Metformin Reprograms Tryptophan Metabolism to Stimulate CD8+ T-cell Function in Colorectal Cancer.

Colorectal carcinogenesis coincides with immune cell dysfunction. Metformin has been reported to play a role in stimulating antitumor immunity, suggesting it could be used to overcome immunosuppression in colorectal cancer. Herein, using single-cell RNA sequencing (scRNA-seq), we showed that metformin remodels the immune landscape of colorectal cancer. In particular, metformin treatment expanded the proportion of CD8+ T cells and potentiated their function. Analysis of the metabolic activities of cells in the colorectal cancer tumor microenvironment (TME) at a single-cell resolution demonstrated that metformin reprogrammed tryptophan metabolism, which was reduced in colorectal cancer cells and increased in CD8+ T cells. Untreated colorectal cancer cells outcompeted CD8+ T cells for tryptophan, leading to impaired CD8+ T-cell function. Metformin in turn reduced tryptophan uptake by colorectal cancer cells, thereby restoring tryptophan availability for CD8+ T cells and increasing their cytotoxicity. Metformin inhibited tryptophan uptake in colorectal cancer cells by downregulating MYC, which led to a reduction in the tryptophan transporter SLC7A5. This work highlights metformin as an essential regulator of T-cell antitumor immunity by reprogramming tryptophan metabolism, suggesting it could be a potential immunotherapeutic strategy for treating colorectal cancer. SIGNIFICANCE: Analysis of the impact of metformin on the colorectal cancer immunometabolic landscape at a single-cell resolution shows that metformin alters cancer cell tryptophan metabolism to stimulate CD8+ T-cell antitumor activity.

Urea cycle activation triggered by host-microbiota maladaptation driving colorectal tumorigenesis.

Maladaptation of host-microbiota metabolic interplay plays a critical role in colorectal cancer initiation. Here, through a combination of single-cell transcriptomics, microbiome profiling, metabonomics, and clinical analysis on colorectal adenoma and carcinoma tissues, we demonstrate that host's urea cycle metabolism is significantly activated during colorectal tumorigenesis, accompanied by the absence of beneficial bacteria with ureolytic capacity, such as Bifidobacterium, and the overabundance of pathogenic bacteria lacking ureolytic function. Urea could enter into macrophages, inhibit the binding efficiency of p-STAT1 to SAT1 promotor region, and further skew macrophages toward a pro-tumoral phenotype characterized by the accumulation of polyamines. Treating a murine model using urea cycle inhibitors or Bifidobacterium-based supplements could mitigate urea-mediated tumorigenesis. Collectively, this study highlights the utility of urea cycle inhibitors or therapeutically manipulating microbial composition using probiotics to prevent colorectal cancer.

Metformin abrogates Fusobacterium nucleatum-induced chemoresistance in colorectal cancer by inhibiting miR-361-5p/sonic hedgehog signaling-regulated stemness.

BACKGROUND: Chemotherapy resistance is the major cause of recurrence in patients with colorectal cancer (CRC). A previous study found that Fusobacterium (F.) nucleatum promoted CRC chemoresistance. Additionally, metformin rescued F. nucleatum-induced tumorigenicity of CRC. Here, we aimed to investigate whether metformin could revert F. nucleatum-induced chemoresistance and explore the mechanism. METHODS: The role of metformin in F. nucleatum-infected CRC cells was confirmed using cell counting kit 8 assays and CRC xenograft mice. Stemness was identified by tumorsphere formation. Bioinformatic analyses were used to explore the regulatory molecules involved in metformin and F. nucleatum-mediated regulation of the sonic hedgehog pathway. RESULTS: We found that metformin abrogated F. nucleatum-promoted CRC resistance to chemotherapy. Furthermore, metformin attenuated F. nucleatum-stimulated stemness by inhibiting sonic hedgehog signaling. Mechanistically, metformin diminished sonic hedgehog signaling proteins by targeting the MYC/miR-361-5p cascade to reverse F. nucleatum-induced stemness, thereby rescuing F. nucleatum-triggered chemoresistance in CRC. CONCLUSIONS: Metformin acts on F. nucleatum-infected CRC via the MYC/miR-361-5p/sonic hedgehog pathway cascade, subsequently reversing stemness and abolishing F. nucleatum-triggered chemoresistance. Our results identified metformin intervention as a potential clinical treatment for patients with chemoresistant CRC with high amounts of F. nucleatum.

Oral pathogen in the pathogenesis of colorectal cancer.

The human body contains more than 100 trillion microorganisms, including the oral cavity, the skin, and the gastrointestinal tract. After the gastrointestinal tract, the oral cavity harbors one of the most diverse microbial communities within the human body and harbors more than 770 species of bacteria. The composition of the oral and gut microbiomes is quite different, but there may be a microbiological link between the two mucosal sites during the course of disease. More studies indicate that oral bacteria can disseminate to the distal gut via enteral or hematogenous routes. This is mostly obvious in periodontitis, where specific bacteria, such as Fusobacterium nucleatum and Porphyromonas gingivalis, show this pathogenic feature. The translocation of oral microbes to the gut may give rise to a variety of gastrointestinal diseases, including colorectal cancer. However, the precise role that oral microbe play in colorectal cancer has not been fully illustrated. Here, we summarize the current researches on possible pathways of ectopic gut colonization by oral bacteria and their possible contribution to the pathogenesis of colorectal cancer. Understanding the correlation of the oral-to-gut microbial axis in the pathogenesis of colorectal cancer will contribute to precise diagnosis and effective treatment.

F. nucleatum targets lncRNA ENO1-IT1 to promote glycolysis and oncogenesis in colorectal cancer.

OBJECTIVE: Microbiota disorder promotes chronic inflammation and carcinogenesis. High glycolysis is associated with poor prognosis in patients with colorectal cancer (CRC). However, the potential correlation between the gut microbiota and glucose metabolism is unknown in CRC. DESIGN: 18F-FDG (18F-fluorodeoxyglucose) PET (positron emission tomography)/CT image scanning data and microbiota PCR analysis were performed to measure the correlation between metabolic alterations and microbiota disorder in 33 patients with CRC. Multiple colorectal cancer models, metabolic analysis and Seahorse assay were established to assess the role of long non-coding RNA (lncRNA) enolase1-intronic transcript 1 (ENO1-IT1) in Fusobacterium (F.) nucleatum-induced glucose metabolism and colorectal carcinogenesis. RNA immunoprecipitation and chromatin immunoprecipitation sequencing were conducted to identify potential targets of lncRNA ENO1-IT1. RESULTS: We have found F. nucleatum abundance correlated with high glucose metabolism in patients with CRC. Furthermore, F. nucleatum supported carcinogenesis via increasing CRC cell glucose metabolism. Mechanistically, F. nucleatum activated lncRNA ENO1-IT1 transcription via upregulating the binding efficiency of transcription factor SP1 to the promoter region of lncRNA ENO1-IT1. Elevated ENO1-IT behaved as a guider modular for KAT7 histone acetyltransferase, specifying the histone modification pattern on its target genes, including ENO1, and consequently altering CRC biological function. CONCLUSION: F. nucleatum and glucose metabolism are mechanistically, biologically and clinically connected to CRC. Targeting ENO1 pathway may be meaningful in treating patients with CRC with elevated F. nucleatum.

PD-L1 Overexpression on Tumor-Infiltrating Lymphocytes Related to Better Prognosis of Colorectal Cancer.

BACKGROUND: PD-L1 expression on tumor-infiltrating lymphocytes (TILs) has recently been reported as a biomarker for colorectal cancer (CRC). However, the prognostic and clinical significance of PD-L1 on TILs in CRC remains controversial. We performed this meta-analysis to evaluate the association between the PD-L1 expression on TILs and clinicopathological features and prognosis of CRC patients. METHODS: A comprehensive literature search for relevant studies published up to Feb 2020 was performed using Medline, Embase, and Web of Science. Odds ratio (OR) with 95% CI was selected to appraise the correlation between PD-L1 expression on TILs with prognostic and clinicopathological characteristics of CRC patients. Begg's and Egger's test were used to assess publication bias. The statistical analysis was conducted using Stata software. RESULTS: A total of 19 studies including 5,213 CRC cases were included in this meta-analysis. The pooled results showed that PD-L1 overexpression on TILs was relevant to longer OS (OR = 1.36, 95% CI = 1.19 - 1.55, p < 0.01) and longer DFS/RFS (OR = 1.22, 95% CI = 1.03 - 1.44, p = 0.02). Moreover, CRC patients with high expression of PD-L1 on TILS was associated with lower T stage (OR = 2.30, 95% CI = 1.85 - 2.87, p < 0.01), less lymph node in-vasion (OR = 1.48, 95% CI = 1.03 - 2.13, p = 0.03), less distant metastasis (OR = 2.56, 95% CI = 1.81 - 3.64, p < 0.01), earlier TNM stage (OR = 1.93, 95% CI = 1.34 - 2.66, p < 0.01), later tumor grade (OR = 0.38, 95% CI = 0.23 - 0.62, p < 0.01) and high MSI status (OR = 0.36, 95% CI = 0.25 - 0.52, p < 0.01). But it is not related to tumor size, tumor differentiation, MMR status, BRAF mutant, and KRAS mutant. CONCLUSIONS: This meta-analysis revealed that PD-L1 expression on TILs can serve as a significant biomarker for positive prognosis and clinicopathological features of CRC. Our results may provide some useful information when using PD-L1 expression to predict the survival of CRC patients and to select the beneficial CRC patients from PD-1/PD-L1 antibody treatment.

Metformin elicits antitumour effect by modulation of the gut microbiota and rescues Fusobacterium nucleatum-induced colorectal tumourigenesis.

BACKGROUND: The effect of metformin on gut microbiota has been reported, but whether metformin can suppress colorectal cancer (CRC) by affecting gut microbiota composition and rescue F. nucleatum-induced tumourigenicity remains unclear. METHODS: To identify microbiota associated with both CRC occurrence and metformin treatment, first, we reanalyzed the gut microbiome of our previous data on two human cohorts of normal and CRC individuals. Subsequently, we summarized microbiota altered by metformin from published literatures. Several taxa, including Fusobacterium, were associated with both CRC occurrence and metformin treatment. We investigated the effect of metformin on APCMin/+ mice given with or without F. nucleatum. 16S rRNA gene sequencing was performed. FINDINGS: We summarized 131 genera altered by metformin from 18 published literatures. Five genera reported to be changed by metformin, including Bacteroides, Streptococcus, Achromobacter, Alistipes and Fusobacterium, were associated with CRC in both of our human cohorts. Metformin relieved the symptoms caused by F. nucleatum administration in APCMin/+ mice, and showed promise in suppressing intestinal tumour formation and rescuing F. nucleatum-induced tumourigenicity. Administration of F. nucleatum and/or metformin had effect on gut microbiome structure, composition and functions of APCMin/+ mice. INTERPRETATION: This study pioneers in predicting critical CRC-associated taxa contributing to the antitumour effect of metformin, and correlating gut microbiome with the antitumour effect of metformin in experimental animals. We presented a basis for future investigations into metformin's potential effect on suppressing F. nucleatum-induced tumor formation in vivo. FUNDING: This work was supported by grants from the National Natural Science Foundation of China (31701250).

RING-Finger Protein 6 Amplification Activates JAK/STAT3 Pathway by Modifying SHP-1 Ubiquitylation and Associates with Poor Outcome in Colorectal Cancer.

Objective: The E3 ubiquitin ligase RNF6 (RING-finger protein 6) plays a crucial role in carcinogenesis. However, the copy number and expression of RNF6 were rarely reported in colorectal cancer. We aimed to explore the mechanical, biological, and clinical role of RNF6 in colorectal cancer initiation and progression.Design: The copy number and expression of RNF6 were analyzed from Tumorscape and The Cancer Genome Atlas (TCGA) datasets. Gene expressions were examined by real-time PCR, Western blot, and immunohistochemical staining. Gene expression profiling studies were performed to identify pivotal genes regulated by RNF6. Biological function of RNF6 on tumor growth and metastasis was detected in vivo and in vitro Role of RNF6 in modulating SHP-1 expression was examined by coimmunoprecipitation and confocal microscopy, respectively.Results: The copy number of RNF6 was significantly amplified in colorectal cancer, and the amplification was associated with RNF6 expression level. Amplification and overexpression of RNF6 positively correlated with patients with colorectal cancer with poor prognosis. The gene set enrichment analysis (GSEA) revealed cell proliferation, and invasion-related genes were enriched in RNF6 high-expressed colorectal cancer cells as well as in patients from TCGA dataset. Downregulation of RNF6 impaired the colorectal cancer cell proliferation and invasion in vitro and in vivo RNF6 may activate the JAK/STAT3 pathway and increase pSTAT3 levels by inducing the ubiquitination and degradation of SHP-1.Conclusions: Genomic amplification drives RNF6 overexpression in colorectal cancer. RNF6 may be a novel biomarker in colorectal carcinogenesis, and RNF6 may increase pSTAT3 level via promoting SHP-1 ubiquitylation and degradation. Targeting the RNF6/SHP-1/STAT3 axis provides a potential therapeutic option for RNF6-amplified tumors. Clin Cancer Res; 24(6); 1473-85. ©2017 AACR.

Fusobacterium nucleatum Promotes Chemoresistance to Colorectal Cancer by Modulating Autophagy.

Gut microbiota are linked to chronic inflammation and carcinogenesis. Chemotherapy failure is the major cause of recurrence and poor prognosis in colorectal cancer patients. Here, we investigated the contribution of gut microbiota to chemoresistance in patients with colorectal cancer. We found that Fusobacterium (F.) nucleatum was abundant in colorectal cancer tissues in patients with recurrence post chemotherapy, and was associated with patient clinicopathological characterisitcs. Furthermore, our bioinformatic and functional studies demonstrated that F. nucleatum promoted colorectal cancer resistance to chemotherapy. Mechanistically, F. nucleatum targeted TLR4 and MYD88 innate immune signaling and specific microRNAs to activate the autophagy pathway and alter colorectal cancer chemotherapeutic response. Thus, F. nucleatum orchestrates a molecular network of the Toll-like receptor, microRNAs, and autophagy to clinically, biologically, and mechanistically control colorectal cancer chemoresistance. Measuring and targeting F. nucleatum and its associated pathway will yield valuable insight into clinical management and may ameliorate colorectal cancer patient outcomes.

LncRNA GClnc1 Promotes Gastric Carcinogenesis and May Act as a Modular Scaffold of WDR5 and KAT2A Complexes to Specify the Histone Modification Pattern.

UNLABELLED: Long noncoding RNAs (lncRNA) play a role in carcinogenesis. However, the function of lncRNAs in human gastric cancer remains largely unknown. In this study, we identified a novel lncRNA, GClnc1, which was upregulated and associated with tumorigenesis, tumor size, metastasis, and poor prognosis in gastric cancer. GClnc1 affected gastric cancer cell proliferation, invasiveness, and metastasis in multiple gastric cancer models. Mechanistically, GClnc1 bound WDR5 (a key component of histone methyltransferase complex) and KAT2A histone acetyltransferase, acted as a modular scaffold of WDR5 and KAT2A complexes, coordinated their localization, specified the histone modification pattern on the target genes, including SOD2, and consequently altered gastric cancer cell biology. Thus, GClnc1 is mechanistically, functionally, and clinically oncogenic in gastric cancer. Targeting GClnc1 and its pathway may be meaningful for treating patients with gastric cancer. SIGNIFICANCE: This report documents a novel lncRNA, GClnc1, which may act as a scaffold to recruit the WDR5 and KAT2A complex and modify the transcription of target genes. This study reveals that GClnc1 is an oncogenic lncRNA in human gastric cancer. Cancer Discov; 6(7); 784-801. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 681.

Emerging Roles of Hydrogen Sulfide in Inflammatory and Neoplastic Colonic Diseases.

Hydrogen sulfide (H2S) is a toxic gas that has been recognized as an important mediator of many physiological processes, such as neurodegeneration, regulation of inflammation, blood pressure, and metabolism. In the human colon, H2S is produced by both endogenous enzymes and sulfate-reducing bacteria (SRB). H2S is involved in the physiological and pathophysiological conditions of the colon, such as inflammatory bowel disease (IBD) and colorectal cancer (CRC), which makes the pharmacological modulation of H2S production and metabolism a potential chemical target for the treatment of colonic diseases. However, the exact mechanisms and pathways by which H2S-mediates normal physiological function and disease in the colon are not fully understood. Besides, the production and release of H2S are modulated by both endogenous and exogenous factors. This review will discuss the production and storage of H2S, its biological roles and the emerging importance in physiology and pathology of IBD and CRC.

Berberine may rescue Fusobacterium nucleatum-induced colorectal tumorigenesis by modulating the tumor microenvironment.

BACKGROUND: Accumulating evidence links colorectal cancer (CRC) with the intestinal microbiota. However, the disturbance of intestinal microbiota and the role of Fusobacterium nucleatum during the colorectal adenoma-carcinoma sequence have not yet been evaluated. METHODS: 454 FLX pyrosequencing was used to evaluate the disturbance of intestinal microbiota during the adenoma-carcinoma sequence pathway of CRC. Intestinal microbiota and mucosa tumor-immune cytokines were detected in mice after introducing 1,2-dimethylhydrazine (DMH), F. nucleatum or Berberine (BBR), using pyrosequencing and Bio-Plex Pro™ cytokine assays, respectively. Protein expressions were detected by western blotting. RESULTS: The levels of opportunistic pathogens, such as Fusobacterium, Streptococcus and Enterococcus spp. gradually increased during the colorectal adenoma-carcinoma sequence in human fecal and mucosal samples. F. nucleatum treatment significantly altered lumen microbial structures, with increased Tenericutes and Verrucomicrobia (opportunistic pathogens) (P < 0.05 = in wild-type C57BL/6 and mice with DMH treatment). BBR intervention reversed the F. nucleatum-mediated increase in opportunistic pathogens, and the secretion of IL-21/22/31, CD40L and the expression of p-STAT3, p-STAT5 and p-ERK1/2 in mice, compared with mice fed with F. nucleatum alone. CONCLUSIONS: F. nucleatum colonization in the intestine may prompt colorectal tumorigenesis. BBR could rescue F. nucleatum-induced colorectal tumorigenesis by modulating the tumor microenvironment and blocking the activation of tumorigenesis-related pathways.