Colorectal cancer cells induce the formation of cancer-associated fibroblasts by activating the ERK signaling pathway in fibroblasts / 南方医科大学学报

OBJECTIVE@#To investigate the mechanism by which conditioned medium of colorectal cancer cells promotes the formation of cancer-associated fibroblasts (CAFs).@*METHODS@#Normal human colorectal fibroblasts (CCD-18Co cells) in logarithmic growth phase were treated with the conditioned media of colorectal cancer HCT116 cells (HCT116-CM) or Caco-2 cells (Caco-2-CM) alone or in combination with 300 nmol/L ERK inhibitor SCH772984. The expression levels of CAFs-related molecular markers were detected in the treated cells with real-time quantitative PCR (RT- qPCR) and immunofluorescence assay, and the changes in cell proliferation, colony formation and migration were assessed with RTCA, colony formation and wound healing assays; Western blotting was performed to detect the activated signaling pathways in the fibroblasts and the changes in CAFs formation after blocking of the signaling pathway.@*RESULTS@#HCT116-CM and Caco-2-CM significantly upregulated mRNA expression levels of CAFs markers (including α-SMA, FAP, FN and TGF-β) in CCD-18Co cells, and strongly promoted fibroblast transformation into CAFs (P < 0.05). The two conditioned media also promoted the proliferation, colony formation and migration of CCD-18Co cells (P < 0.05) and significantly increased the levels of α-SMA protein and ERK phosphorylation in the cells (P < 0.05). The ERK inhibitor SCH772984 obviously inhibited the expression of α-SMA and the transformation of CCD-18Co cells into CAFs induced by the conditioned medium of colorectal cancer cells (P < 0.05).@*CONCLUSION@#Colorectal cancer cells may induce the formation of colorectal CAFs by activating the ERK pathway in the fibroblasts.

Research on applying genetic and environmental risk score in risk-adapted colorectal cancer screening / 中华流行病学杂志

Objective: To evaluate the effectiveness of a risk-adapted colorectal cancer screening strategy constructed utilizing genetic and environmental risk score (ERS). Methods: A polygenic risk score (PRS) was constructed based on 20 previously published single nucleotide polymorphisms for colorectal cancer in East Asian populations, using 2 160 samples with MassARRAY test results from a multicenter randomized controlled trial of colorectal cancer screening in China. The ERS was calculated using the Asia-Pacific Colorectal Screening Score system. Logistic regression was used to analyze the association between PRS alone and PRS combined with ERS and colorectal neoplasms risk, respectively. We also designed a risk-adapted screening strategy based on PRS and ERS (high-risk participants undergo a single colonoscopy, low-risk participants undergo an annual fecal immunochemical test, and those with positive results undergo further diagnostic colonoscopy) and compared its effectiveness with the all-acceptance colonoscopy strategy. Results: The high PRS group had a 26% increased risk of colorectal neoplasms compared with the low PRS group (OR=1.26, 95%CI: 1.03-1.54, P=0.026). Participants with the highest PRS and ERS were 3.03 times more likely to develop advanced colorectal neoplasms than those with the lowest score (95%CI: 1.87-4.90, P<0.001). As the risk-adapted screening simulation reached the third round, the detection rate of the PRS combined with ERS strategy was not statistically different from the all-acceptance colonoscopy strategy (8.79% vs. 10.46%, P=0.075) and had a higher positive predictive value (14.11% vs. 10.46%, P<0.001) and lower number of colonoscopies per advanced neoplasms detected (7.1 vs. 9.6, P<0.001). Conclusion: The risk-adapted screening strategy combining PRS and ERS helps achieve population risk stratification and better effectiveness than the traditional colonoscopy-based screening strategy.

Molecular mechanism of resveratrol combined with irinotecan in treatment of colorectal cancer / 中国中药杂志

This study aimed to investigate the mechanism of resveratrol(RES) combined with irinotecan(IRI) in the treatment of colorectal cancer(CRC). The targets of RES, IRI, and CRC were obtained from databases, and the targets of RES combined with IRI in the treatment of CRC were acquired by Venn diagram. The protein functional cluster analysis, GO and KEGG enrichment analyses were performed. In addition, the protein-protein interaction(PPI) network was constructed. The core target genes were screened out and the target-signaling pathway network was set up. IGEMDOCK was used to dock the core target gene molecules. Besides, the relationship between the expression level of key target genes and the prognosis and immune infiltration of CRC was analyzed. Based on the in vitro cell experiment, the molecular mechanism of RES combined with IRI in the treatment of CRC was explored and analyzed. According to the results, 63 potential targets of RES combined with IRI were obtained for CRC treatment. Furthermore, cluster analysis revealed that protein functions included 23% transmembrane signal receptors, 22% protein modifying enzymes, and 14% metabolite converting enzymes. GO analysis indicated that BPs were mainly concentrated in protein autophosphorylation, CCs in receptor complex and plasma membrane, and MFs in transmembrane receptor protein tyrosine kinase activity. Moreover, KEGG signaling pathways were mainly enriched in central carbon metabolism in cancer. The key targets of RES combined with IRI in the treatment of CRC were PIK3CA, EGFR, and IGF1R, all of which were significantly positively correlated with the immune infiltration of CRC. As shown by the molecular docking results, PIK3CA had the most stable binding with RES and IRI. Compared with the results in the control group, the proliferation ability and EGFR protein expression of CRC cells in the RES-treated group, the IRI-treated group, and the RES+IRI treated group significantly decreased. Moreover, the cell proliferation ability and EGFR protein expression level of CRC cells in the RES+IRI treated group were remarkably lower than those in the IRI-treated group. In conclusion, PIK3CA, EGFR, and IGF1R are the key targets of RES combined with IRI in CRC treatment. In addition, RES can inhibit the proliferation of CRC cells and improve IRI chemoresistance by downregulating the EGFR signaling pathway.

Decursin affects proliferation, apoptosis, and migration of colorectal cancer cells through PI3K/Akt signaling pathway / 中国中药杂志

We investigated the effects of decursin on the proliferation, apoptosis, and migration of colorectal cancer HT29 and HCT116 cells through the phosphatidylinositol 3-kinase(PI3K)/serine-threonine kinase(Akt) pathway. Decursin(10, 30, 60, and 90 μmol·L~(-1)) was used to treat HT29 and HCT116 cells. The survival, colony formation ability, proliferation, apoptosis, wound hea-ling area, and migration of the HT29 and HCT116 cells exposed to decursin were examined by cell counting kit-8(CCK8), cloning formation experiments, Ki67 immunofluorescence staining, flow cytometry, wound healing assay, and Transwell assay, respectively. Western blot was employed to determine the expression levels of epithelial cadherin(E-cadherin), neural cadherin(N-cadherin), vimentin, B-cell lymphoma/leukemia-2(Bcl-2), Bcl-2-associated X protein(Bax), tumor suppressor protein p53, PI3K, and Akt. Compared with the control group, decursin significantly inhibited the proliferation and colony number and promoted the apoptosis of HT29 and HCT116 cells, and it significantly down-regulated the expression of Bcl-2 and up-regulated the expression of Bax. Decursin inhibited the wound healing and migration of the cells, significantly down-regulated the expression of N-cadherin and vimentin, and up-regulated the expression of E-cadherin. In addition, it significantly down-regulated the expression of PI3K and Akt and up-regulated that of p53. In summary, decursin may regulate epithelial-mesenchymal transition(EMT) via the PI3K/Akt signaling pathway, thereby affecting the proliferation, apoptosis, and migration of colorectal cancer cells.

Neutrophil extracellular traps activates focal adhesion kinase by upregulating MMP9 expression to promote proliferation and migration of mouse colorectal cancer cells / 细胞与分子免疫学杂志

Objective To investigate how the neutrophil extracellular traps (NETs) affect the proliferation and migration of mouse MC38 colorectal cancer cells and its mechanism. Methods Spleen neutrophils were extracted in mouse, followed by collection of NETs after ionomycin stimulation in vitro. The proliferation of MC38 cell was detected by CCK-8 assay, and migration ability were detected by TranswellTM and cell scratch assay, after co-incubation with MC38 cells. The mRNA expression of cellular matrix metalloproteinase 2 (MMP2) and MMP9 were detected by real-time fluorescence quantitative PCR, and the expression of MMP2, MMP9 and focal adhesion kinase (FAK), phosphorylated FAK protein were detected by Western blot. After silencing MMP9 using small interfering RNA (siRNA), the effect of NETs on the proliferation and migration ability of MC38 cells and the altered expression of related molecules were examined by previous approach. Results NETs promoted the proliferation and migration of MC38 cells and up-regulated the MMP9 expression and FAK phosphorylation. Silencing MMP9 inhibited the promotion of MC38 proliferation and migration by NETs and suppressed FAK phosphorylation. Conclusion NETs up-regulates MMP9 expression in MC38 cells, activates FAK signaling pathway and promotes tumor cell proliferation and migration.

The efficacy and side effects of rigosertib combined with chemotherapy in KRAS mutant colorectal cancer mice / 中华肿瘤杂志

Objective: To investigate the effect of rigosertib (RGS) combined with classic chemotherapy drugs including 5-fluorouracil, oxaliplatin, and irinotecan in colorectal cancer. Methods: Explore the synergy effects of RGS and 5-fluorouracil (5-FU), oxaliplatin (OXA), and irinotecan (IRI) on colorectal cancer by subcutaneously transplanted tumor models of mice. The mice were randomly divided into control group, RGS group, 5-FU group, OXA group, IRI group, 5-FU+ RGS group, OXA+ RGS group and IRI+ RGS group. The synergy effects of RGS and OXA on KRAS mutant colorectal cancer cell lines in vitro was detected by CCK-8. Ki-67 immunohistochemistry and TdT-mediated dUTP nick-end labeling (TUNEL) staining were performed on the mouse tumor tissue sections, and the extracted tumor tissue was analyzed by western blot. The blood samples of mice after chemotherapy and RGS treatment were collected, blood routine and liver and kidney function analysis were conducted, and H&E staining on liver sections was performed to observe the side effects of chemotherapy and RGS. Results: The subcutaneously transplanted tumor models were established successfully in all groups. 55 days after administration, the fold change of tumor size of OXA+ RGS group was 37.019±8.634, which is significantly smaller than 77.571±15.387 of RGS group (P=0.029) and 92.500±13.279 of OXA group (P=0.008). Immunohistochemical staining showed that the Ki-67 index of tumor tissue in control group, OXA group, RGS group and OXA+ RGS group were (100.0±16.8)%, (35.6±11.3)%, (54.5±18.1)% and (15.4±3.9)%, respectively. The Ki-67 index of OXA+ RGS group was significantly lower than that in control group (P=0.014), but there was no significant difference compared to OXA group and RGS group (OXA: P=0.549; RGS: P=0.218). TUNEL fluorescence staining showed that the apoptotic level of OXA+ RGS group was 3.878±0.547, which was significantly higher than 1.515±0.442 of OXA group (P=0.005) and 1.966±0.261 of RGS group (P=0.008). Western blot showed that the expressions of apoptosis related proteins such as cleaved-PARP, cleaved-caspase 3 and cleaved-caspase 8 in the tumor tissues of mice in the OXA+ RGS group were higher than those in control group, OXA group and RGS group. After the mice received RGS combined with chemotherapy drugs, there was no significant effect on liver and kidney function indexes, but the combined use of oxaliplatin and RGS significantly reduced the white blood cells [(0.385±0.215)×10(9)/L vs (5.598±0.605)×10(9)/L, P<0.001] and hemoglobin[(56.000±24.000)g/L vs (153.333±2.231)g/L, P=0.001] of the mice. RGS, chemotherapy combined with RGS and chemotherapy alone did not significantly increase the damage to liver cells. Conclusions: The combination of RGS and oxaliplatin has a stronger anti-tumor effect on KRAS mutant colorectal cancer. RGS single agent will not cause significant bone marrow suppression and hepatorenal injury in mice, but its side effects may increase correspondingly after combined with chemotherapy.

Alterations in APC, BECN1, and TP53 gene expression levels in colon cancer cells caused by monosodium glutamate / Alterações nos níveis de expressão gênica APC, BECN1 e TP53 em células de câncer de cólon causadas por glutamato monossódico

Colorectal cancer (CRC) is a disease with high incidence worldwide. As of 2018, it is the second leading cause of cancer deaths in the world. In Saudi Arabia, the incidence of this disease has been increasing in the younger population. Both genetic and lifestyle factors may have contributed to its increased incidence and pathogenesis. Monosodium glutamate (MSG) is a food flavor enhancer that can be found in many commercial foods, and it can sometimes be used as a substitute to table salt. MSG has been investigated for its possible genotoxicity, yielding controversial results. In the present study, the effect of MSG on cell viability and its effect on expression of APC, BECN1, and TP53 genes in SW620 and SW480 colon cancer cell lines were studied. TP53 is a tumor suppressor gene that functions in modifying DNA errors and/or inducing apoptosis of damaged cells, and both APC and BECN1 genes are involved in CRC and are of importance in cellular growth and metastasis. Cancer cell viability was analyzed using MTT assay, and the results showed a significant increase in the number of viable cells after 24h of treatment with MSG with different concentrations (0.5, 1.0, 10, 50, and 100mM). Moreover, gene expression results showed a significant increase in the expression levels of APC and BECN1 under specified conditions in both cell lines; conversely, TP53 showed a significant decrease in expression in SW620 cells. Thus, it can be concluded that MSG possibly confers a pro-proliferative effect on CRC cells.

O câncer colorretal (CCR) é uma doença com alta incidência mundial. Desde 2018, é a segunda principal causa de mortes por câncer no mundo. Na Arábia Saudita, a incidência dessa doença vem aumentando na população mais jovem. Tanto fatores genéticos quanto de estilo de vida podem ter contribuído para o aumento da sua incidência e patogênese. O glutamato monossódico (MSG) é um intensificador de sabor de alimentos que pode ser encontrado em muitos alimentos comerciais e às vezes pode ser usado como um substituto do sal de cozinha. O MSG tem sido investigado por sua possível genotoxicidade, produzindo resultados controversos. Neste estudo, foram estudados o efeito do MSG na viabilidade celular e seu efeito na expressão dos genes APC, BECN1 e TP53 em linhas de células de câncer de cólon SW620 e SW480. TP53 é um gene supressor de tumor que atua modificando erros de DNA e/ou induzindo apoptose de células danificadas, estando os genes APC e BECN1 envolvidos no CRC e sendo importantes no crescimento celular e metástase. A viabilidade das células cancerosas foi analisada por meio do ensaio MTT, e os resultados mostraram um aumento significativo no número de células viáveis após 24 h de tratamento com MSG em diferentes concentrações (0,5; 1,0; 10; 50 e 100mM). Além disso, os resultados da expressão gênica mostraram um aumento significativo nos níveis de expressão de APC e BECN1 sob condições especificadas em ambas as linhagens celulares. Por outro lado, TP53 mostrou uma diminuição significativa na expressão em células SW620. Assim, pode-se concluir que, possivelmente, o MSG confere um efeito pró-proliferativo às células CRC.

Associations of cholecystectomy with the risk of colorectal cancer: a Mendelian randomization study / 中华医学杂志(英文版)

BACKGROUND@#Cholecystectomy is a standard surgery for patients suffering from gallbladder diseases, while the causal effects of cholecystectomy on colorectal cancer (CRC) and other complications are still unknown.@*METHODS@#We obtained genetic variants associated with cholecystectomy at a genome-wide significant level ( P value <5 × 10 -8 ) as instrumental variables (IVs) and performed Mendelian randomization (MR) to identify the complications of cholecystectomy. Furthermore, the cholelithiasis was also treated as the exposure to compare its causal effects to those of cholecystectomy, and multivariable MR analysis was carried out to judge whether the effect of cholecystectomy was independent of cholelithiasis. The study was reported based on Strengthening the Reporting of Observational Studies in Epidemiology Using Mendelian Randomization guidelines.@*RESULTS@#The selected IVs explained 1.76% variance of cholecystectomy. Our MR analysis suggested that cholecystectomy cannot elevate the risk of CRC (odds ratio [OR] =1.543, 95% confidence interval [CI]: 0.607-3.924). Also, it was not significant in either colon or rectum cancer. Intriguingly, cholecystectomy might decrease the risk of Crohn's disease (OR = 0.078, 95% CI: 0.016-0.368) and coronary heart disease (OR = 0.352, 95% CI: 0.164-0.756). However, it might increase the risk of irritable bowel syndrome (IBS) (OR = 7.573, 95% CI: 1.096-52.318). Cholelithiasis could increase the risk of CRC in the largest population (OR = 1.041, 95% CI: 1.010-1.073). The multivariable MR analysis suggested that genetic liability to cholelithiasis could increase the risk of CRC in the largest population (OR = 1.061, 95% CI: 1.002-1.125) after adjustment of cholecystectomy.@*CONCLUSIONS@#The study indicated that cholecystectomy might not increase the risk of CRC, but such a conclusion needs further proving by clinical equivalence. Additionally, it might increase the risk of IBS, which should be paid attention to in clinical practice.

Caracterización molecular de pacientes con cáncer colorrectal / Molecular characterization of colorectal cancer patients

Introducción. El cáncer colorrectal tiene una alta incidencia en la población mundial. Diversas vías moleculares están involucradas en su desarrollo, entre ellas, la inestabilidad cromosómica, la inestabilidad microsatelital y la epigenética. Objetivo. Hacer la caracterización molecular de 44 individuos con cáncer colorrectal esporádico. Materiales y métodos. El análisis de mutaciones en los genes APC, KRAS, TP53 y BRAF se hizo mediante secuenciación de Sanger; la inestabilidad microsatelital se determinó mediante electroforesis capilar utilizando cinco marcadores de repetición corta en tándem (Short Tandem Repeat) y el estado de metilación del promotor del gen MLH1 se hizo con la técnica MS-PCR (Methylation-Specific PCR). Resultados. La frecuencia de mutación de los genes APC, KRAS y TP53 fue del 18,1, 25 y 4,5 %, respectivamente; las mutaciones detectadas se localizaron con mayor frecuencia en el colon derecho. La frecuencia de inestabilidad microsatelital fue del 27,2 % y el 73,1 % en los tumores con metilación en el gen MHL1, y el 91,6 % de los tumores con inestabilidad microsatelital presentaba metilación en el gen MLH1. En el grupo de tumores con estabilidad microsatelital, las mutaciones en los genes APC, KRAS y TP53 fueron más frecuentes que en el grupo de tumores con inestabilidad microsatelital. La metilación del gen MLH1 fue la alteración más predominante. Conclusiones. En los pacientes con cáncer colorrectal evaluados se demostró la presencia de alteraciones moleculares en las diferentes vías genéticas, las cuales son comunes en su carcinogénesis. Los pacientes presentaron un perfil de mutaciones diferente al de otras poblaciones. Los hallazgos obtenidos en este estudio confirman la heterogeneidad molecular descrita en el desarrollo del cáncer colorrectal.

Introduction: Colorectal cancer has a high incidence in the world population. Different molecular pathways, such as chromosomal instability, microsatellite instability, and epigenetics are involved in its development. Objective: To perform molecular characterization in 44 individuals with sporadic colorectal cancer. Materials and methods: We conducted mutation analyses of the APC, KRAS, TP53 y BRAF genes using Sanger sequencing techniques; microsatellite instability was determined by capillary electrophoresis with five STR genetic markers while the methylation status of the MHL1 promotor gene was analyzed using methylation-specific PCR. Results: APC, KRAS, and TP53 genes mutation frequency was 18.1%, 25%, and 4.5%, respectively; the somatic mutations detected were located more frequently in the right colon. The frequency of microsatellite instability was 27.2% and 73.1% of the tumors had the MHL1 gene methylated while 91.6% of microsatellite instability-positive tumors had the methylated MLH1 gene. The mutation profile of microsatellite stability tumors APC, KRAS, and TP53 genes was more frequent than in the microsatellite instability-positive tumors. The methylation of the MLH1 gene was the most predominant molecular alteration. Conclusions: We identified molecular alterations in different genetic pathways of the colorectal cancer patients evaluated, which are common in the carcinogenesis of this cancer. These patients showed a different mutational profile compared to other populations. Our findings confirm the molecular heterogeneity described in the development of colorectal cancer.

Correlation of NTRK genetic fusions with mismatch repair protein deletion in patients with colorectal cancer / 中华病理学杂志

Objective: To investigate the relationship between the expression of four mismatch repair proteins (MLH1, MSH2, MSH6 and PMS2) and NTRK genetic fusions in colorectal cancer. Methods: The paraffin-embedded tissue blocks of 830 cases of colorectal cancer were collected at the Affiliated Drum Tower Hospital, Nanjing University Medical School, China, from 2015 to 2019. Immunohistochemical and fluorescence in situ hybridization(FISH) method were used respectively to detect the expression of mismatch repair proteins and the break-apart of NTRKs; and the relationship between the expression of mismatch repair proteins and the NTRK genetic fusions was analyzed. Results: The overall mismatch repair protein deficiency (dMMR) rate was 9.88% (82/830), the mismatch repair proteins proficiency (pMMR) rate was 90.12%(748/830). The total deficiency rate of MLH1 protein was 9.04% (75/830), hPMS2 protein deficiency rate was 9.04% (75/830), MSH2 protein deficiency rate was 2.53% (21/830), MSH6 protein deficiency rate was 4.10% (34/830), the deficiency rate of synchronous MLH1 and PMS2 were 8.67% (72/830) and the deficiency rate of synchronous MSH2 and MSH6 were 2.17% (18/830). The dMMR group was associated with tumor location, different histological subgroups, tumor differentiation, AJCC stage and N stage (P<0.05). There were six cases (7.32%) carrying NTRK fusion by FISH among the 82 cases of dMMR, but only seven cases (0.94%) carrying NTRK fusion among the 748 cases of PMMR. The NTRKs translocation by FISH in all 13 cases were further confirmed by next generation sequencing. Among the clinicopathological characteristics, only differentiation showed significant difference between NTRK fusion positive and negative groups (P<0.05). More importantly, NTRK fusion was enriched in dMMR group (7.32% vs. 0.94%). Conclusion: In dMMR colorectal cancer group, the prevalence of NTRK fusion is higher than that in pMMR group.

Genetic variation of YWHAE gene-"Switch" of disease control / 中南大学学报(医学版)

YWHAE gene is located on chromosome 17p13.3, and its product 14-3-3epsilon protein belongs to 14-3-3 protein family. As a molecular scaffold, YWHAE participates in biological processes such as cell adhesion, cell cycle regulation, signal transduction and malignant transformation, and is closely related to many diseases. Overexpression of YWHAE in breast cancer can increase the ability of proliferation, migration and invasion of breast cancer cells. In gastric cancer, YWHAE acts as a negative regulator of MYC and CDC25B, which reduces their expression and inhibits the proliferation, migration, and invasion of gastric cancer cells, and enhances YWHAE-mediated transactivation of NF-κB through CagA. In colorectal cancer, YWHAE lncRNA, as a sponge molecule of miR-323a-3p and miR-532-5p, can compete for endogenous RNA through direct interaction with miR-323a-3p and miR-532-5p, thus up-regulating K-RAS/ERK/1/2 and PI3K-AKT signaling pathways and promoting the cell cycle progression of the colorectal cancer. YWHAE not only mediates tumorigenesis as a competitive endogenous RNA, but also affects gene expression through chromosome variation. For example, the FAM22B-YWHAE fusion gene caused by t(10; 17) (q22; p13) may be associated with the development of endometrial stromal sarcoma. At the same time, the fusion transcript of YWHAE and NUTM2B/E may also lead to the occurrence of endometrial stromal sarcoma. To understand the relationship between YWHAE, NUTM2A, and NUTM2B gene rearrangement/fusion and malignant tumor, YWHAE-FAM22 fusion gene/translocation and tumor, YWHAE gene polymorphism and mental illness, as well as the relationship between 17p13.3 region change and disease occurrence. It provides new idea and basis for understanding the effect of YWHAE gene molecular mechanism and genetic variation on the disease progression, and for the targeted for the diseases.

Construction of endogenous RNA regulatory network for colorectal cancer based on bioinformatics / 中南大学学报(医学版)

OBJECTIVES@#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. This paper aims to construct a circRNA/miRNA/mRNA regulatory network so as to explore the molecular mechanism of CRC.@*METHODS@#The sequencing data of circRNA from CRC were obtained from Gene Expression Omnibus (GEO). The differential circRNA was screened and its structure was identified by Cancer-specific CircRNA Database (CSCD); the sequencing data of miRNA and messenger RNA (mRNAs) were downloaded from The Cancer Genome Atlas (TCGA) database and the differentially expressed genes were screened; the corresponding miRNA of differential circRNAs were predicted by CircInteractome database; DIANA, Miranda, PicTar, and TargetScan databases were used to predict the target genes of different miRNAs; the target genes from Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were enriched by R language; String database combined with Cytoscape 3.7.2 software was used to construct protein-protein interaction (PPI) network and hub genes were screened; the expressions of mRNAs in the Top10 hub genes were verified in CRC. The network diagrams of circRNAs/miRNAs/mRNAs and circRNAs/miRNAs/Top10 hub mRNAs were constructed by Cytoscape3.7.2. Real-time PCR was used to examine the expression levels of hsa_circRNA_0065173, hsa-mir-450b, hsa-mir-582, adenylate cyclase 5 (ADCY5), muscarinic acetylcholine receptor M2 (CHRM2), cannabinoid receptor 1 (CNR1), and lysophosphatidic acid receptor 1 (LPAR1) in the CRC tissues and the adjacent normal tissues.@*RESULTS@#A total of 14 differential circRNAs were identified, and 8 were found in CSCD; 34 miRNAs targeted by circRNAs were obtained. The PPI network was constructed, and the Top10 hub genes were identified, which were CHRM2, melanin concentrating hormone receptor 2 (MCHR2), G-protein gamma 3 subunit (GNG3), neuropeptide Y receptor Y1 (NPY1R), CNR1, LPAR1, ADCY5, adenylate cyclase 2 (ADCY2), gamma 7 (GNG7) and chemokine 12 (CXCL12), respectively. The expressions of Top 10 hub genes were also verified, and the results showed that the Top 10 hub genes were down-regulated in CRC; the constructed network diagram showed that hsa_circRNA_0065173 may regulate ADCY5, CHRM2, and Hsa-mir-450b by modulating hsa-mir-450b and hsa-mir-582. CNR1 and LPAR1 genes might serve as potentially relevant targets for the treatment of CRC. Real-time PCR results showed that the expression levels of hsa_circRNA_0065173, ADCY5, CHRM2, CNR1 and LPAR1 in the CRC tissues were significantly reduced compared with the adjacent normal tissues (all P<0.05); the expression levels of hsa-mir-450b and hsa-miR-582 were significantly increased (both P<0.05).@*CONCLUSIONS@#In this study, a potential circRNAs/miRNAs/mRNAs network is successfully constructed, which provides a new insight for CRC development mechanism through ceRNA mediated by circRNAs.

Screening of differentially expressed genes for colorectal cancer and prediction of potential traditional Chinese medicine: based on bioinformatics / 中国中药杂志

This study screened and analyzed the differentially expressed genes(DEGs) between colorectal cancer(CRC) tissues and normal tissues with bioinformatics techniques to predict biomarkers and Chinese medicinals for the diagnosis and treatment of CRC. The microarray data sets GSE21815, GSE106582, and GSE41657 were downloaded from the Gene Expression Omnibus(GEO), and the DEGs were screened by GEO2 R, followed by the Gene Ontology(GO) tern enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis of the DEGs based on DAVID. The protein-protein interaction network was constructed by STRING, and MCODE and Cytohubba plug-ins were used to screen the significant modules and hub genes in the network. UCSC, cBioPortal, and Oncomine were employed for hierarchical clustering, survival analysis, Oncomine analysis, and correlation analysis of clinical data. Coremine Medical was applied to predict the Chinese medicinals acting on hub genes. A total of 284 DEGs were screened out, with 146 up-regulated and 138 down-regulated. The up-regulated genes were mainly involved in cell cycle, NLRs pathway, and TNF signaling pathway, and the down-regulated genes were related to mineral absorption, nitrogen metabolism, and bicarbonate reabsorption in proximal tubules. The 15 hub genes were CDK1, CDC20, AURKA, MELK, TOP2 A, PTTG1, BUB1, CDCA5, CDC45, TPX2, NEK2, CEP55, CENPN, TRIP13, and GINS2, among which CDK1 and CDC20 were regarded as core genes. The high expression of CDK1 and CDC20 suggested poor prognosis, and they significantly expressed in many cancers, especially breast cancer, lung cancer, and CRC. The expression of CDK1 and CDC20 was correlated with gender, tumor type, TNM stage, and KRAS gene mutation. The potential effective medicinals against CRC were Scutellariae Radix, Scutellariae Barbatae Herba, Arnebiae Radix, etc. The significant expression of CDK1 and CDC20 can help distinguish tumor tissues from normal tissues, and is related to survival prognosis. Thus, the two can be used as biomarkers for the diagnosis and treatment of CRC. This study provides a reference for related drug development.

Effect of Astragali Radix-Curcumae Rhizoma compatibility combined with 5-fluorouracil on Th17/Treg balance and tumor-related mRNA and protein expression in orthotopic xenograft model mice of CT26.WT colorectal carcinoma / 中国中药杂志

Astragali Radix-Curcumae Rhizoma(AR-CR) is a combination commonly used in the clinical treatment of tumors. Based on the T helper 17(Th17)/regulatory T cell(Treg) balance, the present study explored the possible mechanism of AR-CR combined with 5-fluorouracil(5-FU) on the tumor growth of orthotopic xenograft model mice of colorectal carcinoma. Ninety male BALB/c mice were randomly divided into nine groups, i.e., a blank group, a model group, a 5-FU group, high-, medium-, and low-dose AR-CR(2∶1) groups, and high-, medium-, and low-dose AR-CR+5-FU groups, with 10 mice in each group. The orthotopic xenograft model of CT26.WT colorectal carcinoma was induced in mice except those in the blank group. Twenty-four hours after the ope-ration, mice in the blank group and the model group received normal saline by gavage(10 mL·kg~(-1), once per day), and those in the 5-FU group received 5-FU by intraperitoneal injection(25 mg·kg~(-1), once every other day). Mice in the AR-CR groups received AR and CR decoctions by gavage(12, 6, and 3 g·kg~(-1), once a day) and those in the combination groups received AR and CR decoctions and 5-FU(doses and administration methods were the same as above). After intervention for three weeks, all mice were sacrificed and tumor tissues were collected. The tumor mass was weighed and the average tumor weight was calculated. The changing trend of Th17/Treg(%) in the CD4~+T lymphocytes of the spleen tissues of the mice in each group was detected. The mRNA expression in the blood and protein expression in the tumor tissues of transforming growth factor-β(TGF-β), tumor necrosis factor-α(TNF-α), interferon-γ(IFN-γ), Smad4, N-cadherin, matrix metalloproteinase-7(MMP-7) were detected. The experimental results revealed that compared with the model group, the groups with drug intervention showed reduced tumor mass(P<0.01), decreased CD4~+IL-17~+ in the spleen tissues to varying degrees(P<0.001), and increased proportion of CD4~+Foxp3~+(P<0.001 or P<0.05), indicating that Th17/Treg maintained dynamic balance, and the effect of the combination groups was predominant. Additionally, the mRNA expression in the blood and protein expression in the tumor tissues of TGF-β, TNF-α, IFN-γ, Smad4, N-cadherin, and MMP-7 declined to varying degrees in a dose-dependent manner(P<0.01 or P<0.001). The AR-CR combined with 5-FU can inhibit the tumor growth of orthotopic xenograft model mice of CT26.WT colorectal carcinoma. The mechanism may be related to maintenance of Th17/Treg dynamic balance in the body and down-regulation of TGF-β, TNF-α, IFN-γ, Smad4, N-cadherin, and MMP-7 expression.

β-Elemene induces apoptosis and autophagy in colorectal cancer cells through regulating the ROS/AMPK/mTOR pathway / 中国天然药物

β-Elemene is an effective anti-cancer ingredient extracted from the genus Curcuma (Zingiberaceae familiy). In the present study, we demonstrated that β-elemene inhibited the proliferation of colorectal cancer cells and induced cell cycle arrest in the G2/M phase. In addition, β-elemene induced nuclear chromatin condensation and cell membrane phosphatidylserine eversion, decreased cell mitochondrial membrane potential, and promoted the cleavage of caspase-3, caspase-9 and PARP proteins, indicating apoptosis in colorectal cancer cells. At the same time, β-elemene induced autophagy response, and the treated cells showed autophagic vesicle bilayer membrane structure, which was accompanied by up-regulation of the expression of LC3B and SQSTM1. Furthermore, β-elemene increased ROS levels in colorectal cancer cells, promoted phosphorylation of AMPK protein, and inhibited mTOR protein phosphorylation. In the experiments in vivo, β-elemene inhibited the tumor size and induced apoptosis and autophagy in nude mice. In summary, β-elemene inhibited the occurrence and development of colon cancer xenografts in nude mice, and significantly induced apoptosis and autophagy in colorectal cancer cells in vitro. These effects were associated with regulation of the ROS/AMPK/mTOR signaling. We offered a molecular basis for the development of β-elemene as a promising anti-tumor drug candidate for colorectal cancer.

Establishment of a prediction model for colorectal cancer immune cell infiltration based on the cancer genome atlas (TCGA) database / 北京大学学报(医学版)

OBJECTIVE@#To study the correlation between immune cell infiltration in colorectal cancer tissue and clinical prognosis and to explore the levels of some immune cell genes for predicting the prognosis of patients with glioma colorectal cancer.@*METHODS@#In this study, we extracted colorectal cancer data from the cancer genome atlas (TCGA). Based on a deconvolution algorithm (called CIBERSORT) and clinically annotated expression profiles, the analysis assessed the infiltration patterns of 22 immune cells in colorectal cancer tissue to determine the association between each cell type and survival. Differences in five-year survival rate effectively illustrate the clinical prognostic value of each immune cell proportion in colorectal cancer, using a bar graph, correlation-based heatmap to represent the proportion of immune cells in each colorectal cancer sample.@*RESULTS@#A total of 473 colorectal cancer tissues and 41 normal control tissues were extracted from the TCGA database, and the comparative analysis showed that there were differences in the proportion of various TIICs in colorectal cancer tissues, which could characterize individual differences and have prognostic value. Among the cell subsets studied, the proportions of memory B cells, plasma cells, CD4+ T cells, natural killer (NK) cells, M0 macrophages, M2 macrophages, and activated mast cells were significantly different between normal and cancer tissues. Resting NK cells, CD8+ T cells, and plasma cells were associated with T phase, activated dendritic cells were associated with N phase, and eosinophils, M1 macrophages, and activated mast cells were associated with M phase. Survival analysis showed that activated dendritic cells were positively associated with five-year survival rate in colorectal cancer patients. Naive CD4+ T cells were inversely associated with five-year survival rate.@*CONCLUSION@#There are different degrees of immune cell infiltration in colorectal cancer tissues, and these differences may be important determinants of prognosis and treatment response. We conducted a new gene expression-based study of immune cell subtype levels and prognosis in colorectal cancer, which has potential clinical prognostic value in colorectal cancer patients.

Carcinoma colorrectal en niños y adolescentes / Colorectal carcinoma in children and adolescent

Si bien el carcinoma colorrectal (CCR) es la tercera enfermedad oncológica más frecuente en adultos, solo el 1-4 % ocurre en menores de 25-30 años. Su presentación suele confundirse con otras patologías, lo cual lleva a significativas demoras diagnósticas. Dada su baja incidencia, pocos pediatras se enfrentarán con algún caso a lo largo de su carrera. Sin embargo, existen en la niñez múltiples síndromes hereditarios que predisponen al CCR. El objetivo de esta revisión es brindar una actualización sobre los síndromes predisponentes al CCR. Se revisarán las indicaciones de tamizaje, dado que el diagnóstico precoz en estadios localizados es el principal factor pronóstico. Además, se fortalecerán las herramientas de consejería genética para el paciente y su familia. A su vez, se discutirán las manifestaciones clínicas e histológicas y los factores pronósticos propios del CCR en la población pediátrica. Si bien las guías de tratamiento se extrapolan de la experiencia en adultos, se resumirán los lineamientos terapéuticos.

Although colorectal carcinoma (CRC) is the third most common type of cancer in adults, only 1-4 % of cases are reported in individuals younger than 25-30 years. Its presentation is usually confused with other diseases, leading to significant delays in diagnosis. Given its low incidence, few pediatricians will see a case throughout their practice. However, multiple hereditary syndromes during childhood predispose to CRC.The objective of this review is to provide an update on syndromes predisposing to CRC. Screening indications will be reviewed because an early diagnosis during localized stages is the main prognostic factor. In addition, patient and family genetic counseling tools will be enhanced. In turn, the clinical and histological manifestations and prognostic factors typical of CRC in the pediatric population will be discussed. Although treatment guidelines are extrapolated from the adult experience, therapy guidelines will be summarized here.

Expressão do RNAm e metilação do DNA do gene DKK2 em câncer colorretal / Rnam expression and dna methylation of dkk2 gene in colorectal câncer

ABSTRACT BACKGROUND: Colorectal cancer is the third most common neoplasm in the world. Methylation of tumor related genes in CpG islands can cause gene silencing and been involved in the development of cancer. The potential role of DKK2 as a biomarker for early diagnosis of colorectal cancer remains unclear. OBJECTIVE: The aim of the study was to evaluate the profile of methylation and RNAm expression of DKK2 as potential predictors of colorectal cancer diagnosis and prognosis. METHODS: Expression of mRNAs encoding DKK2 in 35 colorectal cancer tissues was quantified using real-time polymerase chain reaction analysis. The DNA methylation was studied by high resolution melting analysis. The general characteristics of the patients were collected. DKK2 methylation and expression were compared to clinical, pathological aspects and overall survival. RESULTS: Among the 35 patients studied, 18 were male, 10 were on right colon and 25 on left colon. Among the 20 patients with high hypermethylation, 15 of them had mRNA low expression of DKK2. There was no significant association between DKK2 promoter methylation and mRNA DKK2 expression and clinical or pathological features. DKK2 promoter methylation (P=0.154) and DKK2 RNA expression (P=0.345) did not show significant correlation with overall survival. CONCLUSION: DKK2 promoter methylation and DKK2 RNA status appear to be biomarkers of cancer diagnosis but not predictors of prognosis.

RESUMO CONTEXTO: O câncer colorretal é a terceira neoplasia mais comum no mundo. A metilação de alguns genes nas ilhas CpG podem causar silenciamento gênico e estar envolvida no desenvolvimento de câncer. O potencial papel de DKK2 como um biomarcador no diagnóstico precoce de CCR permanece incerto. OBJETIVO: O objetivo do estudo foi avaliar o perfil de metilação e expressão de RNAm do gene DKK2 para identificar preditores potenciais de diagnóstico e prognóstico de CCR. MÉTODOS: A expressão de mRNAs que codificam DKK2 em 35 tecidos de câncer colorretal foi quantificada por reação em cadeia da polimerase em tempo real e a metilação do DNA foi verificada por análise de alta resolução. As características gerais dos pacientes foram coletadas. A metilação e expressão de DKK2 foram comparadas aos aspectos clínicos, patológicos e à sobrevida global. RESULTADOS: Entre os 35 pacientes estudados, 18 eram do sexo masculino, 10 tumores eram do cólon ascendente ou transverso e 25 do descendente ou reto. Entre os 20 pacientes com hipermetilação, 12 deles apresentaram baixa expressão de RNAm do gene DKK2. Não houve associação significativa entre a metilação do promotor de DKK2 e a expressão de RNAm de DKK2 e características clínicas ou patológicas. A metilação do promotor de DKK2 e a expressão do RNA de DKK2 não mostraram correlação com sobrevida global dos pacientes com CCR. CONCLUSÃO: A metilação do gene promotor e a expressão do RNAm do gene DKK2 parecem ser biomarcadores de diagnóstico de câncer, mas não se mostraram úteis na avaliação prognóstica.

The effect of mutation status, pathological features and tumor location on prognosis in patients with colorectal cancer

SUMMARY OBJECTIVE: Colorectal cancer is the most common malignancy of the gastrointestinal tract. It is the third most common tumor in both genders and the second reason of cancer-related deaths. In recent years, tumor location has gained importance as a prognostic indicator. In this study, we aimed to analyze if there was a prognostic effect of tumor location, the pathological features, and the mutation status of patients on survival. METHODS: Two-hundred and ten colorectal cancer patients aged 18 years and older were included into the study. One-hundred and forty-two patients had left-sided tumor and 68 patients had right-sided tumor. Patients who had other malignancies rather than squamous cell skin cancer and in situ cervical cancer were excluded. All statistical tests were carried out using two-sided process, and a p≤0.05 was considered statistically significant. RESULTS: There were 140 men and 70 women in the study. The median age of the patients was 62 years old. There was no statistically significant difference according to tumor location and survival of patients. The overall survival of patients with right-sided tumors was 60.5 months and 47.2 months for left-sided tumors. Disease-free survival of patients was 63.7 months for right-sided tumors and 46 months for left-sided ones. Perineural invasion, grade and stage were crucial prognostic parameters. Disease-free survival was longer for female colorectal cancer patients. CONCLUSION: According to our study, survival of patients was similar regardless of tumor location. This can be explained by the different sequencing of treatment strategies and divergent population genetics.

Consensus of Chinese experts on clinical detection of molecular markers of colorectal cancer / 中华胃肠外科杂志

Colorectal cancer is one of the malignant tumors with the highest morbidity and mortality in China. With the research of precision medicine concept and tumor-related molecular markers, appropriate detection and application of colorectal cancer-related molecular markers has become an important part of current clinical practice. In order to effectively solve the current clinical problems and improve clinicians' understanding and application of molecular markers on colorectal cancer, the Chinese Society of Clinical Oncology(CSCO) Colorectal Cancer Expert Committee organized experts in related fields to write an expert consensus on molecular markers of colorectal cancer based on recent domestic and international clinical trial and clinical experience. The consensus mainly provides guidance on testing specimens, molecular markers and testing methods, and interpretation of testing results. It aims to provide clinicians with standardized clinical reference for diagnosis and treatment, and standard and effective treatment for patients with colorectal cancer.