Immunotherapy can elicit robust anticancer responses in the clinic. However, a large proportion of patients with colorectal cancer do not benefit from treatment. Although previous studies have shown that hydrogen sulfide (H2S) is involved in colorectal cancer development and immune escape, further insights into the mechanisms and related molecules are needed to identify approaches to reverse the tumor-supportive functions of H2S. Here, we observed significantly increased H2S levels in colorectal cancer tissues. Decreasing H2S levels by using CBS+/− mice or feeding mice a sulfur amino acid-restricted diet (SARD) led to a marked decrease in differentiated CD4+CD25+Foxp3+ Tregs and an increase in the CD8+ T-cell/Treg ratio. Endogenous or exogenous H2S depletion enhanced the efficacy of anti–PD-L1 and anti–CTLA4 treatment. H2S promoted Treg activation through the persulfidation of ENO1 at cysteine 119. Furthermore, H2S inhibited the migration of CD8+ T cells by increasing the expression of AAK-1 via ELK4 persulfidation at cysteine 25. Overall, reducing H2S levels engenders a favorable immune microenvironment in colorectal cancer by decreasing the persulfidation of ENO1 in Tregs and ELK4 in CD8+ T cells. SARD represents a potential dietary approach to promote responses to immunotherapies in colorectal cancer. Significance: H2S depletion increases the CD8+ T-cell/Treg ratio and enhances the efficacy of anti–PD-L1 and anti–CTLA4 treatment in colon cancer, identifying H2S as an anticancer immunotherapy target.

Cancer-associated fibroblasts (CAFs) play important roles in tumor progression. However, the behaviors of activated CAFs in gastric cancer remain to be determined. The aim of the present study was to investigate the correlations between activated gastric CAFs and the prognosis of patients with gastric cancer, and to determine the effects of activated CAFs on the malignant phenotype and 5-fluorouracil resistance in this cancer. Ninety-five patients with primary gastric cancer were enrolled in this study. Activation states of gastric CAFs were evaluated by immunohistochemistry. A modified method for the primary culture of gastric CAFs was employed. Types of CAFs and activation states were identified by immunocytochemical and immunofluorescent staining. Cell co-culture and gastric CAF conditioned medium transfer models were established to investigate the paracrine effects of activated CAFs on the migration and invasion of gastric cell lines. The half maximal inhibitory concentration of 5-fluorouracil and levels of cell apoptosis were examined using cell viability assay and flow cytometry, respectively. Protein expression levels of associated molecules were measured by Western blotting. Kaplan–Meier survival curves showed that activated gastric CAFs identified via fibroblast activation protein were significantly related to poorer cumulative survival in gastric cancer patients. Five strains of CAFs were successfully cultured via the modified culture method, and three gastric CAFs strains were identified as activated gastric CAFs. The migration and invasion abilities of gastric cells were significantly enhanced in both the co-culture group and the conditioned medium group. The half maximal inhibitory concentration for 5-fluorouracil in BGC-823 cells was elevated after treatment with conditioned medium, and early apoptosis was inhibited. Additionally, an obvious elevation of epithelial–mesenchymal transition level was observed in the conditioned medium group. Activated gastric CAFs correlate with a poor prognosis of cancer patients and may contribute to the malignant phenotype and the development of resistance to 5-fluorouracil via paracrine action in gastric cancer. Gastric CAFs with a specific activation state might be used as a tumor biomarker within the microenvironment for prognosis and as a new therapeutic target for chemoresistant gastric cancer.

The aim of the present study was to clarify the correlations between SPARC expression in gastric cancer-associated fibroblasts (GCAFs) and the prognosis of patients with gastric cancer and to elucidate the role of GCAF-derived SPARC in stemness transformation and 5-fluorouracil resistance in gastric cancer. One hundred ninety-two patients were enrolled in the present study. SPARC expression levels were evaluated by immunohistochemical staining. Primary GCAFs were obtained and cultured from cancer patients for in vitro study, and a lentivirus infection method was employed to knock down SPARC expression in GCAFs. The stemness phenotype and 5-fluorouracil (5-FU) response of gastric cancer cells were assessed via a 3D co-culture model. The apoptotic status and stemness alterations were monitored by flow cytometry and western blotting. Additionally, label-free quantification proteomics was used to identify the differentially expressed proteins and potential pathways in gastric cancer cells treated with GCAF-derived SPARC. Low expression of GCAF-derived SPARC was associated with decreased differentiation and reduced 5-year overall survival and was an independent predictive factor for prognosis in gastric cancer. The 3D tumour growth and 5-FU resistance abilities of gastric cancer cells were elevated after treatment with GCAFs with SPARC knockdown relative to these abilities in negative control cells. Additionally, suppressing SPARC expression in GCAFs facilitated the phenotypic alteration of gastric cancer cells towards CD44+/CD24− cancer stem cell (CSC)-like cells. Quantification proteomics analysis revealed that the differentially expressed proteins in gastric cancer cells were mainly involved in the AKT/mTOR and MEK/ERK signalling pathways. SPARC expression in GCAFs is a useful prognostic factor in patients with gastric cancer. Low expression of GCAF-derived SPARC can lead to CSC transformation and 5-FU resistance. Additionally, the AKT/mTOR and MEK/ERK signalling pathways may participate in the malignant process.

Lipopolysaccharide (LPS) causes lesions of the epithelial barrier, which allows translocation of pathogens from the intestinal lumen to the host's circulation. Hydrogen sulfide (H₂S) regulates multiple physiological and pathological processes in colonic epithelial tissue, and CBS-H₂S axis involved in multiple gastrointestinal disorder. However, the mechanism underlying the effect of the CBS-H₂S axis on the intestinal and systemic inflammation in colitis remains to be illustrated. To investigate the effect of CBS-H₂S axis on the intestinal and systematic inflammation related injuries in LPS induced colitis and the underlying mechanisms. Wild type and CBS^-/+ mice were used to evaluate the effect of endogenous and exogenous H₂S on LPS-induced colitis in vivo. Cytokine quantitative antibody array, western blot and real-time PCR were applied to detect the key cytokines in the mechanism of action. Biotin switch of S-sulfhydration, CRISPR/Cas9 mediated knockout, immunofluorescence and ActD chase assay were used in the in vitro experiment to further clarify the molecular mechanisms. H₂S significantly alleviated the symptoms of LPS-induced colitis in vivo and attenuated the increase of COX-2 expression. The sulfhydrated HuR increased when CBS express normally or GYY4137 was administered. While after knocking kown CBS, the expression of COX-2 in mice colon increased significantly, and the sulfhydration level of HuR decreased. The results in vitro illustrated that HuR can increase the stability of COX-2 mRNA, and the decrease of COX-2 were due to increased sulfhydration of HuR rather than the reduction of total HuR levels. These results indicated that CBS-H₂S axis played an important role in protecting intestinal barrier function in colitis. CBS-H₂S axis increases the sulfhydration level of HuR, by which reduces the binding of HuR with COX-2 mRNA and inhibited the expression of COX-2.

Background: Cisplatin enhances the antitumor T cell response, and the combination of PD-L1 blockade produces a synergistic therapeutic effect. However, the clinical correlation between cisplatin and immunotherapy in colon cancer (CC) is unknown.Methods: Using the “pRRophetic” package, we calculated the IC50 of cisplatin. The correlation between cisplatin IC50, cisplatin resistance–related genes (CCL18 and BCL2A1), and immunotherapy were preliminarily verified in TCGA and further validated in independent cohorts (GSE39582 and GSE17538), cisplatin-resistant CC cell line DLD1, and our own clinical specimens. Classification performance was evaluated using the AUC value of the ROC curve. Scores of immune signatures, autophagy, ferroptosis, and stemness were quantified using the ssGSEA algorithm.Results: Based on respective medians of three CC cohorts, patients were divided into high- and low-IC50 groups. Compared with the high IC50 group, the low-IC50 group had significantly higher tumor microenvironment (TME) scores and lower tumor purity. Most co-signaling molecules were upregulated in low IC50 group. CC patients with good immunotherapy efficacy (MSI, dMMR, and more TMB) were more attributable to the low-IC50 group. Among seven shared differentially expressed cisplatin resistance–related genes, CCL18 and BCL2A1 had the best predictive efficacy of the above immunotherapy biomarkers. For wet experimental verification, compared with cisplatin-resistant DLD1, similar to PD-L1, CCL18 and BCL2A1 were significantly upregulated in wild-type DLD1. In our own CC tissues, the mRNA expression of CCL18, BCL2A1, and PD-L1 in dMMR were significantly increased. The high group of CCL18 or BCL2A1 had a higher proportion of MSI, dMMR, and more TMB. IC50, CCL18, BCL2A1, and PD-L1 were closely related to scores of immune-related pathways, immune signatures, autophagy, ferroptosis, and stemness. The microRNA shared by BCL2A1 and PD-L1, hsa-miR-137, were significantly associated with CCL18, BCL2A1, and PD-L1, and downregulated in low-IC50 group. The activity of the TOLL-like receptor signaling pathway affected the sensitivity of CC patients to cisplatin and immunotherapy. For subtype analysis, immune C2, immune C6, HM-indel, HM-SNV, C18, and C20 were equally sensitive to cisplatin chemotherapy and immunotherapy.Conclusions: CC patients sensitive to cisplatin chemotherapy were also sensitive to immunotherapy. CCL18 and BCL2A1 were novel biomarkers for cisplatin and immunotherapy.