important immune subsets including CD4 and myeloid cells between untreated reference samples and resistant samples.Conclusions Our results leverage existing datasets to create the largest HNSCC SCRNAseq cohort to date. Preliminary findings in immunotherapy resistant tumors compared to treatment-naïve HNSCC revealed differences in relevant immune subsets and pathways. Further analyses at the protein and spatial level are needed to investigate drivers of immunotherapy resistance among HNSCC.Ethics Approval Mount Sinai PPHS IRB approved STUDY-21-01683