Single-cell RNA-seq Reveals Chemoradiotherapy-induced Immune Microenvironment Modulation in Locally Advanced Cervical Cancer

Abstract

Single-cell RNA sequencing (scRNA-seq) could reshape cancer research by allowing comprehensive analysis of cellular heterogeneity and cell-type classification. We sought to investigate the immune microenvironment modulation induced by concurrent chemoradiotherapy (CCRT) in locally advanced cervical cancer at single-cell resolution. Using scRNA-seq and bioinformatics, we analyzed 3,674 and 6,541 CD45+ tumor-infiltrating immune cells from 3 pairs of tumor biopsies before and at the middle of CCRT in locally advanced cervical cancer patients. Immune cells were sorted by flow cytometry and sequenced by 10X genomics at single-cell resolution. Immunostaining and bulk sequencing dataset were used to validate our findings of scRNA-seq. A total of 14 clusters of tumor-infiltrating immune cells were observed in cervical cancer, including 6 subsets of macrophages, 2 subsets of natural killer (NK) cells, CD8+ T cells, regulatory T cells, proliferated T cells, B cells, dendritic cells, plasma cells. We found 2 transitional macrophages subsets between 2 subsets of M1 and 2 subsets of M2 macrophages in tumors. CCRT for cervical cancer induced the transition of macrophages from M2 to transitional-like to M1 macrophages. Interestingly, 2 subsets of M1 macrophages were predominant in tumors during CCRT, but not before CCRT. Additionally, most of NK cells undergoing exhaustion in pre-CCRT samples, while another cluster of NK cells with cytotoxic and activated phenotype were prevalent in tumors during CCRT. Our study reveals the anti-tumor immune response activated by CCRT at single-cell resolution in locally advanced cervical cancer, which may help further understand the mechanism of anti-tumor effect of CCRT and optimize treatment strategies.