Identification of Key Prognostic Alternative Splicing Events of Costimulatory Molecule-Related Genes in Colon Cancer.

OBJECTIVE: This study aimed to explore the key alternative splicing events in costimulatory molecule-related genes in colon cancer and to determine their correlation with prognosis. METHODS: Gene expression RNA-sequencing data, clinical data, and SpliceSeq data of colon cancer were obtained from The Cancer Genome Atlas. Differentially expressed alternative splicing events in genes were identified, Followed by correlation analysis of genes corresponding to differentially expressed alternative splicing events with costimulatory molecule-related genes. Survival analysis was conducted using differentially expressed alternative splicing events in these genes and a prognostic model was constructed. Functional enrichment, proteinprotein interaction network, and splicing factor analyses were performed. RESULTS: In total, 6504 differentially expressed alternative splicing events in 3949 genes were identified between tumor and normal tissues. Correlation analysis revealed 3499 differentially expressed alternative splicing events in 2168 costimulatory molecule-related genes. Moreover, 328 differentially expressed alternative splicing events in 288 costimulatory molecule-related genes were associated with overall survival. The prognostic models constructed using these showed considerable power in predicting survival. The ubiquitin A-52 residue ribosomal protein fusion product 1 and ribosomal protein S9 were the hub nodes in the protein-protein interaction network. Furthermore, one splicing factor, splicing factor proline and glutamine-rich, was significantly associated with patient prognosis. Four splicing factor-alternative splicing pairs were obtained from four alternative splicing events in three genes: TBC1 domain family member 8 B, complement factor H, and mitochondrial fission 1. CONCLUSION: The identified differentially expressed alternative splicing events of costimulatory molecule-related genes may be used to predict patient prognosis and immunotherapy responses in colon cancer.

LncRNA cancer susceptibility 20 regulates the metastasis of human gastric cancer cells via the miR-143-5p/MEMO1 molecular axis.

BACKGROUND: Gastric cancer (GC) is considered as one of the most widespread malignancies. Emerging evidence has shown that lncRNAs can function as important oncogenes or tumor suppressors during GC progression. AIM: To investigate the effect and mechanism of lncRNA cancer susceptibility 20 (CASC20) in the proliferation and metastasis of GC cells. METHODS: Data mining and clinical samples were used to evaluate the expression of CASC20 in GC and adjacent tissues. CASC20 was down-regulated in GC cells by short-interfering RNA. Cell proliferation was evaluated by CCK-8 assay, and cell migration and invasion were detected by wound healing and Transwell assays. The expressions of proteins related to epithelial-mesenchymal transition were detected by western blot assay. RESULTS: The expression of CASC20 was increased in GC tumor tissues and various GC cell lines. High CASC20 expression was correlated with a high risk of lymphatic metastasis and poor prognosis in GC patients. In vitro assays showed that silencing CASC20 reduced cell proliferation, migration, and invasion in GC cells. Mechanistic studies revealed that CASC20 exhibits oncogenic functions by regulating MEMO1 expression through competitive endogenous binding to miR-143-5p, leading to induction of epithelial-mesenchymal transition. CONCLUSION: Our findings indicate that CASC20 serves as a tumor promoter by regulating metastasis in GC via the miR-143-5p/MEMO1 axis. CASC20 may be a potential therapeutic target for GC.

Circular RNA circPFKP suppresses the proliferation and metastasis of gastric cancer cell via sponging miR-644 and regulating ADAMTSL5 expression.

The treatment of gastric cancer (GC) is extremely challenging; however, the specific pathogenesis of GC remains unclear. Circular RNAs (CircRNAs) are non-coding RNAs that can regulate gene expression both transcriptionally and post-transcriptionally. However, little is known about the circRNAs that are important in the progression of GC. This study identified significantly dysregulated circRNAs by analyzing gastric cancer patients and normal control tissues. The target gene was predicted using online bioinformatics tools and verified using RNA pull-down and luciferase reporter assays. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were used to evaluate gene and protein expression. The malignant behavior of GC cells was determined using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay, wound healing assay, Transwell invasion assay, and flow cytometry. CircPFKP is downregulated in GC tissues, and overexpression of circPFKP inhibits malignant behavior in GC cells. Bioinformatics predicted that circPFKP could bind to miR-644, and miR-644 could target disintegrin-like and metalloprotease domain-containing thrombospondin type 1 motif-like 5 (ADAMTSL5). Overexpression of circPFKP enhances the expression of ADAMTSL5 by decreasing the expression of miR-644 to suppress the growth of xenograft GC tumors in vivo and in vitro. In conclusion, the circPFKP/miR-644/ADAMTSL5 regulatory pathway inhibited the malignant progression of GC. These findings may extend our understanding of the effects of circRNAs on cancer development and provide novel targets for the diagnosis of GC.

CCDC12 promotes tumor development and invasion through the Snail pathway in colon adenocarcinoma.

Integrative expression Quantitative Trait Loci (eQTL) analysis found that rs8180040 was significantly associated with Coiled-coil domain containing 12 (CCDC12) in colon adenocarcinoma (COAD) patients. Immunohistochemical staining and western blotting confirmed CCDC12 was highly expressed in COAD tissues, which was consistent with RNA-Seq data from the TCGA database. Knockdown of CCDC12 could significantly reduce proliferation, migration, invasion, and tumorigenicity of colon cancer cells, while exogenous overexpression of CCDC12 had the opposite effect. Four plex Isobaric Tags for Relative and Absolute Quantitation assays were performed to determine its function and potential regulatory mechanism and demonstrated that overexpression of CCDC12 would change proteins on the adherens junction pathway. Overexpressed Snail and knocked down CCDC12 subsequently in SW480 cells, and we found that overexpression of Snail did not significantly change CCDC12 levels in SW480 cells, while knockdown of CCDC12 reduced that of Snail. CCDC12 plays a significant role in tumorigenesis, development, and invasion of COAD and may affect the epithelial to mesenchymal transformation process of colon cancer cells by regulating the Snail pathway.

One-stitch method vs. traditional method of protective loop ileostomy for rectal cancer: the impact of BMI obesity.

PURPOSE: Protective loop ileostomy is an effective diversion measure often used to reduce the risk of anastomotic leakage. The purpose of the present study was to evaluate the surgical outcomes of the one-stitch method (OM) of protective loop ileostomy in laparoscopic low anterior resection for BMI obesity patients with rectal cancer compared with the traditional method (TM). METHODS: The patients diagnosed as rectal adenocarcinoma cases by preoperative pathology were included in this retrospective study. The subjects underwent protective loop ileostomy in laparoscopic low anterior resection from January 2016 to June 2019 in the Shandong Provincial Hospital affiliated to Shandong University. The data of loop ileostomy and stoma closure operation were retrieved from the medical cases system of the hospital. RESULTS: 242 patients were included in the present study. In the BMI obese cohort, the OM group showed a shorter operative time both in the loop ileostomy (232.5 vs. 250.0 min, p = 0.04) and stoma closure operation (102.5 vs. 115.0 min, p = 0.001) and a lower peristomal adhesion extent (p = 0.02) and a shorter median postoperative stay (6 vs. 7 days, p = 0.03) during stoma closure operation than that of the TM group. In the TM group, obese cases showed a higher operative time of stoma closure operation (115.0 vs. 95.0, p < 0.001), a higher parastomal hernia rate (p = 0.04), a higher peristomal adhesion extent (p = 0.005) and a longer postoperative stay of stoma closure operation (p = 0.02) compared with the non-obese cases, while in the OM group, no significant differences were observed between the obese and non-obese cases in terms of the above-mentioned factors. CONCLUSIONS: The OM exhibited more advantages than TM, notably in BMI obesity patients.

Association between R1 resection and oncological outcome in resectable gastrointestinal stromal tumors without tumor rupture: A systematic review and meta-analysis.

BACKGROUND: The influence of positive microscopic margin (R1) resection on the prognosis of gastrointestinal stromal tumors (GISTs) is controversial. Tumor rupture is significantly associated with the occurrence of R1 resection and may be a confounder of R1 resection in GISTs. The present meta-analysis evaluated the real influence of R1 resection on the prognosis of GISTs by excluding the confounding effect of tumor rupture. METHODS: The PubMed, Embase, Web of Science, Cochrane Central Register of Controlled Trials (CENTRAL), and ClinicalTrials.gov databases were searched. Studies that compared R1 with negative microscopic margin (R0) resection in GIST patients and reported the time-to-event data of recurrence-free survival (RFS) or disease-free survival (DFS) were eligible for inclusion. The quality of the observational studies was assessed using the Newcastle-Ottawa scale. RESULTS: Of the 4896 records screened, 23 retrospective studies with 6248 participants were selected. In the overall analysis, R1 resection resulted in a significantly shorter RFS/DFS than R0 resection for GISTs (HR = 1.80, 95% CI = 1.54-2.10, P < 0.001, I2 = 14%). However, the inferior RFS/DFS vanished when tumor rupture cases were excluded (HR = 1.34, 95% CI = 0.98-1.83, P = 0.07, I2 = 33%). Sensitivity analysis by high-quality studies brought about a more robust HR of 1.15 (95% CI = 0.88-1.50, P = 0.29), with low heterogeneity (I2 = 0%). The qualities of evidence for the outcomes were high. CONCLUSIONS: This meta-analysis shows that R1 resection did not influence the survival outcome of GISTs. Reresection may not be necessary when positive microscopic margins exist. This analysis could provide high-quality evidence for the development of guidelines.

Genetic Fine Mapping and Genomic Annotation Defines Causal Mechanisms at A Novel Colorectal Cancer Susceptibility Locus in Han Chinese.

Genome-wide association studies of colorectal cancer (CRC) have identified two risk SNPs. The characterization of these risk regions in diverse racial groups with different linkage disequilibrium structure would aid in localizing the causal variants. Herein, fine mapping of the established CRC loci was carried out in 1,508 cases and 1,482 controls obtained from the Han Chinese population. One distinct association signal was identified at these loci, where fine mapping implicated rs1010208 as a functional locus. Next, the candidate target genes of functional SNP rs1010208 were analyzed using data from TCGA databases by expression quantitative trait loci analysis method; the data from Peking University People's Hospital were utilized for verification. The dual-luciferase reporter system analysis confirmed that rs1010208 is a regulatory region that can be mutated to decrease the expression of HINT1, resulting in proliferation and invasiveness of CRC.

Fibroblast growth factor receptor-like-1: a new therapeutic target and unfavorable prognostic indicator for rectal adenocarcinoma.

Fibroblast growth factor receptor-like-1 (FGFRL1) is important to cell motility and links with tumorigenic potential in various types of cancers. To investigate the biological function and underlying mechanism of FGFRL1 in rectal adenocarcinoma, we conducted this study. TCGA and Oncomine databases were used to analyze FGFRL1 expression and its association with clinical characteristics or overall survival (OS) in rectal adenocarcinoma patients. siRNA strategy was implemented to knockdown FGFRL1 expression in rectal adenocarcinoma cells. CCK8, colony formation, wound healing, and transwell assays were implemented to measure cell behaviors. qRT-PCR and western blot were utilized to identify mRNA and protein expression levels. FGFRL1 was significantly increased in rectal adenocarcinoma tissue samples, either colon or rectum. High-regulation of FGFRL1 expression induced poorer outcome of rectal adenocarcinoma patients. Downregulation of FGFRL1 inhibited the proliferation, colony formation, migration, and invasion of SW837 cells. The MAPK pathway-related proteins, phosphorylation of MEK and ERK, were also decreased after si-FGFRL1 transfection. These findings demonstrated that FGFRL1, acting as a potential inducator, may promote the progression of rectal adenocarcinoma via activating the MAPK signaling pathway.

Silencing Livin improved the sensitivity of colon cancer cells to 5-fluorouracil by regulating crosstalk between apoptosis and autophagy.

Colorectal cancer (CRC) is the third most common cause of cancer-associated mortality worldwide. Currently, 5-fluorouracil (5-FU) remains a widely used chemotherapeutic drug in the treatment of CRC; however, 5-FU resistance during treatment has become a common problem. Livin, a member of the inhibitor of apoptosis protein family, is considered to be associated with tumor resistance to chemotherapy. In the present study, Livin-silenced cells were generated by introducing a lentivirus into HCT116 and SW620 colon cancer cell lines. Acridine orange/ethidium bromide staining was used as an indicator of cell death. Western blot analysis was performed to detect protein expression levels, and transmission electron microscopy was used to assess autophagy. The half-maximal inhibitory concentration of 5-FU in colon cancer cells was evaluated using a Cell Counting Kit-8 assay. The results of the present study confirmed that silencing Livin significantly enhanced colon cancer cell death in the presence of 5-FU, increased expression levels of various apoptosis- and autophagy-associated proteins and augmented chemotherapeutic sensitivity to 5-FU. Furthermore, the present study demonstrated that this effect may be reversed when autophagy or apoptosis was inhibited, indicating that apoptosis and autophagy were involved in this process. The protein kinase B signaling pathway and B-cell lymphoma-2 expression levels significantly decreased following Livin knockdown, suggesting they may contribute to the regulation of apoptosis and autophagy crosstalk, which caused the Livin knockdown-induced cell death observed.

[Expression of transgelin-2 and clinical significance in colorectal cancer].

OBJECTIVES: To investigate the relationship between the expression of transgelin-2 and the clinicopathological factors of colorectal carcinoma and evaluate the value of transgelin-2 in prognostic assessment of the colorectal cancer patients. METHODS: Using tissue microarray and immunohistochemical methods, we examined transgelin-2 of 120 colorectal cancer patients received surgical treatment from September 2002 to April 2004, including 74 male and 46 female, age from 26 to 89 years. Analyzed the relationship between transgelin-2 and both the clinicopathological features and prognosis of the colorectal cancer by using χ² test and Kaplan-Meier survival analysis. Cox proportion hazard regression analysis was used to study the independent prognostic factors. RESULTS: The positive rate of transgelin-2 expression was 69.2% in colorectal carcinoma. The transgelin-2 expression correlated with differentiation degree (χ² = 5.420), lymph nodes metastasis (χ² = 45.577), distant metastasis (χ² = 12.009), and TNM staging (χ² = 47.577). The survival time was (39 ± 5) months in patients with positive expression of the transgelin-2, while (59 ± 3) months in patients with negative expression. The patient's survival time was statistically correlated with the transgelin-2 expression (P = 0.003). Distant metastasis (RR = 8.318, 95%CI: 4.119 - 16.790), lymph nodes metastasis (RR = 2.794, 95%CI: 1.246 - 6.263) and transgelin-2 expression (RR = 1.834, 95%CI: 1.118- 2.973) were independent prognostic factors in patients with colorectal cancer (P < 0.05). CONCLUSIONS: The expression of transgelin-2 is correlated with clinicopathological features and prognosis in colorectal cancer, may be the potential marker of metastasis and the prognosis of colorectal cancer patients.

[Study on the tumor infiltration in mesorectum of rectal cancer by spiral computed tomography and histopathology].

OBJECTIVE: To evaluate the value of spiral computed tomography in the preoperative assessment of the degree of tumor infiltration in mesorectum and circumferential resection margin status of rectal cancer compared with large tissue slice technique. METHODS: Fifty-seven patients with rectal cancer diagnosed by fiber colonoscopy and pathology from March 2007 to December 2007 underwent preoperative 64-layers spiral CT examination. The degree of tumor infiltration in mesorectum and circumferential resection margin status were evaluated. Large tissue slice technique was applied in the pathologic study after the total mesorectal excision of the rectum to determine the degree of tumor infiltration in mesorectum and the circumferential resection margin status. The spiral CT findings were compared with pathologic results. The accuracy, sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of the CT results were assessed respectively. RESULTS: The overall spiral CT accuracy was 93.0% (53/57) for the degree of tumor infiltration in mesorectum, and it was 94.7%, 94.7% and 96.5% for degree I, II, III infiltration, respectively. Fifty-three cases (93.0%) were accurately predicated with the circumferential resection margin status. The sensitivity, specificity, PPV and NPV of spiral CT measurement was 80.0%, 97.6%, 92.3% and 93.2%, respectively, and was consistent well with the histopathological diagnosis. CONCLUSIONS: Spiral CT provides accurate preoperative assessment for the degree of tumor infiltration in mesorectum and circumferential resection margin status of rectal cancer.

[Prognostic analysis of middle and lower rectal cancer with different degrees of infiltration in mesorectum].

OBJECTIVE: To study the relationship between tumor infiltration in mesorectum and prognosis of middle and lower rectal cancer. METHODS: 49 patients with middle and lower rectal cancer underwent total mesorectal excision. Specimens were obtained during operation and underwent large slice pathologic technique to observe the degree of tumor infiltration in mesorectum and circumferential resection margin. Follow-up was conducted for 61 (9 - 66) months to observe the local recurrence rate, metastasis rate, and five-year survival rate. RESULTS: Follow-up showed a local recurrence rate of 12.2% (6/49), distant metastasis rate of 26.5% (13/49), and five-year survival rate of 67.3% (33/49). The rate of degree I of tumor infiltration in mesorectum was 40.8% (20/49), the degree II rate was 26.5% (13/49), and the degree III rate was 32.7% (16/49) with the corresponding local recurrence rates of 0, 7.7% (1/13), and 31.3% (5/16) respectively (chi(2) = 7.357, P = 0.015), metastatic rates of 10% (2/20), 23.1% (3/13), and 50% (8/16) respectively (chi(2) = 7.405, P = 0.025), and the 5-year survival rates of 90% (18/20), 69.2% (9/13), and 37.5% (6/16) respectively. Kaplan-Meier survival analysis showed that the survival time was correlated with the degree of tumor infiltration in mesorectum (P = 0.012). The rate of circumferential resection margin involvement was 24.5% (12/49). In the 12 patients with positive circumferential resection margin, the local recurrence rate was 33.3% (4/12), whereas 5.4% (2/37) in those with negative circumferential resection margin (chi(2) = 6.577, P = 0.010). Distant metastasis rate was 50% (6/12) in the patients with positive circumferential resection margin, whereas 18.9% (7/37) in those with negative one (chi(2) = 4.491, P = 0.034). The 5-year survival rate of the patients with positive circumferential resection margin was 33.3% (4/12), significantly lower than that of the patients with negative circumferential resection margin [78.4% (29/37)]. Kaplan- Meier survival analysis showed survival time was correlated with the circumferential resection margin status (P = 0.009). CONCLUSION: The degree of tumor infiltration in mesorectum and circumferential resection margin status are important predictors of local and distant recurrence as well as survival of patients with middle and lower rectal carcinoma.

[A clinical study on the degree of mesorectal tumor invasion in middle-low rectal cancer].

OBJECTIVE: To study the relationship between the degree of mesorectal tumor invasion and prognosis of the patients with middle-low rectal cancer. METHODS: Specimens from 49 patients with middle-low rectal cancer, undergone total mesorectal excision in our hospital from April 2003 to December 2003,were studied by large slice pathologic technique. The thickness of mesorectum and the depth of tumor infiltration were measured, and the degree of mesorectal tumor invasion was calculated. The local recurrence rate, metastasis rate and 5-year survival rate were investigated respectively. Possible clinicopathological influence factors were also analyzed. RESULTS: The local recurrence rate was 12.2%(6/49), the distant metastasis rate was 26.5%(13/49). In three different degrees of mesorectal tumor invasion(I(,II(,III(), the local recurrence rates were 0, 7.7% and 31.3% (chi(2)=7.357, P =0.015); the metastasis rates were 10%, 23.1% and 50%(chi(2)=7.405, P =0.025); the 5-year survival rates were 90.9%, 69.2% and 28.6%(log-rank, P =0.013). Tumor diameter, T and N staging were risk factors influencing the degree of mesorectal tumor invasion(chi(2)=6.849 P=0.033, chi(2)=34.845 P =0.000, chi(2)=17.266 P =0.002). CONCLUSIONS: The degree of mesorectal tumor invasion is an important predictor of local and distant metastasis as well as survival of patients in middle-low rectal carcinoma. The degree of mesorectal tumor invasion in the middle-low rectal carcinoma is significantly correlated with tumor diameter, T and N stage.