Cancer immunotherapy has revolutionized cancer treatment, and it relies heavily on the comprehensive understanding of the immune landscape of the tumor microenvironment (TME). Here, we obtain a detailed immune cell atlas of esophageal squamous cell carcinoma (ESCC) at single-cell resolution. Exhausted T and NK cells, regulatory T cells (Tregs), alternatively activated macrophages and tolerogenic dendritic cells are dominant in the TME. Transcriptional profiling coupled with T cell receptor (TCR) sequencing reveal lineage connections in T cell populations. CD8 T cells show continuous progression from pre-exhausted to exhausted T cells. While exhausted CD4, CD8 T and NK cells are major proliferative cell components in the TME, the crosstalk between macrophages and Tregs contributes to potential immunosuppression in the TME. Our results indicate several immunosuppressive mechanisms that may be simultaneously responsible for the failure of immuno-surveillance. Specific targeting of these immunosuppressive pathways may reactivate anti-tumor immune responses in ESCC.

Immunotherapy targeting checkpoint blockade to rescue T cells from exhaustion has become an essential therapeutic strategy in treating cancers. Till now, little is known about the PD-L1 graphic pattern and characteristics in CD8⁺ T cells. We combined cytometry by time-of-flight (CyTOF) and imaging mass cytometry (IMC) approaches to analyze CD8⁺ T cells from primary lung cancers and discovered that PD-L1⁺CD8⁺ T cells were enriched in tumor lesions, spatially localized with PD-1⁺CD8⁺ T cells. Furthermore, PD-L1⁺CD8⁺ T cells exerted regulatory functions that inhibited CD8⁺ T cells proliferation and cytotoxic abilities through the PD-L1/PD-1 axis. Moreover, tumor-derived IL-27 promotes PD-L1⁺CD8⁺ T cells development through STAT1/STAT3 signaling. Single-cell RNA sequencing data analysis further clarified PD-L1⁺CD8⁺ T cells elevated in the components related to downregulation of adaptive immune response. Collectively, our data demonstrated that PD-L1⁺CD8⁺ T cells enriched in lung cancer engaged in tolerogenic effects and may become a therapeutic target in lung cancer.

Proinflammatory cytokine interleukin 32 (IL-32) is involved in infectious diseases and cancer, but what subtypes of immune cells express IL-32 and its roles in tumor microenvironment (TME) have not been well discussed. In this study, we applied bioinformatics to analyze single-cell RNA sequencing data about tumor-infiltrating immune cells from esophageal squamous cell carcinoma (ESCC) TME and analyzed IL-32 expression in different immune cell types. We found CD4⁺regulatory T cells (Treg cells) express the highest level of IL-32, while proliferating T and natural killer cells expressed relatively lower levels. Knocking down of IL-32 reduced Foxp3 and interferon gamma (IFNγ) expressions in CD4⁺and CD8⁺T cells, respectively. IL-32 was positively correlated with Foxp3, IFNG, and GZMB expression but was negatively correlated with proliferation score. IL-32 may have a contradictory role in the TME such as it promotes IFNγ expression in CD8⁺T cells, which enhances the antitumor activity, but at the same time induces Foxp3 expression in CD4⁺T cells, which suppresses the tumor immune response. Our results demonstrate different roles of IL-32 in Treg cells and CD8⁺T cells and suggest that it can potentially be a target for ESCC cancer immunosuppressive therapy.

Increased expression of the kinesin family member 23 (KIF23) has been verified in gastric cancer (GC) and its upregulation contributes to cell proliferation. Even though, the role of KIF23 has not been fully elucidated in GC, and the mechanisms of KIF23 as an oncogene remain unknown. To further identify its potential role in GC, we analyzed gene expression data from GC patients in GEO and TCGA datasets. KIF23 was upregulated in GC, and increased expression of KIF23 correlated with poor prognosis. Importantly, KIF23 inhibition not only suppressed GC cell proliferation, tumorigenesis, but also migration and invasion, and arrested the cell cycle in the G2/M phase. Mechanistic investigations confirmed that KIF23 activated the Wnt/β-catenin signaling pathway by directly interacting with APC membrane recruitment 1 (Amer1). Furthermore, KIF23 exhibited competitive binding with Amer1 to block the association of Amer1 with adenomatous polyposis coli (APC), thus relocating Amer1 from the membrane and cytoplasm to the nucleus and attenuating the ability of Amer1 to negatively regulate Wnt/β-catenin signaling, resulting in activation of this signaling pathway. Collectively, our findings demonstrated that KIF23 promoted GC cell proliferation by directly interacting with Amer1 and activating the Wnt/β-catenin signaling pathway.

Regulatory IL-10-producing B cells (B10 cells) play a critical role in preventing and curing autoimmune diseases in experimental mouse models. However, the precise cellular and molecular mechanisms of action of B10 cells in humans, especially in patients with Crohn’s disease (CD), remain to be determined. miR-155 regulates many physiological and pathological conditions, including inflammation such as that in CD. In this study, we aimed to explore the effect of miRNA-155 on IL-10 production by B cells in healthy controls (HCs) and CD patients. Interestingly, we found that CD24hiCD27+ B cells express high levels of miRNA-155 and IL-10, which are positively correlated. Additionally, CD24hiCD27+ B cells express higher levels of Toll-like receptor 9 than those found in other B cell subsets. Overexpression of miRNA-155 promotes IL-10 production, while inhibition of miRNA-155 decreases IL-10 production. We determined that miR-155 directly inhibits the expression of Jarid2, which reduces H3K27me3 binding to the IL10 promoter and increases IL-10 gene expression. In co-culture systems, the CD24hiCD27+ B cells from HCs suppressed the secretion of TNFa and IFNg by monocytes and T cells, respectively. However, the number and function of CD24hiCD27+ B cells from CD patients were decreased. Moreover, we found that miR-155 induces CD24hiCD27+ B cells to produce higher levels of TNFa instead of IL-10 in CD patients than in the controls and that the increased number of IL-10+TNFa+ B cells reduces the induction of Foxp3 expression and the inhibition of IFN production by CD4+CD25- T cells, as well as TNFa production by monocytes. Our study demonstrates the critical role of miRNA-155 in the regulation of IL-10 production by B cells and reveals the novel molecular mechanism underlying the functional impairment of B10 cells in CD patients. Our study has the potential to drive the development of B10 cell-based strategies to ameliorate disease progression in CD patients.

Protein arginine methyltransferase 5 (PRMT5) participates in the symmetric dimethylation of arginine residues of proteins and contributes to a wide range of biological processes. However, how PRMT5 affects the transcriptional and epigenetic programs involved in the establishment and maintenance of T cell subset differentiation and roles in antitumor immunity is still incompletely understood. In this study, using single-cell RNA and chromatin immunoprecipitation sequencing, we found that mouse T cell–specific deletion of PRMT5 had greater effects on CD8⁺ than CD4⁺ T cell development, enforcing CD8⁺ T cell differentiation into Klrg1⁺ terminal effector cells. Mechanistically, T cell deficiency of PRMT5 activated Prdm1 by decreasing H4R3me2s and H3R8me2s deposition on its loci, which promoted the differentiation of Klrg1⁺CD8⁺ T cells. Furthermore, effector CD8⁺ T cells that transited to memory precursor cells were decreased in PRMT5-deficient T cells, thus causing dramatic CD8⁺ T cell death. In addition, in a mouse lung cancer cell line–transplanted tumor mouse model, the percentage of CD8⁺ T cells from T cell–specific deletion of PRMT5 mice was dramatically lost, but CD8⁺Foxp3⁺ and CD8⁺PDL1⁺ regulatory T cells were increased compared with the control group, thus accelerating tumor progression. We further verified these results in a mouse colon cancer cell line–transplanted tumor mouse model. Our study validated the importance of targeting PRMT5 in tumor treatment, because PRMT5 deficiency enforced Klrg1⁺ terminal CD8⁺ T cell development and eliminated antitumor activity. Key Points T cell–specific deletion of PRMT5 induced SLEC⁺CD8⁺ T cell differentiation. T cell PRMT5 deficiency decreased H4R3me2s and H3R8me2s deposition on Prdm1 loci. T cell–specific deletion of PRMT5 promoted tumor progression.