Comprehensive serum proteomics profiles and potential protein biomarkers for the early detection of advanced adenoma and colorectal cancer.

BACKGROUND: The majority of colorectal cancer (CRC) cases develop from precursor advanced adenoma (AA). With the development of proteomics technologies, blood protein biomarkers have potential applications in the early screening of AA and CRC in the general population. AIM: To identify serum protein biomarkers for the early screening of AA and CRC. METHODS: We collected 43 serum samples from 8 normal controls (NCs), 19 AA patients and 16 CRC patients at China-Japan Friendship Hospital. Quantitative proteomic analysis was performed using liquid chromatography-mass spectrometry/mass spectrometry and data independent acquisition, and differentially expressed proteins (DEPs) with P-values < 0.05 and absolute fold changes > 1.5 were screened out, followed by bioinformatics analysis. Prognosis was further analyzed based on public databases, and proteins expression in tissues were validated by immunohistochemistry. RESULTS: A total of 2132 proteins and 17365 peptides were identified in the serum samples. There were 459 upregulated proteins and 118 downregulated proteins in the NC vs AA group, 289 and 180 in the NC vs CRC group, and 52 and 248 in the AA vs CRC group, respectively. Bioinformatic analysis revealed that these DEPs had different functions and participated in extensive signaling pathways. We also identified DIAPH1, VASP, RAB11B, LBP, SAR1A, TUBGCP5, and DOK3 as important proteins for the progression of AA and CRC. Furthermore, VASP (P < 0.01), LBP (P = 0.01), TUBGCP5 (P < 0.01), and DOK3 (P < 0.01) were associated with a poor prognosis. In addition, we propose that LBP and VASP may be more promising protein biomarkers for the early screening of colorectal tumors. CONCLUSION: Our study elucidated the serum proteomic profiles of AA and CRC patients, and the identified proteins, such as LBP and VASP, may contribute to the early detection of AA and CRC.

Association Between Vitamin B12 Intake and Mortality in Patients with Colorectal Cancer: The US National Health and Nutrition Examination Survey, 1999-2018.

Vitamin B12 plays a role in DNA methylation, influencing the 1-carbon cycle; However, its effect on colorectal cancer (CRC) mortality remains uncertain. This study assessed the relationship between vitamin B12 intake and all-cause and cancer-specific mortality among CRC patients. We analyzed data from the NHANES from 1999 to 2018, using multivariable Cox regression, competing risk model, Kaplan-Meier survival curves, and stratified analysis with interaction effects. The studied involved 4,554 cancer patients (mean age 65.8 years, 47.6% males). Results from multivariate Cox regression indicated that each additional 1 mcg/day of dietary vitamin B12 independently increased the risk of all-cause (HR, 1.07; 95% CI: 1.04-1.09, p < 0.001) and cancer-specific mortality (HR, 1.04; 95% CI, 1.02-1.06; p < 0.001). Kaplan-Meier curves indicated a higher risk of all-cause mortality with increased vitamin B12 intake (Log rank p = 0.01). Subgroup analysis suggested that higher vitamin B12 intake correlated with increased all-cause mortality risk, especially in individuals with higher protein (HR, 1.04; 95% CI, 1.02-1.06; p = 0.019) or carbohydrate intake (HR, 1.03; 95% CI, 1.01-1.05; p = 0.04). Thus, higher vitamin B12 intake correlates with increased all-cause and cancer-specific mortality in CRC patients, particularly those with higher protein or carbohydrate intake.

Clinicopathologic and endoscopic features of sessile serrated lesions and conventional adenomas: a large inpatient population-based study in China.

Objectives: Sessile serrated lesions (SSLs) are precursors of sporadic colorectal cancer (CRC) and have distinct characteristics compared with conventional adenomas (CAs). Several lifestyle and environmental factors may play critical roles in the development of advanced lesions. Our aim is to describe the features of SSLs and CAs and further explore risk factors for advanced lesions. Methods: This is an observational study that collected demographic, endoscopic, and histological data from the China-Japan Friendship Hospital among the inpatient population with pathologically reported as SSL or CA between 2015 and 2022. We analyzed the clinicopathology and endoscopic differences between SSL alone, CA alone, and synchronous SSL+CA groups, and identified risk factors using multiple regression analysis. Results: A total of 9236 polyps from 6598 patients were included in the cohort. Patients with SSL+CA were more likely to be older (p=0.008), while individuals with SSL alone had a higher proportion of early-onset polyps (p<0.001), and SSLs were more common in advanced polyps than CAs (p<0.001). A greater proportion of advanced polyps in the SSL and CA groups were diagnosed as Yamada III, Yamada IV, and laterally spreading tumor (p=0.002, p<0.001, respectively), and multiple SSLs and CAs were more represented in nonadvanced polyps than in advanced polyps. In multiple regression analysis, older patients were more likely to develop advanced SSLs (aOR 1.05, 95% CI 1.02-1.09, p=0.005). Conclusion: SSLs and CAs have diverse demographic, endoscopic, and histological characteristics, and their advanced lesions share different risk factors, which advances the understanding of the etiology and progression of SSLs.

Leukocyte immunoglobulin-like receptor B2 overexpression as a promising therapeutic target and noninvasive screening biomarker for colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) has become the second most deadly malignancy in the world, and the exploration of screening markers and precise therapeutic targets is urgent. Our previous research identified leukocyte immunoglobulin-like receptor B2 (LILRB2) protein as a characteristic protein of CRC, but the association between LILRB2 expression and clinicopathological features, the internal mechanism related to CRC progression, and screening diagnostic efficacy are not clear. Therefore, we hypothesized that LILRB2 is significantly highly expressed in CRC tissues, correlated with advanced stage and a poor prognosis, and could be used as a therapeutic target and potential screening biomarker for CRC. AIM: To explore whether LILRB2 can be used as a potential therapeutic target and noninvasive screening biomarker for CRC. METHODS: Patients who underwent radical surgery for CRC at China-Japan Friendship Hospital between February 2021 and October 2022 were included. Cancer and paracancerous tissues were collected to verify LILRB2 expression, and the association between LILRB2 expression and clinicopathological features was analysed. Serum was collected from CRC patients, adenoma patients and healthy controls during the same period to assess the diagnostic value of LILRB2 as a noninvasive screening biomarker, and its diagnostic value was further compared with that of the traditional markers carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9). RESULTS: A total of 58 CRC patients were included, and LILRB2 protein was significantly overexpressed in cancer tissues compared with paracancerous tissues (P < 0.001). Angiopoietin-like protein 2 (ANGPTL2) protein, as the ligand of LILRB2, was synergistically overexpressed in CRC tissues (P < 0.001), and overexpression of LILRB2 and ANGPTL2 protein was significantly correlated with poor to moderate differentiation, vascular involvement, lymph node metastasis, distant metastasis, advanced tumor-node-metastasis stage and a poor prognosis (P < 0.05), which suggested that LILRB2 and ANGPTL2 are closely associated with CRC progression. In addition, serum LILRB2 concentrations increased stepwise in healthy individuals, adenoma patients and CRC patients with statistically significant differences. The sensitivity of serum LILRB2 for the diagnosis of CRC was 89.74%, the specificity was 88.89%, the area under the curve was 0.95, and the diagnostic efficacy was better than that of conventional CEA and CA19-9. CONCLUSION: LILRB2 protein can be used as a potential novel therapeutic target and noninvasive screening biomarker for CRC, which is beneficial for early screening and precise treatment.

Proteomics-based identification of proteins in tumor-derived exosomes as candidate biomarkers for colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the second leading cause of cancer-related death, with high morbidity worldwide. There is an urgent need to find reliable diagnostic biomarkers of CRC and explore the underlying molecular mechanisms. Exosomes are involved in intercellular communication and participate in multiple pathological processes, serving as an important part of the tumor microenvironment. AIM: To investigate the proteomic characteristics of CRC tumor-derived exosomes and to identify candidate exosomal protein markers for CRC. METHODS: In this study, 10 patients over 50 years old who were diagnosed with moderately differentiated adenocarcinoma were recruited. We paired CRC tissues and adjacent normal intestinal tissues (> 5 cm) to form the experimental and control groups. Purified exosomes were extracted separately from each tissue sample. Data-independent acquisition mass spectrometry was implemented in 8 matched samples of exosomes to explore the proteomic expression profiles, and differentially expressed proteins (DEPs) were screened by bioinformatics analysis. Promising exosomal proteins were verified using parallel reaction monitoring (PRM) analysis in 10 matched exosome samples. RESULTS: A total of 1393 proteins were identified in the CRC tissue group, 1304 proteins were identified in the adjacent tissue group, and 283 proteins were significantly differentially expressed between them. Enrichment analysis revealed that DEPs were involved in multiple biological processes related to cytoskeleton construction, cell movement and migration, immune response, tumor growth and telomere metabolism, as well as ECM-receptor interaction, focal adhesion and mTOR signaling pathways. Six differentially expressed exosomal proteins (NHP2, OLFM4, TOP1, SAMP, TAGL and TRIM28) were validated by PRM analysis and evaluated by receiver operating characteristic curve (ROC) analysis. The area under the ROC curve was 0.93, 0.96, 0.97, 0.78, 0.75, and 0.88 (P < 0.05) for NHP2, OLFM4, TOP1, SAMP, TAGL, and TRIM28, respectively, indicating their good ability to distinguish CRC tissues from adjacent intestinal tissues. CONCLUSION: In our study, comprehensive proteomic profiles were obtained for CRC tissue exosomes. Six exosomal proteins, NHP2, OLFM4, TOP1, SAMP, TAGL and TRIM28, may be promising diagnostic markers and effective therapeutic targets for CRC, but further experimental investigation is needed.

Ubiquitinomics revealed disease- and stage-specific patterns relevant for the 3PM approach in human sigmoid colon cancers.

Objective: The patients with sigmoid colorectal cancer commonly show high mortality and poor prognosis. Increasing evidence has demonstrated that the ubiquitinated proteins and ubiquitination-mediated molecular pathways influence the growth and aggressiveness of colorectal cancer. It emphasizes the scientific merits of quantitative ubiquitinomics in human sigmoid colon cancer. We hypothesize that the ubiquitinome and ubiquitination-mediated pathway networks significantly differ in sigmoid colon cancers compared to controls, which offers the promise for in-depth insight into molecular mechanisms, discovery of effective therapeutic targets, and construction of reliable biomarkers in the framework of predictive, preventive, and personalized medicine (PPPM; 3P medicine). Methods: The first ubiquitinome analysis was performed with anti-K-ε-GG antibody beads (PTMScan ubiquitin remnant motif [K-ε-GG])-based label-free quantitative proteomics and bioinformatics to identify and quantify ubiquitination profiling between sigmoid colon cancer tissues and para-carcinoma tissues. A total of 100 human sigmoid colon cancer samples that included complete clinical information and the corresponding gene expression data were obtained from The Cancer Genome Atlas (TCGA). Ubiquitination was the main way of protein degradation; the relationships between differentially ubiquitinated proteins (DUPs) and their differently expressed genes (DEGs) and between DUPs and their differentially expressed proteins (DEPs) were analyzed between cancer tissues and control tissues. The overall survival of those DUPs was obtained with Kaplan-Meier method. Results: A total of 1249 ubiquitinated sites within 608 DUPs were identified in human sigmoid colon cancer tissues. KEGG pathway network analysis of these DUPs revealed 35 statistically significant signaling pathways, such as salmonella infection, glycolysis/gluconeogenesis, and ferroptosis. Gene Ontology (GO) analysis of 608 DUPs revealed that protein ubiquitination was involved in 98 biological processes, 64 cellular components, 51 molecule functions, and 26 immune system processes. Protein-protein interaction (PPI) network of 608 DUPs revealed multiple high-combined scores and co-expressed DUPs. The relationship analysis between DUPs and their DEGs found 4 types of relationship models, including DUP-up (increased ubiquitination level) and DEG-up (increased gene expression), DUP-up and DEG-down (decreased gene expression), DUP-down (decreased ubiquitination level) and DEG-up, and DUP-down and DEG-down. The relationship analysis between DUPs and their DEPs found 4 types of relationship models, including DUP-up and DEP-up (increased protein expression), DUP-up and DEP-down (decreased protein expression), DUP-down and DEP-up, and DUP-down and DEP-down. Survival analysis found 46 overall survival-related DUPs in sigmoid colon cancer, and the drug sensitivity of overall survival-related DUPs were identified. Conclusion: The study provided the first differentially ubiquitinated proteomic profiling, ubiquitination-involved signaling pathway network changes, and the relationship models between protein ubiquitination and its gene expression and between protein ubiquitination and its protein expression, in human sigmoid colon cancer. It offers the promise for deep insights into molecular mechanisms of sigmoid colon cancer, and discovery of effective therapeutic targets and biomarkers for patient stratification, predictive diagnosis, prognostic assessment, and personalized treatment in the context of 3P medicine. Supplementary Information: The online version contains supplementary material available at 10.1007/s13167-023-00328-2.

microRNA-627-5p inhibits colorectal cancer cell proliferation, migration and invasion by targeting Wnt2.

BACKGROUND: microRNA-627-5p (miR-627-5p) dysregulation has been observed in several cancer types, such as hepatocellular carcinoma, oral squamous cell carcinoma, glioblastoma multiforme, and gastric cancer. The biological function of miR-627-5p in colorectal cancer (CRC) growth and metastasis is yet unclear. AIM: To investigate the effects of miR-627-5p on the malignant biological properties of colorectal malignant tumour cells by targeting Wnt2. METHODS: The levels of miR-627-5p in colorectal tumour tissues were assessed in Gene Expression Omnibus datasets. In order to identify Wnt2 transcript expression in CRC tissues, quantitative real-time polymerase chain reaction (qRT-PCR) analysis was used. Luciferase reporter tests were used to explore whether miR-627-5p might potentially target Wnt2. Wnt2 transcript and protein levels were detected in CRC cells with high miR-627-5p expression. To learn more about how miR-627-5p affects CRC development, migration, apoptosis, and invasion, functional experiments were conducted. Cotransfection with the overexpression vector of Wnt2 and miR-627-5p mimics was utilized to verify whether overexpression of Wnt2 could cancel the impact of miR-627-5p in CRC. Western blot and qRT-PCR were conducted to investigate the effects of miR-627-5p on the Wnt/ß-catenin signalling pathway. RESULTS: miR-627-5p was notably decreased in colorectal tumour tissues, while the gene level of Wnt2 was notably upregulated. A dual luciferase reporter assay revealed that miR-627-5p specifically targets the 3'-untranslated regions of Wnt2 and miR-627-5p upregulation markedly reduced the protein and gene expression of Wnt2 in CRC cells. In vitro gain-of-function assays displayed that miR-627-5p overexpression decreased CRC cells' capabilities to invade, move, and remain viable while increasing apoptosis. Wnt2 overexpression could reverse the suppressive functions of miR-627-5p. Moreover, upregulation of miR-627-5p suppressed the transcript and protein levels of the downstream target factors in the canonical Wnt/ß-catenin signalling, such as c-myc, CD44, ß-catenin, and cyclinD1. CONCLUSION: miR-627-5p acts as a critical inhibitory factor in CRC, possibly by directly targeting Wnt2 and negatively modulating the Wnt/ß-catenin signalling, revealing that miR-627-5p could be a possible treatment target for CRC.

Age-stratified proteomic characteristics and identification of promising precise clinical treatment targets of colorectal cancer.

Colorectal cancer (CRC) is an extremely lethal disease worldwide. However, the underlying pathogenesis remains unclear. This study aimed to reveal the distinct characteristics of age-stratified CRC at the protein level and explore precise treatment targets. Patients who underwent surgical removal with pathologically confirmed CRC at China-Japan Friendship Hospital from January 2020 to October 2021 were recruited, cancer and para-carcinoma tissues (> 5 cm) were detected by mass spectrometry. Ninety-six clinical samples were collected and divided into three groups according to age: young (≤ 50 years), middle-aged (51-69 years), and old (≥ 70 years). Quantitative proteomic analysis was performed, as well as comprehensive bioinformatic analysis based on the Human Protein Atlas, Clinical Proteomic Tumor Analysis Consortium and Connectivity Map databases. The numbers of upregulated and downregulated proteins were 1315 and 560 in the young group, 757 and 311 in the old group, and 1052 and 468 in the middle-aged group, respectively. Bioinformatic analysis showed that these differentially expressed proteins had different molecular functions and participated in extensive signaling pathways. We also revealed ADH1B, ARRDC1, GATM, GTF2H4, MGME1, and LILRB2 as possible cancer-promoting molecules, which might serve as potential prognostic biomarkers and precise therapeutic targets for CRC. SIGNIFICANCE: This study comprehensively characterized the proteomic profiles of age-stratified colorectal cancer patients, focusing on the differentially expressed proteins between cancer and paracancerous tissues in different age groups, in an effort to find corresponding potential prognostic biomarkers and therapeutic targets. In addition, this study provides potentially valuable clinical small molecule inhibitory agents.

Atypical Whipple's disease with special endoscopic manifestations: A case report.

BACKGROUND: Whipple's disease is a rare systemic infection caused by Tropheryma whipplei. Most patients present with nonspecific symptoms, and routine laboratory and imaging examination results also lack specificity. The diagnosis often relies on invasive manipulation, pathological examination, and molecular techniques. These difficulties in diagnosing Whipple's disease often result in misdiagnosis and inappropriate treatments. CASE SUMMARY: This paper reports on the case of a 58-year-old male patient who complained of fatigue and decreased exercise capacity. The results of routine blood tests indicated hypochromic microcytic anemia. Results of gastroscopy and capsule endoscopy showed multiple polypoid bulges distributed in the duodenal and proximal jejunum. A diagnosis of small intestinal adenomatosis was initially considered; hence, the Whipple procedure, a pylorus-preserving pancreaticoduodenectomy, was performed. Pathological manifestations showed many periodic acid-Schiff-positive macrophages aggregated in the intestinal mucosa of the duodenum, upper jejunum, and surrounding lymph nodes. Based on comprehensive analysis of symptoms, laboratory findings, and pathological manifestations, the patient was finally diagnosed with Whipple's disease. After receiving 1 mo of antibiotic treatment, the fatigue and anemia were significantly improved. CONCLUSION: This case presented with atypical gastrointestinal manifestations and small intestinal polypoid bulges, which provided new insight on the diagnosis of Whipple's disease.

Comprehensive proteomic signature and identification of CDKN2A as a promising prognostic biomarker and therapeutic target of colorectal cancer.

BACKGROUND: The carcinogenesis of colorectal cancer (CRC) involves many different molecules and multiple pathways, and the specific mechanism has not been elucidated until now. Existing studies on the proteomic signature profiles of CRC are relatively limited. Therefore, we herein aimed to provide a more comprehensive proteomic signature profile and discover new prognostic markers and therapeutic targets by performing proteomic analysis of CRC and paired normal tissues. AIM: To investigate the proteomic signature and identify novel protein prognostic biomarkers of CRC. METHODS: Cancer tissues and paired normal tissues were collected from 48 patients who underwent surgical removal at the China-Japan Friendship Hospital from January 2020 to June 2021. Data independent acquisition (DIA) quantitative proteomic analysis was performed using high-performance liquid chromatography-mass spectrometry/mass spectrometry (nano-UHPLC-MS/MS) to identify differentially expressed proteins, among which those with a P adj value (t test, BH correction) < 0.05 and an absolute fold change (|log2FC|) > 2 were identified as potential markers. Differentially expressed proteins were selected by bioinformatics analysis and validated by immunohistochemical tissue microarrays, and their association with prognosis was further analyzed with the Gene Expression Profiling Interactive Analysis database to identify prognostic protein biomarkers of CRC. RESULTS: Significantly differential protein expression was observed between cancer tissues and normal tissues. Compared with normal tissues, 1115 proteins were upregulated and 705 proteins were downregulated in CRC based on P adj < 0.05 and |log2FC| > 2, and bioinformatics analysis revealed that the differentially expressed proteins were involved in multiple biological processes associated with tumorigenesis, including ribosome biogenesis in eukaryotes, focal adhesion, extracellular matrix-receptor interactions and other tumor metabolism processes. Moreover, cyclin-dependent kinase inhibitor 2A (CDKN2A) expression was markedly upregulated in CRC, as validated by immunohistochemistry (0.228 vs 0.364, P = 0.0044), and was significantly enriched in tumor proliferation and signal transduction pathways such as the cell cycle and p53 signaling pathways. High CDKN2A expression was significantly correlated with poor prognosis (P = 0.021). These results demonstrated that CDKN2A functions as a driver of CRC. CONCLUSION: Our study provides a comprehensive proteomic signature of CRC and highlights CDKN2A as a potential powerful prognostic marker and precision therapeutic target.

Circulating miR-627-5p and miR-199a-5p are promising diagnostic biomarkers of colorectal neoplasia.

BACKGROUND: Early detection of colorectal neoplasms, including colorectal cancers (CRCs) and advanced colorectal adenomas (AAs), is crucial to improve patient survival. Circulating microRNAs (miRNAs) in peripheral blood are emerging as noninvasive diagnostic markers for multiple cancers, but their potential for screening colorectal neoplasms remains ambiguous. AIM: To identify candidate circulating cell-free miRNAs as diagnostic biomarkers in patients with colorectal neoplasms. METHODS: The study was divided into three phases: (1) Candidate miRNAs were selected from three public miRNA datasets using differential gene expression analysis methods; (2) an independent set of serum samples from 60 CRC patients, 60 AA patients and 30 healthy controls (HCs) was included and analyzed by quantitative real-time polymerase chain reaction for miRNAs, and their diagnostic power was detected by receiver operating characteristic (ROC) analysis; and (3) the origin and function of miRNAs in cancer patients were investigated in cancer cell lines and tumor tissues. RESULTS: Based on bioinformatics analysis, miR-627-5p and miR-199a-5p were differentially expressed in both the serum and tissues of patients with colorectal neoplasms and HCs and were selected for further study. Further validation in an independent cohort revealed that both circulating miR-627-5p and miR-199a-5p were sequentially increased from HCs and AAs to CRCs. The diagnostic power of miR-672-5p yielded an area under the curve (AUC) value of 0.90, and miR-199a-5p had an AUC of 0.83 in discriminating colorectal neoplasms from HCs. A logistic integrated model combining miR-199a-5p and miR-627-5p exhibited a higher diagnostic performance than either miRNA. Additionally, the levels of serum miR-627-5p and miR-199a-5p in CRC patients were significantly lower after surgery than before surgery and the expression of both miRNAs was increased with culture time in the culture media of several CRC cell lines, suggesting that the upregulated serum expression of both miRNAs in CRC might be tumor derived. Furthermore, in vitro experiments revealed that miR-627-5p and miR-199a-5p acted as tumor suppressors in CRC cells. CONCLUSION: Serum levels of miR-199a-5p and miR-627-5p were markedly increased in patients with colorectal neoplasms and showed strong potential as minimally invasive biomarkers for the early screening of colorectal neoplasms.

Identification of circ_0000375 and circ_0011536 as novel diagnostic biomarkers of colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) imposes a tremendous burden on human health, with high morbidity and mortality. Circular ribonucleic acids (circRNAs), a new type of noncoding RNA, are considered to participate in cancer pathogenesis as microRNA (miRNA) sponges. However, the dysregulation and biological functions of circRNAs in CRC remain to be explored. AIM: To identify potential circRNA biomarkers of CRC and explore their functions in CRC carcinogenesis. METHODS: CircRNAs and miRNAs differentially expressed in CRC tissues were identified by analyzing expression profiles from the Gene Expression Omnibus (GEO) database. Circ_0000375 and circ_0011536 were selected as CRC biomarker candidates. Quantitative real-time polymerase chain reaction was utilized to evaluate the expression of these 2 circRNAs in CRC tissues, serums and cell lines. Receiver operating characteristic curves were generated to assess the diagnostic performances of these 2 circRNAs. Then, functional experiments, including cell counting kit-8, wound healing and Transwell invasion assays, were performed after the overexpression of circ_0000375 and circ_0011536 in CRC cell lines. Furthermore, candidate target miRNAs of circ_0000375 and circ_0011536 were predicted via bioinformatics analysis. The expression levels of these miRNAs were explored in CRC cell lines and tissues from GEO datasets. A luciferase reporter assay was developed to examine the interactions between circRNAs and miRNAs. Based on the target miRNAs and downstream genes, functional enrichment analyses were applied to reveal the critical signaling pathways involved in CRC carcinogenesis. RESULTS: Downregulated circ_0000375 and circ_0011536 expression was observed in CRC tissues in GSE126095, clinical CRC tissue and serum samples and CRC cell lines. The areas under the curve for circ_0000375 and circ_0011536 were 0.911 and 0.885 in CRC tissue and 0.976 and 0.982 in CRC serum, respectively. Moreover, the serum levels of these 2 circRNAs were higher in patients at 30 d postsurgery than in patients before surgery, suggesting that the serum expression of circ_0000375 and circ_0011536 is related to CRC tumorigenesis. Circ_0000375 and circ_0011536 overexpression inhibited the proliferation, migration and invasion of CRC cells. Furthermore, miR-1182 and miR-1246, which were overexpressed in CRC tissues in GSE41655, GSE49246 and GSE115513, were verified as target miRNAs of circ_0000375 and circ_0011536, respectively, by luciferase reporter assays. The downstream genes of miR-1182 and miR-1246 were enriched in some CRC-associated pathways, such as the Wnt signaling pathway. CONCLUSION: Circ_0000375 and circ_0011536 may function as tumor suppressors in CRC progression, serving as novel biomarkers for CRC diagnosis and as promising candidates for therapeutic exploration.

Identification of the circRNA-miRNA-mRNA regulatory network and its prognostic effect in colorectal cancer.

BACKGROUND: The high morbidity and mortality of colorectal cancer (CRC) have posed great threats to human health. Circular RNA (CircRNA) and microRNA (miRNA), acting as competing endogenous RNAs (ceRNAs), have been found to play vital roles in carcinogenesis. However, the biological function of ceRNAs in CRC pathogenesis and prognosis remains largely unexplored. AIM: To identify the CRC-specific circRNA-miRNA-mRNA regulatory network and uncover the subnetwork associated with its prognosis. METHODS: CircRNAs, miRNAs and mRNAs differentially expressed (DE) in CRC tissues were selected by expression file analysis in the Gene Expression Omnibus (GEO) database, and the downstream target molecules of circRNAs and miRNAs were predicted. Then, the intersection of differentially expressed RNA molecules with the predicted targets was determined to obtain a ceRNA network. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted to elucidate the possible mechanism of pathogenesis. A survival analysis using the gene profiles and clinical information in The Cancer Genome Atlas (TCGA) database was performed to identify the mRNAs associated with the clinical outcome of CRC patients and construct a prognostic subnetwork. RESULTS: We downloaded three datasets (GSE126095, GSE41655 and GSE41657) of large-scale CRC samples from the GEO database. There were 55 DEcircRNAs, 114 DEmiRNAs and 267 DEmRNAs in CRC tissues compared with normal tissues. After intersecting these molecules with predicted targets, 19 circRNAs, 13 miRNAs and 28 mRNAs were chosen to develop a circRNA-miRNA-mRNA network. GO and KEGG functional enrichment analyses indicated that the retinol metabolic process, leukocyte chemotaxis, extracellular matrix remodeling, endoplasmic reticulum stress, alcohol dehydrogenase activity, gastric acid secretion, nitrogen metabolism and NOD-like receptor signaling pathway might participate in the tumorigenesis of CRC. After verifying the identified mRNA effect in the TCGA database, we finally recognized 3 mRNAs (CA2, ITLN1 and LRRC19) that were significantly associated with the overall survival of CRC patients and constructed a ceRNA subnetwork including 5 circRNAs (hsa_circ_0080210, hsa_circ_0007158, hsa_circ_0000375, hsa_circ_0018909 and hsa_circ_0011536) and 3 miRNAs (hsa-miR-601, hsa-miR-671-5p and hsa-miR-765), which could contain innovative and noninvasive indicators for the early screening and prognostic prediction of CRC. CONCLUSION: We proposed a circRNA-miRNA-mRNA regulatory network closely associated with the progression and clinical outcome of CRC that might include promising biomarkers for carcinogenesis and therapeutic targets.

Alteration of fecal tryptophan metabolism correlates with shifted microbiota and may be involved in pathogenesis of colorectal cancer.

BACKGROUND: Gut tryptophan (Trp) metabolites are produced by microbiota and/or host metabolism. Some of them have been proven to promote or inhibit colorectal cancer (CRC) in vitro and animal models. We hypothesized that there is an alteration of gut Trp metabolism mediated by microbiota and that it might be involved in the pathogenesis of cancer in patients with CRC. AIM: To investigate the features of Trp metabolism in CRC and the correlation between fecal Trp metabolites and gut microbiota. METHODS: Seventy-nine patients with colorectal neoplastic lesions (33 with colon adenoma and 46 with sporadic CRC) and 38 healthy controls (HCs) meeting the inclusion and exclusion criteria were included in the study. Their demographic and clinical features were collected. Fecal Trp, kynurenine (KYN), and indoles (metabolites of Trp metabolized by gut microbiota) were examined by ultraperformance liquid chromatography coupled to tandem mass spectrometry. Gut barrier marker and indoleamine 2,3-dioxygenase 1 (IDO1) mRNA were analyzed by quantitative real-time polymerase chain reaction. Zonula occludens-1 (ZO-1) protein expression was analyzed by immunohistochemistry. The gut microbiota was detected by 16S ribosomal RNA gene sequencing. Correlations between fecal metabolites and other parameters were examined in all patients. RESULTS: The absolute concentration of KYN [1.51 (0.70, 3.46) nmol/g vs 0.81 (0.64, 1.57) nmol/g, P = 0.036] and the ratio of KYN to Trp [7.39 (4.12, 11.72) × 10-3 vs 5.23 (1.86, 7.99) × 10-3, P = 0.032] were increased in the feces of patients with CRC compared to HCs, while the indoles to Trp ratio was decreased [1.34 (0.70, 2.63) vs 2.46 (1.25, 4.10), P = 0.029]. The relative ZO-1 mRNA levels in patients with CRC (0.27 ± 0.24) were significantly lower than those in HCs (1.00 ± 0.31) (P < 0.001), and the relative IDO1 mRNA levels in patients with CRC [1.65 (0.47-2.46)] were increased (P = 0.035). IDO1 mRNA levels were positively associated with the KYN/Trp ratio (r = 0.327, P = 0.003). ZO-1 mRNA and protein levels were positively correlated with the indoles/Trp ratio (P = 0.035 and P = 0.009, respectively). In addition, the genera Asaccharobacter (Actinobacteria) and Parabacteroides (Bacteroidetes), and members of the phylum Firmicutes (Clostridium XlVb, Fusicatenibacter, Anaerofilum, and Anaerostipes) decreased in CRC and exhibited a positive correlation with indoles in all subjects. CONCLUSION: Alteration of fecal Trp metabolism mediated by microbiota is associated with intestinal barrier function and tissue Trp metabolism, and may be involved in the pathogenesis of CRC.