MCCC2 is a novel mediator between mitochondria and telomere and functions as an oncogene in colorectal cancer.

BACKGROUND: The mitochondrial gene MCCC2, a subunit of the heterodimer of 3-methylcrotonyl-CoA carboxylase, plays a pivotal role in catabolism of leucine and isovaleric acid. The molecular mechanisms and prognostic value still need to be explored in the context of specific cancers, including colorectal cancer (CRC). METHODS: In vitro and in vivo cell-based assays were performed to explore the role of MCCC2 in CRC cell proliferation, invasion, and migration. Mitochondrial morphology, membrane potential, intracellular reactive oxygen species (ROS), telomerase activity, and telomere length were examined and analyzed accordingly. Protein complex formation was detected by co-immunoprecipitation (CO-IP). Mitochondrial morphology was observed by transmission electron microscopy (TEM). The Cancer Genome Atlas (TCGA) CRC cohort analysis, qRT-PCR, and immunohistochemistry (IHC) were used to examine the MCCC2 expression level. The association between MCCC2 expression and various clinical characteristics was analyzed by chi-square tests. CRC patients' overall survival (OS) was analyzed by Kaplan-Meier analysis. RESULTS: Ectopic overexpression of MCCC2 promoted cell proliferation, invasion, and migration, while MCCC2 knockdown (KD) or knockout (KO) inhibited cell proliferation, invasion, and migration. MCCC2 KD or KO resulted in reduced mitochondria numbers, but did not affect the gross ATP production in the cells. Mitochondrial fusion markers MFN1, MFN2, and OPA1 were all upregulated in MCCC2 KD or KO cells, which is in line with a phenomenon of more prominent mitochondrial fusion. Interestingly, telomere lengths of MCCC2 KD or KO cells were reduced more than control cells. Furthermore, we found that MCCC2 could specifically form a complex with telomere binding protein TRF2, and MCCC2 KD or KO did not affect the expression or activity of telomerase reverse transcriptase (TERT). Finally, MCCC2 expression was heightened in CRC, and patients with higher MCCC2 expression had favorable prognosis. CONCLUSIONS: Together, we identified MCCC2 as a novel mediator between mitochondria and telomeres, and provided an additional biomarker for CRC stratification.

Aquaporin 9 is involved in CRC metastasis through DVL2-dependent Wnt/ß-catenin signaling activation.

Background: Aquaporin 9 (AQP9) is permeable to water or other small molecules, and plays an important role in various cancers. We previously found that AQP9 was related to the efficacy of chemotherapy in patients with colorectal cancer (CRC). This study aimed to identify the role and regulatory mechanism of AQP9 in CRC metastasis. Methods: The clinical significance of AQP9 was analysed by using bioinformatics and tissue microarray. Transcriptome sequencing, Dual-Luciferase Reporter Assay, Biacore, and co-immunoprecipitation were employed to demonstrate the regulatory mechanism of AQP9 in CRC. The relationship between AQP9 and CRC metastasis was verified in vitro and in vivo by using real-time cell analysis assay, high content screening, and liver metastasis models of nude mice. Results: We found that AQP9 was highly expressed in metastatic CRC. AQP9 overexpression reduced cell roundness and enhanced cell motility in CRC. We further showed that AQP9 interacted with Dishevelled 2 (DVL2) via the C-terminal SVIM motif, resulting in DVL2 stabilization and the Wnt/ß-catenin pathway activation. Additionally, we identified the E3 ligase neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) as a modulator regulating the ubiquitination and degradation of AQP9. Conclusions: Collectively, our study revealed the important role of AQP9 in regulating DVL2 stabilization and Wnt/ß-catenin signaling to promote CRC metastasis. Targeting the NEDD4L-AQP9-DVL2 axis might have therapeutic usefulness in metastatic CRC treatment.

CXCL16 inhibits epithelial regeneration and promotes fibrosis during the progression of radiation enteritis.

Radiation enteritis (RE) is a prevalent complication of radiotherapy for pelvic malignant tumors, characterized by severe intestinal epithelial destruction and progressive submucosal fibrosis. However, little is known about the pathogenesis of this disease, and so far, there is no specific targeted therapy. Here, we report that CXCL16 is upregulated in the injured intestinal tissues of RE patients and in a mouse model. Genetic deletion of Cxcl16 mitigates fibrosis and promotes intestinal stem cell-mediated epithelial regeneration after radiation injury in mice. Mechanistically, CXCL16 functions on myofibroblasts through its receptor CXCR6 and activates JAK3/STAT3 signaling to promote fibrosis and, at the same time, to transcriptionally modulate the levels of BMP4 and hepatocyte growth factor (HGF) in myofibroblasts. Moreover, we find that CXCL16 and CXCR6 auto- and cross-regulate themselves in positive feedback loops. Treatment with CXCL16 neutralizing monoclonal antibody attenuates fibrosis and improves the epithelial repair in RE mouse model. Our findings emphasize the important role of CXCL16 in the progression of RE and suggest that CXCL16 signaling could be a potential therapeutic target for RE. © 2022 The Pathological Society of Great Britain and Ireland.

Epigenetic silencing and tumor suppressor gene of HAND2 by targeting ERK signaling in colorectal cancer.

BACKGROUND: The screening biomarkers for early detection of colorectal cancer (CRC) is lacking. The aim is to identify epigenetic silenced genes and clarify its roles and underlying mechanism in CRC. We conducted integrative analyses of epigenome-wide Human Methylation 450 K arrays and transcriptome to screen out candidate epigenetic driver genes with transcription silencing. Methylated silencing HAND2 were identified and verified in large CRC cohort. The mechanism of HAND2 expression by promoter inhibition were clarified both in vitro and vivo assays. Cell biofunctional roles of HAND2 methylation was investigated in CRC cells. HAND2 reconstitution were constructed by lentivirus plasmid and tumor xenograft model of HAND2 were built subcutaneously. Genomic mRNA analysis by RNA-sequencing and subsequent GSEA analysis were performed to identify potential target of HAND2 and qPCR/WB was conducted to identify the results. RESULTS: We firstly reported high frequency of HAND2 methylation in promoter in CRC and hypermethylation was negatively correlated with expression silencing and leaded to poor survival in several CRC cohort patients. 5-Aza treatment to demethylated HAND2 could revert its expression in CRC cells. Functionally, HAND2 reconstitution can inhibit cell proliferation, invasion and migration in vitro. In tumor xenograft, HAND2 reconstruction significantly repressed tumor growth when compared to control vector. Thousands of aberrant expressed genes were observed in the heatmap of RNA-sequencing data. HAND2 reconstitution could bind to ERK and reduce its phosphorylation by CoIP assay. These above results showed HAND2 reconstitution perturbed the activation of MAPK/ERK signaling by reduction of ERK phosphorylation. CONCLUSIONS: HAND2 is one tumor suppressor by targeting ERK signaling and one potential epigenetic driver gene in CRC. Video Abstract.

Laparoscopic Proximally Extended Colorectal Resection With Two-Stage Turnbull-Cutait Pull-Through Coloanal Anastomosis for Late Complications of Chronic Radiation Proctopathy.

Background: Chronic radiation proctopathy (CRP) is a common complication after radiation therapy for pelvic malignancies. Compared with diversion surgery, resection surgery removes the damaged tissue completely to avoid the risks of recurrence and improve patients' outcome. Hence, resection surgery could be an optimal surgical approach when CRP is complicated by late complications. This study aimed to describe a modified surgical procedure of resection surgery and report its preliminary efficacy and safety in treating patients with CRP with late complications. Methods: We retrospectively reviewed the patients who were diagnosed with CRP with late complications and underwent the modified surgical procedure of laparoscopic proximally extended colorectal resection with two-Stage Turnbull-Cutait pull-through coloanal anastomosis (PE-Bacon) between November 2019 and October 2020 in the Sixth Affiliated Hospital of Sun Yat-sen University. Results: A total of 15 patients were performed the modified laparoscopic procedure of PE-Bacon, of which 1 patient underwent conversion from laparoscopic to open operation for intraoperative massive hemorrhage. Overall, the major (Clavien-Dindo III-V) postoperative complications occurred in 1 patient, anastomotic leakage was observed in 2 (13.3%) patients, and anastomotic stricture was observed in 4 (26.7%) patients. No patient had to be reoperated and died. Up to now, at the average follow-up of (524.40 ± 108.39) days, the preoperative symptoms of 93.3% (14/15) patients were relieved, with nine patients achieved complete remission, five patients only suffered minor symptoms. Because of the progression of radiation uropathy, one patient still had a vesicovaginal fistula as pre-operative complication. Colostomy reversal has been performed on 8 (53.3%) patients at an average postoperative duration of 299.5 ± 92.68 days, among whom only 2 patients suffered from major Low Anterior Resection Syndrome (LARS) until now. Conclusions: Laparoscopic PE-Bacon surgery is a safe and feasible surgical procedure for late complications of CRP with low morbidity and high symptom remission rate.

TSG-6 promotes Cancer Cell aggressiveness in a CD44-Dependent Manner and Reprograms Normal Fibroblasts to create a Pro-metastatic Microenvironment in Colorectal Cancer.

Tumor necrosis factor α stimulated gene 6 (TSG-6), a 30-KD secretory protein, plays an essential role in modulating inflammatory responses and extracellular matrix remodeling. However, little is known regarding the role of TSG-6 in human cancers. Here, we investigated the mechanism of action and the role of TSG-6 in colorectal cancer (CRC) metastasis. We found that TSG-6 was highly expressed in tumor tissues and was associated with poor prognosis and metastasis in CRC. Mechanistically, TSG-6 overexpression in CRC cells resulted in ERK activation and epithelial-mesenchymal transition by means of stabilizing CD44 and facilitating the CD44-EGFR complex formation on the cell membrane. Consequently, this resulted in the promotion of tumor migration and invasion both in vitro and in vivo. Notably, our data showed that CRC cells secreted TSG-6 could trigger a paracrine activation of JAK2-STAT3 signaling and reprogram normal fibroblasts into cancer-associated fibroblasts, which exhibited upregulation of pro-metastatic cytokines (CCL5 and MMP3) and higher movement ability. In animal models, the co-injection of cancer cells and TSG6-reprogrammed fibroblasts led to a significant increase in tumor metastasis. Our findings indicated that TSG-6 overexpression in CRC cells could promote cancer metastasis in both an autocrine and paracrine manner. Therefore, targeting TSG-6 might be a potential therapeutic strategy for the treatment of metastatic CRC.

Metabolomics reveals that CAF-derived lipids promote colorectal cancer peritoneal metastasis by enhancing membrane fluidity.

Patients with peritoneal metastasis (PM) of colorectal cancer (CRC) have poorer overall survival outcomes than those without PM. Cancer-associated fibroblasts (CAFs) are a major component of the tumor microenvironment and mediate CRC progression and PM. It is imperative to identify and develop novel therapeutic targets for PM-CRC driven by CAFs. Using lipidomics, we reveal that the abundance of phosphatidylcholine (PC) with unsaturated acyl chains was increased in clinical PM-CRC specimens. Additionally, we found that CAFs were present at a higher relative abundance in primary PM-CRC tumors and that membrane fluidity in CRC cells was increased after incubation with CAF-conditioned medium (CM) through three independent methods: lipidomics, fluorescence recovery after photobleaching (FRAP), and generalized polarization. Then, we found that increased membrane fluidity can enhance glucose uptake and metabolism, as supported by real-time bioenergetics analysis and U-13C glucose labeling. Interestingly, stearoyl-CoA desaturase 1 (SCD), the rate-limiting enzyme in the biosynthesis of unsaturated fatty acids (uS-FAs), was expressed at low levels in PM and associated with poor prognosis in CRC patients. Importantly, by untargeted metabolomics analysis and fatty acid ([U-13C]-stearic acid) tracing analyses, we found that CRC cells take up lipids and lipid-like metabolites secreted from CAFs, which may compensate for low SCD expression. Both in vitro and in vivo experiments demonstrated that sodium palmitate (C16:0) treatment could decrease the CAF-induced change in cell membrane fluidity, limit glucose metabolism, suppress cell invasiveness, and impair tumor growth and intraperitoneal dissemination. An increased C16:0 concentration was shown to induce apoptosis linked to lipotoxicity. Furthermore, C16:0 effectively enhanced the antitumor activity of 5-fluorouracil (5-FU) in vitro and was well tolerated in vivo. Taken together, these findings suggest that adding the saturated fatty acid (S-FA) C16:0 to neoadjuvant chemotherapy may open new opportunities for treating PM-CRC in the future.

A PDX model combined with CD-DST assay to evaluate the antitumor properties of KRpep-2d and oxaliplatin in KRAS (G12D) mutant colorectal cancer.

Patient-derived xenograft (PDX) models are more faithful in maintaining the characteristics of human tumors than cell lines and are widely used in drug development, although they have some disadvantages, including their relative low success rate, long turn-around time, and high costs. The collagen gel droplet embedded culture drug sensitivity test (CD-DST) has been used as an in-vitro drug sensitivity test for patients with cancer because of its high success rate of primary cell culture, high sensitivity, and good clinical relevance, but it is based on an in-vitro cell culture and may not simulate the tumor microenvironment accurately. This study aims to combine a PDX model with CD-DST to evaluate the efficiency of antitumor agents. KRpep-2d, a small peptide targeting KRAS (G12D), and oxaliplatin were used to verify the feasibility of this approach. Whole-exome sequencing and Sanger sequencing were first applied to test and validate the KRAS mutation status of a panel of colorectal cancer PDX tissues. One PDX model was verified to carry KRAS (G12D) mutation and was used for in-vivo and the CD-DST drug tests. We then established the PDX mouse model from the patient with the KRAS (G12D) mutation and obtained viable cancer cells derived from the same PDX model. Next, the antitumor abilities of KRpep-2d and oxaliplatin were estimated in the PDX model and the CD-DST. We found that KRpep-2d showed no significant antitumor effect on the xenograft model or on cancer cells derived from the same PDX model. In contrast, oxaliplatin showed significant inhibitory effects in both tests. In conclusion, the PDX model in combination with the CD-DST assay is a comprehensive and feasible method of evaluating the antitumor properties of compounds and could be applied for new drug discovery.

The predictive value of high-sensitive troponin I for perioperative risk in patients undergoing gastrointestinal tumor surgery.

BACKGROUND: The incidence of cardiovascular events in perioperative period of gastrointestinal tumor surgery cannot be ignored, and studies have shown that level of postoperative troponin is related to the postoperative risk of non-cardiac surgery. However, the relationship between pre-operative troponin levels and perioperative risk of gastrointestinal tumor surgery is unclear. Thus, we aimed to evaluate the value of high-sensitive cardiac troponin I (hs-cTnI) prior to gastrointestinal tumor surgery for perioperative risk assessment. METHODS: In this retrospective cohort study, 1259 patients who underwent gastrointestinal tumor surgery and had been tested for hs-cTnI on admission within 7 days prior to surgery were retrospectively recruited from January 2018 to June 2020. The primary combined endpoint including in-hospital all-cause mortality, acute myocardial infarction, cardiac arrest or ventricular fibrillation and acute decompensated heart failure. The secondary endpoint included total hospital stay and requirement of intensive care treatment. FINDINGS: Compared with patients with normal hs-cTnI, those with elevated hs-cTnI (> 0·028 ng/ml) were more likely to experience the combined endpoint (28·2% versus 2·7%, P < 0·001) and there was also an increasing rate of in mortality in elevated hs-cTnI group (2·4% versus 0·3%, P = 0·057). The length of total hospital stay was significantly longer in patients with elevated hs-cTnI (24·8 ± 16·3 versus 19·5 ± 7·9, P = 0·003) and the number of patients requiring intensive care treatment was also higher (22·6% versus 4·2%, P < 0·001). The area under the ROC curve assessing hs-cTnI in predicting in-hospital mortality was 0·787 [95% confidence interval (CI) 0·612-0·963, P = 0·015] and for combined endpoint was 0·822 [95% CI 0·766-0·879, P < 0·001]. Hs-cTnI > 0·028 ng/ml was associated with significantly higher cardiovascular event rate in patients with the revised cardiac index ≤ 1. The positive likelihood ratio of hs-cTnI (> 0·028 ng/ml) for predicting combined endpoint reaches 10.5 in patients with Lee index = 0. In multivariate logistic analyses, hs-cTnI was one of the best predictors for the combined endpoint [odds ratio (OR) 5·924 (95%CI: 2·869-12·233), P < 0·001]. INTERPRETATION: Hs-cTnI provides powerful prognostic information for patients undergoing gastrointestinal tumor surgery, and therefore provides reliable prognostic information incremental to revised cardiac index.

ORMDL1 is upregulated and associated with favorable outcomes in colorectal cancer.

BACKGROUND: The ORMDL1 gene is known as a crucial negative regulator of sphingolipid biogenesis. However, the ORMDL1 gene has rarely been studied in a tumor-related context. Therefore, its prognostic value and functional significance in colorectal cancer (CRC) remain to be explored. METHODS: TCGA CRC cohort analysis, qRT-PCR, and immunohistochemistry (IHC) were used to examine the ORMDL1 expression level. The association between ORMDL1 expression and various clinical characteristics was analyzed by chi-square tests. The overall survival (OS) of CRC patients was analyzed by Kaplan-Meier analysis. In vitro and in vivo cell-based assays were performed to explore the role of ORMDL1 in cell proliferation, invasion and migration. Transcriptional changes in cells with either ORMDL1 knockdown or overexpression were compared and analyzed. RESULTS: ORMDL1 was upregulated in CRC tissues in both the TCGA and our cohort. Interestingly, its expression was significantly lower in patients with metastasis than in patients without metastasis, and the high expression group had longer OS than the low expression group. Knockdown of ORMDL1 expression can promote proliferation, colony formation and invasion, while attenuating migration in CRC cell lines. In contrast, forced overexpression of ORMDL1 reduced cell proliferation, colony formation and invasion, while enhancing cell migration. Stable knockdown of ORMDL1 can promote cancer cell proliferation in vivo to some extent. Finally, Rho GTPase activity was influenced by ORMDL1, and the expression of ORMDL1 was enhanced by DTT treatment. CONCLUSION: ORMDL1 is upregulated and may serve as a biomarker to predict favourable outcomes in colorectal cancer.

Enhancing cancer-associated fibroblast fatty acid catabolism within a metabolically challenging tumor microenvironment drives colon cancer peritoneal metastasis.

Most cancer-related deaths result from the progressive growth of metastases. Patients with peritoneal metastatic (PM) colorectal cancer have reduced overall survival. Currently, it is still unclear why colorectal cancer (CRC) cells home to and proliferate inside the peritoneal cavity, and there is no effective consolidation therapy for improved survival. Using a proteomic approach, we found that key enzymes of fatty acid oxidation (FAO) were decreased in patients with PM colorectal cancer. Furthermore, we confirmed that carnitine palmitoyltransferase IA (CPT1A), a rate-limiting enzyme of FAO, was expressed at significantly low levels in patients with PM colorectal cancer, as determined by RT-qPCR, IHC, and GEO dataset analysis. However, lipidomics revealed no difference in FFA levels between PM and non-PM primary tumors. Here, we showed that cancer-associated fibroblasts (CAFs) promote the proliferation, migration, and invasion of colon cancer cells via upregulating CPT1A to actively oxidize FAs and conduct minimal glycolysis. In addition, coculture-induced glycolysis increased in cancer cells while fatty acid catabolism decreased with lower adiponectin levels. Importantly, inhibition of glycolysis significantly reduced the survival of CRC cells after incubation with conditioned medium from CAFsCPT1A-OE in vitro and impaired the survival and growth of organoids derived from CRC-PM. Finally, we found that directly blocking FAO in CAFsCPT1A-OE with etomoxir inhibits migration and invasion in vitro and decreases tumor growth and intraperitoneal dissemination in vivo, revealing a role for CAF CPT1A in promoting tumor growth and invasion. In conclusion, our results suggest the possibility of testing FAO inhibition as a novel approach and clinical strategy against CAF-induced colorectal cancer with peritoneal dissemination/metastases.

EGF Relays Signals to COP1 and Facilitates FOXO4 Degradation to Promote Tumorigenesis.

Forkhead-Box Class O 4 (FOXO4) is involved in critical biological functions, but its response to EGF-PKB/Akt signal regulation is not well characterized. Here, it is reported that FOXO4 levels are downregulated in response to EGF treatment, with concurrent elevation of COP9 Signalosome subunit 6 (CSN6) and E3 ubiquitin ligase constitutive photomorphogenic 1 (COP1) levels. Mechanistic studies show that CSN6 binds and regulates FOXO4 stability through enhancing the E3 ligase activity of COP1, and that COP1 directly interacts with FOXO4 through a VP motif on FOXO4 and accelerates the ubiquitin-mediated degradation of FOXO4. Metabolomic studies demonstrate that CSN6 expression leads to serine and glycine production. It is shown that FOXO4 directly binds and suppresses the promoters of serine-glycine-one-carbon (SGOC) pathway genes, thereby diminishing SGOC metabolism. Evidence shows that CSN6 can regulate FOXO4-mediated SGOC gene expression. Thus, these data suggest a link of CSN6-FOXO4 axis and ser/gly metabolism. Further, it is shown that CSN6-COP1-FOXO4 axis is deregulated in cancer and that the protein expression levels of CSN6 and FOXO4 can serve as prognostic markers for cancers. The results illustrate a pathway regulation of FOXO4-mediated serine/glycine metabolism through the function of CSN6-COP1 axis. Insights into this pathway may be strategically designed for therapeutic intervention in cancers.

KRAS and PIK3CA bi-mutations predict a poor prognosis in colorectal cancer patients: A single-site report.

STUDY RATIONALE: The coexistence of KRAS and PIK3CA mutations in cells implies potential synergistic hyperactivation of the Ras/MAPK and PI3K/Akt oncogenic pathways. Therefore, it is desirable to investigate the concomitant mutations of KRAS and PIK3CA in colorectal cancer (CRC) samples and whether the concomitant mutations are associated with a poor prognosis in CRC patients. AIM: To investigate the clinicpathological characteristics and prognostic value of concomitant mutations of KRAS and PIK3CA in CRC samples. METHODS: In this study, a total of 655 CRC patients from the Sixth Affiliated Hospital of Sun Yat-sen University were enrolled from January to December 2015. Sanger sequencing was applied to survey the mutational status of hotspot regions in the open reading frames (ORFs) of the KRAS and PIK3CA genes. Clinicpathological parameters were collected and analyzed. The Kaplan-Meier method and Cox regression model were applied to determine the correlation between the KRAS and PIK3CA mutation statuses and survival. RESULTS: We found that KRAS and PIK3CA bi-mutations were significantly associated with aggressive clinicpathological features. Among the studied CRC patients, those with either KRAS mutations (P = 0.004) or KRAS and PIK3CA bi-mutations (P = 0.033) had poor overall survival (OS). In the multivariable analysis, KRAS mutations in exons 3 and 4 but not exon 2 with concomitant PIK3CA mutations were associated with a high risk of death (univariate HR = 8.05; 95% CI, 1.926-33.64, P = 0.004; multivariate HR = 10.505; 95% CI, 2.304-47.905, P = 0.002). CONCLUSION: The concomitant mutation statuses of KRAS and PIK3CA should be considered when the prognostic value of gene mutations is consulted in CRC patients.

To what extent should the intestinal be resected proximally after radiotherapy: hint from a pathological view.

BACKGROUND: Neoadjuvant chemoradiotherapy (nCRT) is associated with post-operative anastomotic complications in rectal-cancer patients. Anastomosis involving at least one non-irradiated margin reportedly significantly reduces the risk of post-operative anastomotic complications in radiation enteritis. However, the exact scope of radiotherapy on the remaining sigmoid colon remains unknown. METHODS: We evaluated the radiation damage of proximally resected colorectal segments in 44 patients with rectal cancer, who received nCRT followed by conventional resection (nCRT-C, n = 21) or proximally extended resection (nCRT-E, n = 23). The segments from another 13 patients undergoing neoadjuvant chemotherapy (nCT) were used as control. We dissected these samples at a distance of 2 cm between the two adjacent sections. Radiation damage in proximally resected colorectal segments was evaluated using the radiation injury score (RIS) and the concentration and distribution patterns of angiostatin. RESULTS: Compared to those in the nCT group, the nCRT group showed higher RIS, levels of angiostatin, and proportion of diffuse pattern of angiostatin. With increasing distance from the tumor site, these parameters all gradually decreased; and the differences came to be not significant at the site that is over 20 cm from the tumor. The nCRT-E group showed lower RIS (median: 2 vs 4, P = 0.002) and a greater proportion of non-diffuse angiostatin (87% vs 55%, P = 0.039) at the proximal margins compared with the nCRT-C group. CONCLUSIONS: The severity of the radiation damage of the proximal colon is inversely proportional to the proximal-resection margin length. Little damage was left on the proximal margin that was over 20 cm from the tumor. Removal of an initial length of ≥20 cm from the tumor may be beneficial for rectal-cancer patients after nCRT.

Ubiquitin ligase TRIM65 promotes colorectal cancer metastasis by targeting ARHGAP35 for protein degradation.

Tripartite motif-containing protein 65 (TRIM65) is an E3 ubiquitin ligase and a critical regulator of a variety of cellular processes as well as tumor progression. Therefore, more substrates must be identified in the physiology or disease context. Here, we found that TRIM65 is upregulated and associated with poor survival in colorectal cancer (CRC). More specifically, high expression of TRIM65 is associated with CRC metastasis and recurrence. Ectopic overexpression of TRIM65 in CRC cell lines enhanced proliferation, invasion, and migration, while knockdown of TRIM65 expression had the opposite effects. Furthermore, we identified a new substrate of TRIM65, namely ARHGAP35, a Rho GTPase-activating protein (GAP) that is involved in polarized cell migration. Phenotypically, forced expression of TRIM65 induces increased production of migration-related structures, focal adhesions, and/or filopodia and enhances CRC metastasis to the liver or the lung in a mouse model. Mechanistic studies revealed that TRIM65 mediates ubiquitination of ARHGAP35, whose degradation leads to elevated Rho GTPase activity. In addition, we identified several phosphorylation sites on TRIM65. In sum, we reveal a novel TRIM65-GAP-Rho regulatory axis that modulates the actin cytoskeleton and the migration behavior of CRC cells, and the TRIM65-ARHGAP35 interaction might be a valuable therapeutic target in CRC.

Efficacy and complications of argon plasma coagulation for hemorrhagic chronic radiation proctitis.

BACKGROUND: Chronic radiation proctitis (CRP) is a complication which occurs in 1%-5% of patients who undergo radiotherapy for pelvic malignancies. Although a wide range of therapeutic modalities are available, there is no literature to date showing any particularly appropriate therapeutic modality for each disease stage. Argon plasma coagulation (APC) is currently recommended as the first-choice treatment for hemorrhagic CRP, however, its indication based on long-term follow-up is still unclear. On the hypothesis that the long-term efficacy and safety of APC are not fully understood, we reviewed APC treatment for patients with hemorrhagic CRP from a single center. AIM: To assess the long-term efficacy and safety of APC for hemorrhagic CRP. METHODS: This is a retrospective study of consecutive patients treated with APC for hemorrhagic CRP from January 2013 to October 2017. Demographics, clinical variables, and typical endoscopic features were recorded independently. Success was defined as either cessation of bleeding or only occasional traces of bloody stools with no further treatments for at least 12 mo after the last APC treatment. We performed univariate and multivariate analyses to identify factors associated with success and risk factors for fistulas. RESULTS: Forty-five patients with a median follow-up period of 24 mo (range: 12-67 mo) were enrolled. Fifteen (33.3%) patients required blood transfusion before APC. Successful treatment with APC was achieved in 31 (68.9%) patients. The mean number of APC sessions was 1.3 (1-3). Multivariate analysis showed that APC failure was independently associated with telangiectasias present on more than 50% of the surface area [odds ratio (OR) = 6.53, 95% confidence interval (CI): 1.09-39.19, P = 0.04] and ulcerated area greater than 1 cm2 (OR = 8.15, 95%CI: 1.63-40.88, P = 0.01). Six (13.3%) patients had severe complications involving rectal fistulation. The only factor significantly associated with severe complications was ulcerated area greater than 1 cm2 (P = 0.035). CONCLUSION: The long-term efficacy of APC for hemorrhagic CRP is uncertain in patients with telangiectasias present on > 50% of the surface area and ulceration > 1 cm2.

The novel long noncoding RNA u50535 promotes colorectal cancer growth and metastasis by regulating CCL20.

Long noncoding RNAs (lncRNAs) have been emerging as master regulators of tumor growth and metastasis, but the functions and underlying mechanisms of lncRNAs in colorectal cancer (CRC) still need to be clarified. Here, we found a novel lncRNA u50535, which was greatly overexpressed in CRC tissues and was associated with poor prognosis in CRC patients. Function studies showed that u50535 was an oncogene in CRC both in vitro and in vivo. In mechanism, through RNA sequencing and rescue assay, we found that u50535 activates CCL20 signaling to promote cell proliferation and migration in CRC. Taken together, these findings suggest that u50535 can promote CRC growth and metastasis and may serve as a potential biomarker in CRC.

Downregulation of TRIM58 expression is associated with a poor patient outcome and enhances colorectal cancer cell invasion.

TRIM58 is a member of the tripartite motif protein (TRIM) family of E3 ubiquitin ligases. Aberrant gene methylation of TRIM58 has been reported in liver and lung cancer and indicates a poor patient prognosis. However, the expression level and functional role of TRIM58 in colorectal cancer (CRC) have yet to be elucidated. In the present study, we found that TRIM58 expression was significantly suppressed in human CRC and was inversely correlated with CRC progression. Additionally, overall survival was significantly reduced in patients with low TRIM58 expression in CRC tumors. In vitro studies demonstrated that ectopic TRIM58 overexpression strongly inhibited CRC cell invasion but had minimal effects on cell proliferation, colonization and migration. Furthermore, TRIM58 suppression enhanced the expression of epithelial-to-mesenchymal transition (EMT) and matrix metalloproteinase (MMP) genes. Thus, our findings suggest that TRIM58 is a potential prognostic marker of CRC and functions as a tumor-suppressor gene via inhibition of cancer cell invasion through EMT and MMP activation.

Role of Angiogenesis in Chronic Radiation Proctitis: New Evidence Favoring Inhibition of Angiogenesis Ex Vivo.

BACKGROUND: Chronic radiation proctitis (CRP), a common complication after radiotherapy for pelvic malignancies, compromises patient quality of life. Vascular damage and aberrant angiogenesis in the mucosal layer are essential histological features, but changes to the submucosal layer are unclear. Thus, we evaluated the histological characteristics and distribution changes of key angiogenic factors in full-layered human CRP samples. METHODS: Thirty paraffin-embedded CRP and twenty-nine non-CRP tissues were used to evaluate histopathological changes. Immunohistochemistry with anti-CD34 antibody was performed to calculate microvascular density (MVD). Frozen tissues from eight CRP patients and five non-CRP controls were collected and analyzed by antibody array, which contained sixty human angiogenesis-related factors. Quality controls with positive and negative controls were performed during antibody array analysis. Two differentially expressed factors were confirmed by ELISA. RESULTS: CRP lesions showed vasculopathy, fibrosis, mucosal ulceration, edema, and inflammatory cell infiltration. Human angiogenesis antibody array and ELISA confirmed the increased angiostatin in CRP lesions. Immunohistochemical staining showed dispersed distribution of angiostatin throughout the mucosal and submucosal layers in CRP lesions, while angiostatin accumulated within the vessel lumens in non-CRP tissues. MVD significantly decreased in the submucosal layer of CRP, suggesting a potential association with increased angiostatin. CONCLUSIONS: Angiostatin increased and had a distinct distribution in CRP lesions. Compensatory telangiectasia in the mucosa, vessel stenosis, and reduced MVD might attenuate blood flow in the submucosa and contribute to CRP progression. Restoration of vascular functionality by promoting angiogenesis in the submucosal layer may help alleviate CRP in clinical practice.