Integrated Single-Cell and Spatial Analysis Reveals Context-Dependent Myeloid-T Cell Interactions in Response to Immune Checkpoint Blockade in Head and Neck Cancer.

purposeWe conduct a systematic evaluation of cell-cell interactions between tumor-infiltrating immune cells in patients with head and neck squamous cell carcinoma (HNSCC) who have been treated with immune checkpoint blockade (ICB) using spatial and single-cell omics data. EXPERIMENTAL

designWe employed complementary techniques from both Visium spot-based spatial transcriptomics and CosMx Spatial Molecular Imager single-cell spatial omics, utilizing a 64-plex protein panel and a 1,000-gene RNA panel, which includes 435 ligands and receptors. We conducted integrated bioinformatics analyses to identify cellular neighborhoods of colocalizing cell types and ligand-receptor interactions across different single-cell and spatial data modalities.

resultsWith 522,399 single cells profiled for both RNA and protein from 23 patients, along with spot-resolved spatial RNA sequencing (RNA-seq) data from eight patients treated with ICB, and through bioinformatics analysis of publicly available single-cell and bulk RNA-seq, we identified a spatial and cell type-specific context dependency in the differences in myeloid and T-cell interactions between responder and nonresponder samples. We further defined the cellular neighborhood and sources of chemokine CXCL9/10-CXCR3 interactions, emphasizing the specificity of this marker in responder samples, an emerging target in ICB, as well as other underappreciated markers and targets for ICB response in HNSCC, such as CXCL16-CXCR6 and CCL4/5-CCR5.

conclusionsWe have provided a valuable resource for analyzing spatial and cell-cell ligand-receptor interactions, including the cellular and spatial contexts of ICB response markers. Our data suggest that future mechanistic studies should consider this context specificity when evaluating ICB response biomarkers and targets.