Malignant epithelial states drive immune dysfunction in ampulla of Vater carcinoma

Methods

We performed single-cell RNA sequencing on eight primary AoV tumors and four matched normal tissues. Comprehensive clustering and transcriptomic analyses identified cell-type composition, epithelial heterogeneity, and tumor-immune interactions. Findings were validated using deconvolution of bulk RNA-seq data from 62 AoV carcinoma patients.

Results Malignant epithelial cells were categorized into four distinct subtypes: Int-Wnt, PB-KRAS, Int-Hypoxia, and Cycling stage. PB-KRAS cells exhibited stem-like transcriptional programs and high genomic instability. Deconvolution analysis of bulk RNA-seq data from the independent AoV cohort revealed that enrichment of the PB-KRAS subtype correlated with tumor recurrence and poor survival.

Our immune profiling analysis discovered a significant association between PB-KRAS subtype and GZMK+ CD8+ T cells, which are in a pre-dysfunctional state, alongside SPP1⁺ macrophages exhibiting immunosuppressive traits. Spatial transcriptome data further supports the immunosuppressive natures of TME around PB-KRAS subtype malignant epithelial cells in AoV carcinoma. Conclusions Our study presents a single-cell atlas of AoV carcinoma, highlighting the molecular diversity of malignant epithelium and its association with the immune microenvironment.

The PB-KRAS subtype emerges as a stem-like, immunosuppressive tumor state associated with poor prognosis, providing insights for future therapeutic targeting.