Spatiotemporal analysis of tumour-infiltrating immune cells in biliary carcinogenesis

Abstract

Intraductal papillary neoplasms (IPN) and biliary epithelial neoplasia (BilIN) are distinct precursor lesions that may progress to biliary tract carcinoma (BTC). A comprehensive characterization of the inflammatory microenvironment of IPN and BilIN precursor lesions was the main aim of our study. Immunohistochemistochemical staining with anti-CD3, -CD4, -CD8, -CD20, -CD68, -CD163, -CD56, and -MUM1 antibodies was used to investigate tumor-infiltrating immune cell populations in tissue samples from patients in whom coexisting precursor lesions and invasive BTC were identified. Using a triplet sample set of nonneoplastic epithelium, precursor lesions, and invasive BTC, we extensively analyzed the spatiotemporal evolution of the immune microenvironment during IPN- and BilIN-related carcinogenesis. We found that stromal CD3+(P=0.002), CD4+(P=0.007) and CD8+(P<0.001) T cells were significantly reduced in IPN compared to nonneoplastic epithelium. Similarly, fewer CD20+B cells (P=0.008), MUM1+plasma cells (P=0.012) and CD163+M2-like macrophages (P=0.008) were observed in IPN compared to nonneoplastic epithelium. Stromal CD68+(P=0.001) and CD163+(P<0.001) macrophages significantly increased during the transition from IPN to invasive BTC. Intraepithelial CD4+and CD8+T cells conversely decreased from nonneoplastic epithelium to IPN and were similarly distributed in invasive BTC. BilIN-associated biliary carcinogenesis was characterized by a significant reduction of intraepithelial CD8+T-lymphocyte infiltration at the transition from non-tumorous biliary epithelium to BilIN(P=0.008) and a further decrease from BilIN to BTC (P=0.004). We conclude that IPN and BilIN undergo distinct immune-cell changes throughout biliary carcinogenesis