Single-cell RNA sequencing highlights the immunosuppression of IDO1⁺ macrophages in the malignant transformation of oral leukoplakia

Abstract

Rationale: Immunosuppressive tumor microenvironment (iTME) plays an important role in carcinogenesis, and some macrophage subsets are associated with iTME generation.However, the sub-population characterization of macrophages in oral carcinogenesis remains largely unclear.Here, we investigated the immunosuppressive status with focus on function of a macrophage subset that expressed indoleamine 2,3 dioxygenase 1 (Macro-IDO1) in oral carcinogenesis.Methods: We built a single cell transcriptome atlas from 3 patients simultaneously containing oral squamous cell carcinoma (OSCC), precancerous oral leukoplakia (preca-OLK) and paracancerous tissue (PCA).Through single-cell RNA sequencing and further validation using multicolor immunofluorescence staining and the in vitro/in vivo experiments, the immunosuppressive cell profiles were built and the role of a macrophage subset that expressed indoleamine 2,3 dioxygenase 1 (Macro-IDO1) in the malignant transformation of oral leukoplakia was evaluated. Results:The iTME formed at preca-OLK stage, as evidenced by increased exhausted T cells, Tregs and some special subsets of macrophages and fibroblasts.Macro-IDO1 was predominantly enriched in preca-OLK and OSCC, distributed near exhausted T cells and possessed tumor associated macrophage transformation potentials.Functional analysis revealed the established immunosuppressive role of Macro-IDO1 in preca-OLK and OSCC: enriching the immunosuppression related genes; having an established level of immune checkpoint score; exerting strong immunosuppressive interaction with T cells; positively correlating with the CD8-exhausted.The immunosuppression related gene expression of macrophages also increased in preca-OLK/OSCC compared to PCA.The use of the IDO1 inhibitor reduced 4NQO induced oral carcinogenesis in mice.Mechanistically, IFN--JAK-STAT pathway was associated with IDO1 upregulation in OLK and OSCC. Ivyspring