Stanford University Stanford, California, United States
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Background/Purpose: Immune checkpoint inhibitors (ICIs) have transformed cancer therapy, but their use is often limited by immune-related adverse events (irAEs), particularly in barrier tissues such as the skin. The mechanisms underlying cutaneous irAEs remain incompletely understood. We aimed to identify cellular and antigenic drivers of skin irAEs in ICI-treated patients Methods: We performed single-cell RNA sequencing (scRNA-seq) and T cell receptor (TCR) sequencing on paired lesional skin biopsies and peripheral blood mononuclear cells (PBMCs) from ICI-treated cancer patients who developed cutaneous irAEs. In parallel, we conducted ex vivo stimulation of patient PBMCs with microbial antigens, followed by flow cytometry to assess T cell activation and cytokine production Results: Inflamed skin lesions harbored a dominant population of clonally expanded CD8⁺ T cells expressing cytotoxic effectors including GZMB, PRF1, and IFNG, indicative of tissue-damaging potential. Notably, CD8⁺ T cells from irAE lesions exhibited substantial expansion of the GZMB⁺ subset accompanied by a reduction in the less cytotoxic GZMK⁺ subset, highlighting a skew towards a more pathogenic cytotoxic phenotype. TCR clonotype tracking revealed overlap between skin-infiltrating clones and circulating CD8⁺ T cells, suggesting antigen-driven recruitment. Computational analysis using GILPH identified shared sequence motifs between patient TCRs and known anti-pathogen TCRs. Additionally, stimulation of patient PBMCs with Staphylococcus aureus and S. epidermidis antigens led to robust activation of GZMB⁺ CD8⁺ T cells and enhanced IFN-γ production Conclusion: Our study identifies microbial antigens as candidate drivers of cytotoxic and inflammatory T cell responses in cutaneous irAEs. These findings highlight the contribution of microbial-reactive CD8⁺ T cells to irAE pathogenesis and suggest that targeting GZMB⁺ CD8⁺ effector functions may offer a therapeutic strategy to alleviate irAEs without compromising anti-tumor immunity
