Decoding the transcriptional heterogeneity, differentiation lineage, clinical significance in tissue-resident memory CD8 T cell of the small intestine by single-cell analysis.

Resident memory T (Trm) cells which are specifically located in non-lymphoid tissues showed distinct phenotypes and functions compared to circulating memory T cells and were vital for the initiation of robust immune response within tissues. However, the heterogeneity in the transcriptional features, development pathways, and cancer response of Trm cells in the small intestine was not demonstrated. Here, we integrated scRNA-seq and scTCR-seq data pan-tissue T cells to explore the heterogeneity of Trm cells and their development pathways. Trm were enriched in tissue-specific immune response and those in the DUO specially interacted with B cells via TNF and MHC-I signatures. T cell lineage analyses demonstrated that Trm might be derived from the T_CD4/CD8 subset within the same organ or migrated from spleen and mesenteric lymph nodes. We compared the immune repertoire of Trm among organs and implied that clonotypes in both DUO and ILE were less expanded and hydrophilic TRB CDR3s were enriched in the DUO. We further demonstrated that Trm in the intestine infiltrated the colorectal cancer and several effector molecules were highly expressed. Finally, the TCGA dataset of colorectal cancer implied that the infiltration of Trm from the DUO and the ILE was beneficial for overall survival and the response to immune checkpoint blockade.

Aspirin induces immunogenic cell death and enhances cancer immunotherapy in colorectal cancer.

The use of aspirin is associated with reduced incidence of colorectal cancer (CRC). However, the detailed mechanism remains unclear. In this study, we reported that colon cancer cells treated with aspirin showed the hallmarks of immunogenic cell death (ICD), including surface expression of calreticulin (CRT) and heat shock protein 70 (HSP70). Mechanistically, aspirin induced endoplasmic reticulum (ER) stress in colon cancer cells. In addition, aspirin decreased the expression of the glucose transporters, GLUT3, and reduced the key enzyme of glycolysis, including HK2, PFKM, PKM2 and LDHA. The changes of tumor glycolysis after aspirin treatment were associated with c-MYC downregulation. Moreover, aspirin potentiated the antitumor efficacy of anti-PD-1 antibody and anti-CTLA-4 antibody in CT26 tumors. However, this antitumor activity of aspirin in combination with anti-PD-1 antibody was abolished by the depletion of CD8+ T cells. Vaccination with tumor antigens is one of the strategies for activating T-cell response against tumors. Here, we demonstrated that aspirin-treated tumor cells in combination with tumor antigens (AH1 peptide) or protective substituted peptide (A5 peptide) could be served as a potent vaccine to eradicate tumors. Overall, our data indicated that aspirin can be used as an inducer of ICD for CRC therapy.

Taming metabolic competition via glycolysis inhibition for safe and potent tumor immunotherapy.

Metabolic competition between tumors and T cells is fierce in the tumor microenvironment (TME). Tumors usually exhaust glucose and accumulate lactic acid in TME. Nutrient deprivation and lactic acid accumulation in TME blunt T cell functions and antitumor immune responses. Here, we reported that glycolysis-related genes were upregulated in melanoma patients with weak immune responses and T cell poorly infiltrated tumors of BRCA and COAD patients. Dimethyl fumarate (DMF), a GAPDH inhibitor, which is FDA proved to treat autoimmune diseases was identified to promote oxidative pentose phosphate pathway through glucose-6-phosphate dehydrogenase (G6PD) but to suppress aerobic glycolysis and oxidative phosphorylation in tumor cells. Additionally, DMF normalized metabolic competition between tumors and T cells, thus potentiate antitumor responses of tumor infiltrating CD8+ T lymphocytes (TILs). Moreover, DMF optimized the efficiency of immune checkpoint therapy and interleukin-2 (IL-2) therapy while eliminating severe toxicity induced by IL-2 therapy. This study indicates a novel clinically feasible therapy strategy aiming shared metabolic pathway of tumors and T cells for effective and less toxic tumor immunotherapy.

Molecular Cloning, Expression and Macrophage Activation of an Immunoregulatory Protein from Cordyceps militaris.

Protein components of C. militaris have been reported to possess various biological activities. In our previous research, a Cordyceps militaris-derived immunoregulatory protein (CMIP) was naturally isolated and showed the activity of inhibiting the metastasis of breast cancer cells. This study aimed to obtain recombinant CMIP (rCMIP) using recombinant expression and elucidate its ability to activate macrophages. Recombinant CMIP showed one band at approximately 15 kDa or 30 kDa, or two bands at 15 kDa and 30 kDa, under different denaturation conditions of electrophoresis. The cell binding assay showed that rCMIP selectively binds to the surface of macrophages. After adhesion, it did not induce the apoptosis of RAW 264.7 cells, but promoted their proliferation. Moreover, rCMIP significantly induced the expression of M1 macrophage polarization-related molecules. The mean fluorescence intensity (MFI) of CD 86 was enhanced by 2.1-fold and 3.2-fold under 0.64 µM and 1.6 µM of rCMIP treatment, respectively. Cytokines typically expressed in M1 macrophages, such as TNF-α, iNOS, IL-6, CCL 4, CCL 5 and CXCL 10, were also considerably induced by rCMIP, while the expression of cytokines in typical M2 macrophages, like Arg-1, CCL17 and CCL22, were not changed or slightly decreased. Under rCMIP treatment, the release of NO was also appreciably induced. In the present study, we reported cloning, expression and functional characterization of rCMIP, which was naturally isolated from the fruiting body of C. militaris in our previous study. The data imply that rCMIP possesses immunomodulatory activity in macrophages.