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bioRxiv · 10.64898/2026.05.26.727931

Spatial Neuroimmune Crosstalk Driving Perineural Invasion in Head and Neck Squamous Cell Carcinoma

Chhabra, R.·Kao, A.·Ding, R.·Bajaj, S.·Jackson, S. G.·Li, W. T.·John, D. J.·Wang-Rodriguez, J.·Ongkeko, W. M.
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Abstract

BackgroundPerineural invasion (PNI) is a clinically important feature of aggressive head and neck squamous cell carcinoma (HNSCC) and is associated with recurrence and poor survival. However, the spatial organization and molecular programs that characterize tumor-nerve interactions in HNSCC remain incompletely understood. MethodsSingle-cell-resolution Xenium spatial transcriptomic data from 10 HNSCC patients were analyzed to define nerve-associated regions, cell-type composition, tumor-nerve proximity, and perineural transcriptional programs. Complementary Visium spatial transcriptomic datasets were used to assess broader transcriptomic features of nerve-associated regions, including epithelial-mesenchymal transition and inferred ligand-receptor signaling. Clinical relevance was evaluated using bulk transcriptomic validation cohorts from TCGA-HNSC and GSE65858. ResultsSpatial mapping identified a distinct nerve-proximal microenvironment characterized by depletion of mature dendritic cells and altered immune and stromal composition, consistent with localized disruption of antigen-presenting support at the tumor-nerve interface. HPV- tumors exhibited closer tumor-nerve proximity and a higher exploratory PNI index compared with HPV+ tumors, suggesting subtype-specific differences in nerve-associated tumor behavior. Differential expression analysis of perineural versus distal tumor cells identified a spatially enriched gene program, including NFE2L2, MDM2, and PPARG, that demonstrated a nerve-proximal gradient most evident in HPV- disease. A composite three-gene score was associated with worse overall survival in HPV- patients in TCGA-HNSC and validated in GSE65858, while showing limited prognostic value in HPV+ disease. Visium analyses provided complementary evidence of EMT enrichment and inferred tumor-nerve signaling involving neural guidance and adhesion-associated pathways. ConclusionsThese findings support a model in which PNI in HNSCC reflects a spatially organized, transcriptionally distinct tumor-nerve microenvironment, particularly in HPV- disease. The NFE2L2/MDM2/PPARG signature may provide a candidate biomarker for risk stratification and nominates pathways for future mechanistic and therapeutic investigation.

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Chhabra, R., Kao, A., Ding, R., Bajaj, S., Jackson, S. G., Li, W. T., John, D. J., Wang-Rodriguez, J., Ongkeko, W. M.. 2026-05-29. Spatial Neuroimmune Crosstalk Driving Perineural Invasion in Head and Neck Squamous Cell Carcinoma. https://doi.org/10.64898/2026.05.26.727931

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