Inhibited Colorectal Cancer Growth Research Articles

Intestinal metabolite xylulose inhibits colorectal cancer by inducing apoptosis through the MAPK signalling pathway

BackgroundThe intestinal metabolites are involved in the initiation, progression and metastasis of colorectal cancer (CRC). They are a potential source of agents for cancer therapy. Our previous study identified altered faecal metabolites between CRC patients and healthy volunteers. However, no specific metabolite was clearly illustrated for CRC therapy. ResultsWe found that the level of xylulose was lower in the stools of CRC patients than in those of healthy volunteers. Xylulose inhibited cell growth without affecting the cell cycle by inducing apoptosis in CRC cells, which was evidenced by increased expression of the proapoptotic proteins C-PARP and C-Caspase3 and decreased expression of the antiapoptotic protein BCL-2 in CRC cells. Mechanistically, xylulose reduced the activity of the MAPK signalling pathway, represented by reduced phosphorylation of JNK, ERK, and P38. Furthermore, an ALI model was used to show the tumour killing ability of xylulose on human CRC spheres, as well as human colorectal adenoma (AD) spheres. ConclusionXylulose inhibits CRC growth by inducing apoptosis through attenuation of the MAPK signalling pathway. These results suggest that xylulose may serve as an effective agent for CRC therapy.

Combination of miR159 Mimics and Irinotecan Utilizing Lipid Nanoparticles for Enhanced Treatment of Colorectal Cancer.

Colorectal cancer (CRC) ranks as the third most prevalent global malignancy, marked by significant metastasis and post-surgical recurrence, posing formidable challenges to treatment efficacy. The integration of oligonucleotides with chemotherapeutic drugs emerges as a promising strategy for synergistic CRC therapy. The nanoformulation, lipid nanoparticle (LNP), presents the capability to achieve co-delivery of oligonucleotides and chemotherapeutic drugs for cancer therapy. In this study, we constructed lipid nanoparticles, termed as LNP-I-V by microfluidics to co-deliver oligonucleotides miR159 mimics (VDX05001SI) and irinotecan (IRT), demonstrating effective treatment of CRC both in vitro and in vivo. The LNP-I-V exhibited a particle size of 118.67 ± 1.27 nm, ensuring excellent stability and targeting delivery to tumor tissues, where it was internalized and escaped from the endosome with a pH-sensitive profile. Ultimately, LNP-I-V significantly inhibited CRC growth, extended the survival of tumor-bearing mice, and displayed favorable safety profiles. Thus, LNP-I-V held promise as an innovative platform to combine gene therapy and chemotherapy for improving CRC treatment.

Cooperation of MLL1 and Jun in controlling H3K4me3 on enhancers in colorectal cancer

BackgroundEnhancer dysregulation is one of the important features for cancer cells. Enhancers enriched with H3K4me3 have been implicated to play important roles in cancer. However, their detailed features and regulatory mechanisms have not been well characterized.ResultsHere, we profile the landscape of H3K4me3-enriched enhancers (m3Es) in 43 pairs of colorectal cancer (CRC) samples. M3Es are widely distributed in CRC and averagely possess around 10% of total active enhancers. We identify 1322 gain variant m3Es and 367 lost variant m3Es in CRC. The target genes of the gain m3Es are enriched in immune response pathways. We experimentally prove that repression of CBX8 and RPS6KA5 m3Es inhibits target gene expression in CRC. Furthermore, we find histone methyltransferase MLL1 is responsible for depositing H3K4me3 on the identified Vm3Es. We demonstrate that the transcription factor AP1/JUN interacts with MLL1 and regulates m3E activity. Application of a small chemical inhibitor for MLL1 activity, OICR-9429, represses target gene expression of the identified Vm3Es, enhances anti-tumor immunity and inhibits CRC growth in an animal model.ConclusionsTaken together, our study illustrates the genome-wide landscape and the regulatory mechanisms of m3Es in CRC, and reveals potential novel strategies for cancer treatment.

Chidamide and Oxaliplatin Synergistically Inhibit Colorectal Cancer Growth by Regulating the RPS27A-MDM2-P53 Axis.

The present study explored the anti-tumor effects of chidamide plus oxaliplatin on colorectal cancer (CRC) and examined its underlying mechanism. First, the Combination Index (CI) of chidamide and oxaliplatin was evaluated via CCK-8 assay. Second, the effects of chidamide and oxaliplatin monotherapy and the combined treatment on cell proliferation, invasion, migration, and apoptosis were detected. Third, whole-transcriptome RNA sequencing (RNA-seq) was performed to seek the potential targeted gene by which chidamide plus oxaliplatin exerted anti-tumor effects. Fourth, the validation of the targeted gene and the signal pathway it regulated were performed. Finally, the anti-tumor effect of chidamide plus oxaliplatin on mice xenograft was examined. Chidamide and oxaliplatin acted synergistically to inhibit CRC growth in vitro and in vivo (CI<1). Besides, compared with oxaliplatin monotherapy, chidamide could significantly enhance oxaliplatin-induced inhibition in cell proliferation, invasion, and migration, and promotion in HCT-116 and RKO cell apoptosis (P<0.05). The RNA-seq displayed that, compared to oxaliplatin monotherapy, RPS27A mRNA was evidently decreased in HCT-116 cells treated with chidamide plus oxaliplatin (P<0.001). Then, we found RPS27A was highly expressed in CRC tissues and CRC cell lines (P<0.001). Silence of RPS27A attenuated proliferation and induced apoptosis in HCT-116 and RKO cells via downregulation of MDM2 expression and upregulation of P53. Next, RPS27A overexpression could partially reverse chidamide plus oxaliplatin induced growth inhibition and apoptosis in HCT-116 and RKO cells (P<0.01). RPS27A overexpression could promote the upregulation of MDM2 and downregulation of P53 after the combined treatment of chidamide with oxaliplatin. Chidamide and oxaliplatin acted synergistically to suppress CRC growth by the inhibition of the RPS27A-MDM2-p53 axis.

ATP13A2 activates the pentose phosphate pathway to promote colorectal cancer growth though TFEB-PGD axis.

The pentose phosphate pathway (PPP) is an important mechanism by which tumour cells resist stressful environments and maintain malignant proliferation. However, the mechanism by which the PPP regulates these processes in colorectal cancer (CRC) remains elusive. Closely related PPP genes were obtained from the TCGA and GEO databases. The effect of ATP13A2 on CRC cell proliferation was evaluated by performing in vitro assays. The connection between the PPP and ATP13A2 was explored by assessing proliferation and antioxidative stress. The molecular mechanism by which ATP13A2 regulates the PPP was investigated using chromatin immunoprecipitation and dual luciferase experiments. The clinical therapeutic potential of ATP13A2 was explored using patient-derived xenograft (PDX), patient-derived organoid (PDO) and AOM/DSS models. We identified ATP13A2 as a novel PPP-related gene. ATP13A2 deficiency inhibited CRC growth and PPP activity, as manifested by a decrease in the levels of PPP products and an increase in reactive oxygen species levels, whereas ATP13A2 overexpression induced the opposite effect. Mechanistically, ATP13A2 regulated the PPP mainly by affecting phosphogluconate dehydrogenase (PGD) mRNA expression. Subsequent studies showed that ATP13A2 overexpression promoted TFEB nuclear localization by inhibiting the phosphorylation of TFEB, thereby enhancing the transcription of PGD and ultimately affecting the activity of the PPP. Finally, ATP13A2 knockdown inhibited CRC growth in PDO and PDX models. ATP13A2- /- mice had a lower CRC growth capacity than ATP13A2+/+ in the AOM/DSS model.Our findings revealed that ATP13A2 overexpression-driven dephosphorylation of TFEB promotes PPP activation by increasing PGD transcription, suggesting that ATP13A2 may serve as a potential target for CRC therapy.

Targeted DNA demethylation of the ZNF334 promoter inhibits colorectal cancer growth

Colorectal cancer (CRC) is a leading cause of cancer deaths worldwide. Aberrant regulation of DNA methylation in promoters of tumor suppressor genes or proto-oncogenes is one of the fundamental processes driving the initiation and progression of CRC. Zinc-finger proteins (ZNFs) are one of the most abundant groups of proteins and function in many important biological processes related to tumorigenesis. Herein, we detected abnormal hypermethylation of the ZNF334 gene in CRC tissues compared with normal tissues, and this modification downregulated the expression of ZNF334. Furthermore, ten-eleven translocation 1 (TET1) was identified to be involved in regulating the methylation level of ZNF334. Next, a dCas9-multiGCN4/scFv-TET1CD-sgZNF334-targeted demethylation system was constructed to reverse the expression of ZNF334 through sgRNA targeting the ZNF334 promoter. Both in vitro and in vivo experiments demonstrated the targeted demethylation system upregulated ZNF334 expression and inhibited CRC growth. Collectively, targeted DNA demethylation of the ZNF334 promoter sheds light on the precise treatment of CRC.

Berberine promotes IGF2BP3 ubiquitination by TRIM21 to induce G1/S phase arrest in colorectal cancer cells

The increasing incidence of colorectal cancer (CRC) has become a major global public health burden. The natural drug Berberine (BBR) has shown potential in preventing CRC, and IGF2 mRNA binding protein 3 (IGF2BP3) may be a target of BBR. This study aims to investigate the mechanisms of BBR acting on IGF2BP3 to improve CRC. The results showed that IGF2BP3 played an important role in the development of CRC. BBR down-regulated IGF2BP3 expression and inhibited CRC growth in mice. Cell thermodynamic stability analysis (CETSA) and drug affinity responsive target stability (DARTS) analysis showed BBR may bind to IGF2BP3. BBR may induce structural changes in IGF2BP3 and decrease its protein stability in cytoplasm. The results from Co-Immunoprecipitation (Co-IP) suggested that BBR promoted the ubiquitination of IGF2BP3 by tripartite motif-containing protein 21 (TRIM21). Through RNA binding protein Immunoprecipitation (RIP) assay, it was found BBR inhibited the stabilization of CDK4/CCND1 mRNA by IGF2BP3 and promoted G1/S phase arrest in CRC cells. Overexpression of IGF2BP3 in vitro and in vivo attenuated the inhibition of CRC growth by BBR. This work demonstrated the potential of BBR targeting to IGF2BP3 in improving CRC and provided a new strategy for clinical treatment on CRC as well as novel anticancer drug design based on IGF2BP3 and TRIM21.

Akkermansia muciniphila inhibits tryptophan metabolism via the AhR/β-catenin signaling pathway to counter the progression of colorectal cancer.

Akkermansia muciniphila (A. muciniphila), a gram-negative anaerobic bacterium, is selectively decreased in the fecal microbiota of patients with colorectal cancer (CRC), but its molecular mechanism in CRC development remains inconclusive. In this study, we first confirmed the inhibitory effect of A. muciniphila on CRC formation and analyzed the metabolic role of intestinal flora in human Polyps, A-CRA (advanced colorectal adenoma) and CRC samples. To better clarify the role of A. muciniphila in CRC development, a pseudo-germ-free (GF) azoxymethane (AOM)/dextran sulfate sodium (DSS) mouse model was established, followed by infection with or without A. muciniphila. Metabolomic analysis and RNA-seq analysis showed tryptophan-mediated aryl hydrocarbon receptor (AhR) was significantly down-regulated in A. muciniphila-infected CRC mice. Then, mice with intestinal specific AhR deficiency (AhRfl/fl Cre) were generated and were used in 2 murine models: AOM/DSS treatment as a model of carcinogen-induced colon cancer and a genetically induced model using ApcMin/+ mice. Notably, AhR deficiency inhibited CRC growth in the AOM/DSS and ApcMin/+ mouse model. Moreover, AhR deficiency inhibited, rather than enhanced, tumor formation and tumor-derived organoids in Apc-deficient cells both in vivo and in vitro by activating Wnt/β-catenin signaling and TCF4/LEF1-dependent transcription. Furthermore, the antitumor effectiveness of A. muciniphila was abolished either in a human colon cancer tumor model induced by subcutaneous transplantation of AhR-silenced CRC cells, or AhR-deficienty spontaneous colorectal cancer model. In conclusion, supplementation with A. muciniphila. protected mice from CRC development by specifically inhibiting tryptophan-mediated AhR/β-catenin signaling.

Bufalin Inhibits Tumorigenesis, Stemness, and Epithelial-Mesenchymal Transition in Colorectal Cancer through a C-Kit/Slug Signaling Axis.

Colorectal cancer (CRC) is a major source of morbidity and mortality, characterized by intratumoral heterogeneity and the presence of cancer stem cells (CSCs). Bufalin has potent activity against many tumors, but studies of its effect on CRC stemness are limited. We explored bufalin's function and mechanism using CRC patient-derived organoids (PDOs) and cell lines. In CRC cells, bufalin prevented nuclear translocation of β-catenin and down-regulated CSC markers (CD44, CD133, LGR5), pluripotency factors, and epithelial-mesenchymal transition (EMT) markers (N-Cadherin, Slug, ZEB1). Functionally, bufalin inhibited CRC spheroid formation, aldehyde dehydrogenase activity, migration, and invasion. Network analysis identified a C-Kit/Slug signaling axis accounting for bufalin's anti-stemness activity. Bufalin treatment significantly downregulated C-Kit, as predicted. Furthermore, overexpression of C-Kit induced Slug expression, spheroid formation, and bufalin resistance. Similarly, overexpression of Slug resulted in increased expression of C-Kit and identical functional effects, demonstrating a pro-stemness feedback loop. For further study, we established PDOs from diagnostic colonoscopy. Bufalin differentially inhibited PDO growth and proliferation, induced apoptosis, restored E-cadherin, and downregulated CSC markers CD133 and C-Myc, dependent on C-Kit/Slug. These findings suggest that the C-Kit/Slug axis plays a pivotal role in regulating CRC stemness, and reveal that targeting this axis can inhibit CRC growth and progression.

Design, synthesis, and biological evaluation of tetrahydroquinolinones and tetrahydroquinolines with anticancer activity

Colorectal cancer (CRC) is the most commonly diagnosed cancer in Europe and the United States and the second leading cause of cancer related mortality. A therapeutic strategy used for the treatment of CRC involves targeting the intracellular levels of reactive oxygen species (ROS). In this study, we synthesized a series of novel tetrahydroquinolinones and assessed their ability to inhibit CRC growth and proliferation by evoking cellular stress through ROS. Our results revealed that (2-oxo-4-phenyl-5,6,7,8-tetrahydroquinolin-8-yl) N-(3-fluorophenyl)carbamate (20d) exhibited in vitro antiproliferative activity at micromolar concentrations. The compound also suppressed colony formation and the migration of HCT-116 cells, as well as deregulated the expression of several proteins involved in cell proliferation and metastasis. Furthermore, 20d induced massive oxidative stress by disrupting the balance of cells survival resulting in autophagy via the PI3K/AKT/mTOR signaling pathway. These findings suggest that this tetrahydroquinolinone can be an ideal lead compound for drug discovery based on quinone derivatives.

Toosendanin inhibits colorectal cancer cell growth through the Hedgehog pathway by targeting Shh.

Colorectal cancer (CRC) is one of the most common gastrointestinal cancers worldwide. This complex and often fatal disease has a high mortality rate. The Hedgehog (Hh) signaling pathway is crucial in CRC. Many studies have indicated that Shh is overexpressed in cancer stem cells (CSCs), and shh overexpression is positively correlated with CRC tumorigenesis. New drugs that kill CRC cells through the Hh pathway are needed. Toosendanin (TSN), a natural triterpenoid saponin extracted from the bark or fruit of Melia toosendan Sieb. et Zucc, can inhibit various tumors. Here, we investigated the effects of TSN in CRC and explored the possible targets and mechanisms. Shh-Light Ⅱ cells were treated with TSN and tested by dual luciferase reporter assays to determine the relationship with the Hh pathway. Cell Counting Kit-8 (CCK-8) assays were used to test the inhibitory effects of TSN on CRC cells. The expression of Hh components after TSN treatment was detected using western blots and quantitative reverse transcription polymerase chain reaction. Cellular thermal shift assays confirmed the targets of TSN. The same effects of TSN on xenograft tumor growth were investigated in vivo. The average weight, volume of the finally resected tumor, and the expression of Shh in the TSN-treated groups were significantly lower than those of the control group. This result strongly suggested that TSN administration inhibited CRC growth in vivo. Our research preliminarily demonstrated that the target of TSN was Shh and that TSN inhibits CRC cell growth by inhibiting the Hh pathway, identifying a new anticancer molecular mechanism of TSN in CRC.

A novel and potent dihydroorotate dehydrogenase inhibitor suppresses the proliferation of colorectal cancer by inducing mitochondrial dysfunction and DNA damage

AbstractDihydroorotate dehydrogenase (DHODH) is a quite attractive target in cancer therapy. Nevertheless, the antitumor effects of DHODH inhibitors against colorectal cancer (CRC) and the underlying mechanism have seldom been studied. Here, we explored the anti‐CRC efficacy of M62, a novel and potent DHODH inhibitor. In the study, M62 significantly inhibited the proliferation of CRC cells and induced S phase arrest. The antiproliferative effects caused by M62 were rescued by uridine supplementation. Mechanistically, messenger RNA sequencing results showed that M62‐triggered gene changes related to mitochondrial dysfunction, DNA damage, and DNA damage repair. Further validation showed that M62 treatment induced the generation of reactive oxygen species and decreased ΔΨm and ATP production. Meanwhile, M62 induced the accumulation of γ‐H2AX and longer comet tail moment, which were both markers of DNA damage. The downregulated DNA repair proteins and PI3K/ATK pathway were observed after M62 treatment. Furthermore, M62 significantly inhibited CRC xenograft tumor growth without detectable toxicity. Therefore, we conclude that M62 inhibits CRC growth both in vitro and in vivo by causing mitochondrial dysfunction and DNA damage, suggesting that DHODH inhibitors are potential therapeutic strategies for treating CRC.

HOXB9 Overexpression Promotes Colorectal Cancer Progression and Is Associated with Worse Survival in Liver Resection Patients for Colorectal Liver Metastases.

As is known, HOXB9 is an important factor affecting disease progression and overall survival (OS) in cancer. However, its role in colorectal cancer (CRC) remains unclear. We aimed to explore the role of HOXB9 in CRC progression and its association with OS in colorectal liver metastases (CRLM). We analysed differential HOXB9 expression in CRC using the Tissue Cancer Genome Atlas database (TCGA). We modulated HOXB9 expression in vitro to assess its impact on cell proliferation and epithelial-mesenchymal transition (EMT). Lastly, we explored the association of HOXB9 protein expression with OS, using an institutional patient cohort (n = 110) who underwent liver resection for CRLM. Furthermore, HOXB9 was upregulated in TCGA-CRC (n = 644) vs. normal tissue (n = 51) and its expression levels were elevated in KRAS mutations (p < 0.0001). In vitro, HOXB9 overexpression increased cell proliferation (p < 0.001) and upregulated the mRNA expression of EMT markers (VIM, CDH2, ZEB1, ZEB2, SNAI1 and SNAI2) while downregulated CDH1, (p < 0.05 for all comparisons). Conversely, HOXB9 silencing disrupted cell growth (p < 0.0001). High HOXB9 expression (HR = 3.82, 95% CI: 1.59–9.2, p = 0.003) was independently associated with worse OS in CRLM-HOXB9-expressing patients after liver resection. In conclusion, HOXB9 may be associated with worse OS in CRLM and may promote CRC progression, whereas HOXB9 silencing may inhibit CRC growth.

Tumor-suppressive circRHOBTB3 is excreted out of cells via exosome to sustain colorectal cancer cell fitness

Background & AimsTo clarify the biological roles, circularization process and secretion pathway of circRHOBTB3 in colorectal cancer (CRC) progression.MethodsWe performed a comprehensive analysis of circRNA levels in serum exosomes from multiple types of cancer patients in public databases and verified the higher level of circRHOBTB3 in CRC sera versus healthy donors by RT-qPCR. Then, the function of circRHOBTB3 in CRC was investigated in vitro and in vivo. RNA-seq and RNA pull-down assays together with mass spectrometry identified the downstream signals and the binding proteins of circRHOBTB3. Finally, Antisense oligonucleotides (ASOs) were designed to target circularization and secretion elements of circRHOBTB3 for CRC therapy.ResultscircRHOBTB3 levels were increased in the sera but was downregulated in tissue samples in CRC, and the downregulation was associated with poor prognosis. Furthermore, circRHOBTB3 acts a tumor-suppressive circRNA by repressing metabolic pathways, intracellular ROS production in CRC. Several key elements were discovered to regulate circRHOBTB3 circularization and exosomal secretion. Moreover, SNF8 was identified that sorts circRHOBTB3 into exosomes. Interestingly, we found that CRC cells could actively secrete more circRHOBTB3 than normal cells. According to the sequence of regulatory elements for circularization and exosomal secretion, we designed and synthesized ASOs, which increased circRHOBTB3 expression and blocked circRHOBTB3 exosomal secretion. More importantly, ASOs could inhibit CRC growth and metastasis in vitro and in vivo.ConclusionscircRHOBTB3 plays a tumor-suppressive role in CRC and has to be excreted out of cells to sustain cancer cell fitness. ASOs targeting regulatory elements for circularization and exosomal secretion will become a novel antitumor strategy.

Decylubiquinone Inhibits Colorectal Cancer Growth Through Upregulating Sirtuin2.

Colorectal cancer (CRC) is one of the leading causes of cancer-related death worldwide. Decylubiquinone (DUb), a coenzyme Q10 analog, was reported to inhibit breast cancer growth and metastasis by us. However, the influence of DUb on CRC remains unclear. Herein, we found that DUb significantly inhibited CRC growth in the patient-derived xenograft (PDX) and CT26 xenograft models. DUb was further identified to significantly suppress CRC cell proliferation, colony formation, migration and invasion in a dose-dependent manner, while not inhibiting CRC cell apoptosis from flow cytometry assay. Sirtuin2 (SIRT2), a member of the sirtuin protein family, plays a critical role in growth and metastasis in various cancers. Moreover, DUb inhibited CRC progression by upregulating SIRT2. These findings reveal that DUb has the potential to a novel drug for the treatment of CRC by inhibiting CRC cell proliferation.

Degradation of HIF-1α induced by curcumol blocks glutaminolysis and inhibits epithelial-mesenchymal transition and invasion in colorectal cancer cells.

Colorectal cancer (CRC) has high morbidity and mortality. Epithelial-mesenchymal transition (EMT) is associated with CRC progression and metastasis. Glutaminolysis is essential for malignancy of cancer cells. Here, we examined the effects of curcumol on CRC EMT. We observed that curcumol suppressed invasion and migration in human CRC cells associated with upregulation of epithelial markers E-cadherin and Zonula occludens 1 and downregulation of mesenchymal markers N-cadherin and Vimentin as well as EMT-related transcription factors Snail and Twist. Curcumol increased intracellular levels of glutamine but decreased intracellular levels of glutamate, α-ketoglutarate, ATP, glutathione, and tricarboxylic acid cycle metabolites, suggesting interruption of glutaminolysis. Next, curcumol repressed glutaminase 1 (Gls1) mRNA and protein expression, and overexpression of Gls1 promoted EMT and abolished curcumol effects on CRC cell EMT. Molecular examinations showed that curcumol stimulated protein degradation of hypoxia-inducible factor-1α (HIF-1α) and prevented its nuclear accumulation in CRC cells. HIF-1α agonist deferoxamine (DFO) promoted HIF-1α binding to Gls1 promoter and increased Gls1 expression but abolished curcumol's inhibitory effects on Gls1 expression. DFO also enhanced EMT and invasion and migration in CRC cells and eliminated curcumol effects. Furthermore, mouse CRC models were established with in vivo overexpression of HIF-1α and Gls1. Curcumol effectively inhibited CRC growth, metastasis, and EMT in mice, which was abrogated by overexpression of HIF-1α or Gls1. Altogether, stimulation of HIF-1α degradation was required for curcumol to disrupt EMT and repress invasion and migration in CRC cells through inhibiting Gls1-mediated glutaminolysis. Curcumol could be a promising candidate for intervention of CRC metastasis. • Curcumol inhibits EMT and blocks glutaminolysis in CRC cells. • Inhibition of Gls1 is required for curcumol blockade of glutaminolysis and EMT. • Curcumol induces HIF-1α degradation leading to inhibition of Gls1 and blockade of glutaminolysis and EMT. • Curcumol suppresses CRC growth and metastasis via inhibiting HIF-1α, glutaminolysis and EMT in mice.

Exosomal circ_PTPRA inhibits tumorigenesis and promotes radiosensitivity in colorectal cancer by enriching the level of SMAD4 via competitively binding to miR-671-5p.

Accumulating evidence supports that exosomal RNAs are crucial in tumor microenvironment and may be used as diagnostic biomarkers for cancers. This study aimed to determine the role of exosomal circular RNA_protein tyrosine phosphatase receptor type A (circ_PTPRA) in colorectal cancer (CRC). The morphology of exosomes was identified by transmission electron microscopy (TEM), and several exosome-specific proteins were quantified by western blot. The expression of circ_PTPRA, miR-671-5p and SMAD family member 4 (SMAD4) was detected using quantitative polymerase chain reaction (qPCR). Cell cycle was assessed using flow cytometry assay. Cell proliferation was assessed by MTT assay. Radiosensitivity was observed according to colony growth and cell apoptosis rate by colony formation assay and flow cytometry assay. The protein levels of proliferation- and apoptosis-related markers and SMAD4 were measured by western blot. The predicted relationship between miR-671-5p and circ_PTPRA or SMAD4 was verified by dual-luciferase reporter assay. Animal study was performed to investigate the role of exosomal circ_PTPRA in vivo. Circ_PTPRA expression was declined in serumal exosomes from CRC patients and CRC cell lines. Exosomal circ_PTPRA induced CRC cell cycle arrest and inhibited cell proliferation. Besides, exosomal circ_PTPRA promoted radiosensitivity of CRC cells, leading to inhibitory colony formation and increased apoptotic rate. In mechanism, circ_PTPRA functioned as a competing endogenous RNA (ceRNA) to increasing SMAD4 level by binding to miR-671-5p. Rescue experiments concluded that circ_PTPRA inhibited CRC growth and radioresistance by decreasing miR-671-5p expression, and miR-671-5p inhibition also inhibited CRC growth and radioresistance by enriching SMAD4 expression. Moreover, exosomal circ_PTPRA blocked tumor growth in vivo. Exosomal circ_PTPRA enhanced CRC cell radiosensitivity and inhibited CRC malignant development partially by regulating the miR-671-5p/SMAD4 pathway, hinting that exosomal circ_PTPRA might be used as a potential predicted and therapeutic target for CRC.

Choline-induced SLC5A7 impairs colorectal cancer growth by stabilizing p53 protein.

The members of the solute carrier (SLC) superfamily are vital membrane transporters in human cells. In the present study, we determine the expression and function of SLC5 family members in colorectal cancer (CRC). Expression analysis based on The Cancer Genome Atlas database and potential clinical relation analysis based on the Oncomine database indicate that SLC5A7 is downregulated and is predicted to correlate with the staging, and prognosis response of CRC. Additional results demonstrate that SLC5A7 is downregulated and correlates with good prognosis in patients with CRC. Ectopic expression of SLC5A7 either by overexpression, or uptake of choline efficiently inhibits CRC growth. Examination of the molecular mechanism reveals that SLC5A7 promotes p53 protein expression by directly interacting with and modifying p53 and disrupting the interaction between p53 and MDM2 in wild type p53 CRC cells. Our findings establish the clear correlation between SLC5A7 and tumour growth, providing a novel potential therapeutic target for CRC.

Qingjie Fuzheng Granule Inhibits EMT and Induces Autophagy in Colorectal Cancer via mTOR Signaling Pathways.

Qingjie Fuzheng granule (QFG) is a traditional Chinese medicinal formula used extensively as an alternative medicine for cancer treatment, including colorectal cancer (CRC). But its pathological mechanism in CRC is unclear. To study antitumor treatment effects and mechanisms of QFG, we established a CRC HCT-116 xenograft mouse model and assessed QFG on EMT and autophagy progression in vivo. The mice were randomly divided into 2 groups (n = 10 each group) and treated with intragastric administration of 1 g/kg of QFG or saline 6 days a week for 28 days (4 weeks). Body weight was measured every other day with electronic balance. At the end of the treatment, the tumor weight was measured. Immunohistochemical (IHC) and western blot (WB) assay were used to detect the expression level of E-cadherin, N-cadherin, vimentin, and TWIST1 to evaluate the effect of QFG on tumor cell EMT progression. IHC and WB assay were also used to detect the expression level of beclin-1, LC3-II, and p62 to evaluate the effect of QFG on tumor cell autophagy progression. Furthermore, the expression level of relative proteins in mTOR pathway was detected by WB assay to investigate the mechanism of QFG effect on CRC. We discovered that QFG inhibited the rise of tumor weight while it had no effect on mice body weight, which proved that QFG could inhibit CRC growth progression without significant side effects in vivo. In addition, QFG treatment inhibited EMT and induced autophagy progression in CRC tumor cells, including that QFG upregulated the expression of E-cadherin, beclin-1, and LC3-II, but downregulated the expression of N-cadherin, vimentin, TWIST1, and p62. And, QFG decreased the ratio of p-PI3K/PI3K, p-AKT/AKT, and p-mTOR/mTOR, but increased the ratio of p-AMPK/AMPK. All findings from this research proved that QFG can induce autophagy and inhibit EMT progression in CRC via regulating the mTOR signaling pathway.

An MST4-pβ-CateninThr40 Signaling Axis Controls Intestinal Stem Cell and Tumorigenesis.

Elevated Wnt/β‐catenin signaling has been commonly associated with tumorigenesis especially colorectal cancer (CRC). Here, an MST4‐pβ‐cateninThr40 signaling axis essential for intestinal stem cell (ISC) homeostasis and CRC development is uncovered. In response to Wnt3a stimulation, the kinase MST4 directly phosphorylates β‐catenin at Thr40 to block its Ser33 phosphorylation by GSK3β. Thus, MST4 mediates an active process that prevents β‐catenin from binding to and being degraded by β‐TrCP, leading to accumulation and full activation of β‐catenin. Depletion of MST4 causes loss of ISCs and inhibits CRC growth. Mice bearing either MST4T178E mutation with constitutive kinase activity or β‐cateninT40D mutation mimicking MST4‐mediated phosphorylation show overly increased ISCs/CSCs and exacerbates CRC. Furthermore, the MST4‐pβ‐cateninThr40 axis is upregulated and correlated with poor prognosis of human CRC. Collectively, this work establishes a previously undefined machinery for β‐catenin activation, and further reveals its function in stem cell and tumor biology, opening new opportunities for targeted therapy of CRC.