AbstractTherapy using anti‐PD‐1 immune checkpoint inhibitors (ICI) has revolutionized the treatment of many cancers including head and neck squamous cell carcinomas (HNSCC), but only a fraction of patients respond. To better understand the molecular mechanisms driving resistance, we performed extensive analysis of plasma and tumor tissues before and after a 4‐week neoadjuvant trial in which HNSCC patients were treated with the anti‐PD‐1 inhibitor, nivolumab. Luminex cytokine analysis of patient plasma demonstrated that HPV^pos nonresponders displayed high levels of the proinflammatory chemokine, interleukin‐8 (IL‐8), which decreased after ICI treatment, but remained higher than responders. miRNAseq analysis of tetraspanin‐enriched small extracellular vesicles (sEV) purified from plasma of HPV^pos nonresponders demonstrated significantly lower levels of seven miRNAs that target IL‐8 including miR‐146a. Levels of the pro‐survival oncoprotein Dsg2, which has been to down‐regulate miR‐146a, are elevated with HPV^pos tumors displaying higher levels than HPV^neg tumors. Dsg2 levels decrease significantly following ICI in responders but not in nonresponders. In cultured HPV^pos cells, restoration of miR‐146a by forced expression or treatment with miR‐146a‐loaded sEV, reduced IL‐8 level, blocked cell cycle progression, and promoted cell death. These findings identify Dsg2, miR‐146a, and IL‐8 as potential biomarkers for ICI response and suggest that the Dsg2/miR‐146a/IL‐8 signaling axis negatively impacts ICI treatment outcomes and could be targeted to improve ICI responsiveness in HPV^pos HNSCC patients.