Knockdown by MSH2 and EPCAM siRNA suppress Wnt/β-Catenin Pathway in HCT116 Cell Line

Abstract Purpose: Small interfering RNA (siRNA) has the potential as a therapeutic approach against selective pathways in colorectal cancer. EPCAM, a transmembrane glycoprotein mediating cell adhesion, was known to be involved in suppressing Wnt/β-catenin pathway, an important pathway for tumour progression in colon cancer cells. EPCAM deletions caused a transcriptional read-through that may silence its neighbouring gene, MSH2. This study aimed to investigate the effect of co-siRNA targeted genes, MSH2 and EPCAM, in colon cancer cell line, HCT116, and their effect in regulating the Wnt/β-catenin pathway. Methods: Pre-designed siRNA of MSH2 and EPCAM were transfected into HCT116 cells. The cells were divided into six group of treatments: untreated cell group, cells treated with negative control siRNA, MSH2-siRNA treated cells, EPCAM-siRNA treated cells, cells treated with both EPCAM and MSH2-siRNA, and cells treated with transfection reagent (mock control). The mRNA and protein expression following the individual and combined siRNA treatments were assessed by quantitative polymerase chain reaction and Western blot. Results: The mRNA and protein expression levels of MSH2, EPCAM and β-catenin were reduced in the individual MSH2 and EPCAM-siRNA treated samples as compared to the untreated sample. Further reduction of mRNA and protein expressions for MSH2, EPCAM and β-catenin were identified in combined siRNA treatments. Conclusion: Reduction of β-catenin expression by simultaneous silencing of MSH2 and EPCAM suggested that these genes may play a role in supressing the Wnt/β-catenin pathway in cancer cells.

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Expression of Monocyte Chemotactic Protein-1 in Human Endometrial Cancer Cells and the Effect of Treatment with Tamoxifen or Buserelin

Journal of International Medical Research ◽

10.1177/147323000603400307 ◽

2006 ◽

Vol 34 (3) ◽

pp. 284-290 ◽

Cited By ~ 2

Author(s):

L Wang ◽

W Zheng ◽

S Zhang ◽

X Chen ◽

D Hornung

Keyword(s):

Endometrial Cancer ◽

Protein Expression ◽

Cancer Cells ◽

Cell Line ◽

Enzyme Linked Immunosorbent Assay ◽

Releasing Hormone ◽

Monocyte Chemotactic Protein ◽

Chemotactic Protein ◽

Mrna And Protein Expression ◽

Gonadotrophin Releasing Hormone

Monocyte chemotactic protein-1 (MCP-1) is an important determinant of macrophage infiltration in tumours. This study investigated the effect of tamoxifen and the gonadotrophin-releasing hormone agonist buserelin on MCP-1 in the human endometrial cancer cell line EFE-184. Reverse transcription polymerase chain reaction and Western blot analysis were used to determine MCP-1 mRNA and protein expression, respectively. Immunoreactive MCP-1 in the cell culture media was quantified by enzyme-linked immunosorbent assay. Tamoxifen inhibited MCP-1 mRNA and protein expression in endometrial cancer cells and inhibited MCP-1 secretion in a time- and dose-dependent manner at concentrations of 10−7 to 10−5 M. Buserelin had no significant effect on MCP-1 mRNA and protein expression. These results suggest that tamoxifen directly inhibits the expression of MCP-1 in this cell line by blocking the MCP-1 signalling pathways. These findings may contribute to the understanding of the mechanisms underlying the different effects of tamoxifen and gonadotrophin-releasing hormone agonists in the treatment of endometrial cancer.

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Silibinin Inhibits TGF-β-induced MMP-2 and MMP-9 Through Smad Signaling Pathway in Colorectal Cancer HT-29 Cells

Basic & Clinical Cancer Research ◽

10.18502/bccr.v12i2.5752 ◽

2021 ◽

Author(s):

Zahra Zare ◽

Tina Nayerpour dizaj ◽

Armaghan Lohrasbi ◽

Zakieh Sadat Sheikhalishahi ◽

Amirhooman Asadi ◽

...

Keyword(s):

Colorectal Cancer ◽

Protein Expression ◽

Cell Viability ◽

Cancer Cells ◽

Cell Line ◽

Dependent Manner ◽

Smad2 Phosphorylation ◽

Mrna And Protein Expression ◽

Dose Dependent ◽

Ht 29

Background: Metastasis of cancer cells is the primary responsible for death in patients with colorectal cancer (CRC). Transforming growth factor-β (TGF-β)-induced matrix metalloproteinases (MMPs) are essential for the metastasis process. Silibinin is a natural compound extracted from the Silybum marianum that exhibits anti-neoplastic activity in cancer cell lines. In this study, we evaluated the effects of silibinin on MMP-2 and MMP-9 induced by TGF-β in human HT-29 CRC cell line and the potential mechanism underlying the effects. Methods: The present in vitro study was done on the HT-29 cell line. The HT-29 cell line was cultured in RPMI1640 and exposed to TGF- β (5 ng/ml) in the absence and presence of different concentrations of silibinin (10, 25, 50, and 100 μM). The effect of silibinin on HT-29 cell viability was measured with the MTT assay. A real-time polymerase chain reaction (Real-Time PCR) determined the relative mRNA expression of MMP-2 and MMP-9. Western blotting was employed to examine MMP-2 and MMP 9 protein expression and Smad2 phosphorylation. Results: Silibinin inhibits cell viability of HT-29 cell line at 24 hours in a dose-dependent manner. TGF-β increased the mRNA and protein expression of MMP-2, MMP-9, and phosphorylated Smad2 compared to controls. Pharmacological inhibition with silibinin markedly blocked TGF-β–induced MMP-2 and MMP-9 mRNA and protein expression and Smad2 phosphorylation. Conclusion: Silibinin decreased the cell viability of HT-29 cancer cells in a dose-dependent manner. Silibinin also inhibited TGF-β-stimulated MMP-2 and MMP-9 expression in HT-29 cells, possibly mediated with the Smad2 signaling pathway.

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Docking Studies and Antiproliferative Activities of 6-(3-aryl-2-propenoyl)-2(3H)-benzoxazolone Derivatives as Novel Inhibitors of Phosphatidylinositol 3-Kinase (PI3Kα)

Anti-Cancer Agents in Medicinal Chemistry ◽

10.2174/1871520620666200807221731 ◽

2020 ◽

Vol 20 ◽

Author(s):

Sinan Bilginer ◽

Sanaa K. Bardaweel ◽

Dima A. Sabbah ◽

Halise Inci Gul

Keyword(s):

Colon Cancer ◽

Cancer Cells ◽

Cell Line ◽

Caspase 3 ◽

Hct116 Cell ◽

Biological Evaluation ◽

Anticancer Activities ◽

Human Colon Cancer ◽

Chalcone Derivatives ◽

Antiproliferative Activities

Background: Cancer is a life-threatening group of diseases and universally the second main cause of death. Design and development of new scaffolds targeting selective cancer cells is considered a promising goal for cancer treatment. Aim and Objective: Chalcone derivatives; 6-(3-aryl-2-propenoyl)-2(3H)-benzoxazolone, were previously prepared and evaluated against the oral cavity squamous cell carcinoma cell line, HSC-2, and were reported to have remarkably high tumor selectivity. The aim of this study is to further investigate the anticancer activities of the chalcone derivatives against human colon cancer cells with possible elucidation of their mechanism of action. Methods: Computational studies were conducted to explore the potential interaction of the synthesized molecules with the phosphatidylinositol-4,5-bisphosphate 3-kinaseα (PI3Kα). Biological evaluation of the antiproliferative activities associated with compounds 1-23 was carried out against the colon cancer cell line HCT116. Lactate dehydrogenase (LDH) activity was measured to study necrosis while the caspase-3 activation and DNA measurements were used to evaluate apoptosis in the treated cells. Results: Glide studies against PI3Kα kinase domain demonstrated that the 6-(3-aryl-2-propenoyl)-2(3H)-benzoxazolone scaffold forms H-bond with K802, Y836, E849, V851, N853, Q859, and D933, and it fits the fingerprint of PI3Kα active inhibitors. Biological evaluation of the reported compounds in HCT116 cell line confirmed that the series inhibited PI3Kα activity and induced apoptosis via activation of caspase-3 and reduction of DNA content. Conclusion: The recently developed compounds might be employed as lead structures for the design of new antitumor drugs targeting PI3Kα.

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Combination Effects of Gambogic Acid on Imatinib Mesylate Cytotoxicity in Colon Cancer Cells

Key Engineering Materials ◽

10.4028/www.scientific.net/kem.859.27 ◽

2020 ◽

Vol 859 ◽

pp. 27-33

Author(s):

Ei Mon Khaing ◽

Thanika Saenpunya ◽

Pittawas Kerdklai ◽

Sornsawan Pangpongma ◽

Sarisa Vongvijit ◽

...

Keyword(s):

Colon Cancer ◽

Cell Proliferation ◽

Cancer Cells ◽

Cell Line ◽

Imatinib Mesylate ◽

Kinase Inhibitor ◽

Hct116 Cell ◽

Additive Effect ◽

Gambogic Acid ◽

Combination Effects

Imatinib mesylate (IM) is a kinase inhibitor with inhibitory effect on colon cancer cell proliferation. However, some adverse effect of imatinib and drug resistance are challenges for maintenance the therapeutic effect with lowering the dose; thus, the combination with other substances was of interest. Gambogic acid (GA), a natural compound from gamboge, was revealed for inhibition of cell proliferation in many types of cancers. This research aimed to investigate the effect of GA on IM response in colorectal cancer cells, HT29 and HCT116. The 50% inhibitory concentration (IC50) of IM and GA was determined. Concentrations which lower than IC50 of each compound were combined and tested for the combination effects on HT29 and HCT116 cells. The results were analyzed using isobologram to assess the types of interaction. The combination index (CI) of the tests was calculated at the 3 different percentages of inhibition (IC50, IC60 and IC70). The finding indicated that IC50 and IC60 of the combination of 5 and 7 μM IM with 0.2-1.2 μM GA showed antagonism while IC70 showed additive effect in HT29 cell line. In HCT116 cell line, IC50 of 10 μM IM with 0.1-0.8 μM GA showed antagonism while IC60 and IC70 expressed additive effect. For the studies with IC50 and IC60 of 12 μM IM with 0.1-0.8 μM GA showed antagonistic result while IC70 showed additive effect. The result indicated that, at the lower IC studied, the CI obtained from the experiments indicated the inhibitory effects, while the higher IC, the results showed the changing trend from antagonistic to additive and synergistic effects of GA on IM.

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Protein expression following γ-irradiation relevant to growth arrest and apoptosis in colon cancer cells

Journal of Cancer Research and Clinical Oncology ◽

10.1007/s00432-009-0606-4 ◽

2009 ◽

Vol 135 (11) ◽

pp. 1583-1592 ◽

Cited By ~ 14

Author(s):

Daniella Pfeifer ◽

Åsa Wallin ◽

Birgitta Holmlund ◽

Xiao-Feng Sun

Keyword(s):

Colon Cancer ◽

Protein Expression ◽

Cancer Cells ◽

Growth Arrest ◽

Colon Cancer Cells ◽

Γ Irradiation

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Schisandrin B Attenuates Colitis-Associated Colorectal Cancer through SIRT1 Linked SMURF2 Signaling

The American Journal of Chinese Medicine ◽

10.1142/s0192415x21500841 ◽

2021 ◽

pp. 1-17

Author(s):

Zhichen Pu ◽

Weiwei Zhang ◽

Minhui Wang ◽

Maodi Xu ◽

Haitang Xie ◽

...

Keyword(s):

Colorectal Cancer ◽

Colon Cancer ◽

Cell Growth ◽

Protein Expression ◽

Hct116 Cell ◽

Enzyme Linked Immunosorbent Assay ◽

Schisandrin B ◽

In Vivo Models

Colon cancer, a common type of malignant tumor, seriously endangers human health. However, due to the relatively slow progress in diagnosis and treatment, the clinical therapeutic technology of colon cancer has not been substantially improved in the past three decades. The present study was designed to investigate the effects and involved mechanisms of schisandrin B in cell growth and metastasis of colon cancer. C57BL/6 mice received AOM and dextran sulfate sodium. Mice in treatment groups were gavaged with 3.75–30 mg/kg/day of schisandrin B. Transwell chamber migration, enzyme-linked immunosorbent assay (ELISA), Western blot analysis, immunoprecipitation (IP) and immunofluorescence were conducted, and HCT116 cell line was employed in this study. Data showed that schisandrin B inhibited tumor number and tumor size in the AOD+DSS-induced colon cancer mouse model. Schisandrin B also inhibited cell proliferation and metastasis of colon cancer cells. We observed that schisandrin B induced SMURF2 protein expression and affected SIRT1 in vitro and in vivo. SMURF2 interacted with SIRT1 protein, and there was a negative correlation between SIRT1 and SMURF2 expressions in human colorectal cancer. The regulation of SMURF2 was involved in the anticancer effects of schisandrin B in both in vitro and in vivo models. In conclusion, the present study revealed that schisandrin B suppressed SIRT1 protein expression, and SIRT1 is negatively correlated with the induction of SMURF2, which inhibited cell growth and metastasis of colon cancer. Schisandrin B could be a leading compound, which will contribute to finding novel potential agents and therapeutic targets for colon cancer.

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A novel epigenetic regulation of circFoxp1 on Foxp1 in colon cancer cells

Cell Death and Disease ◽

10.1038/s41419-020-03007-6 ◽

2020 ◽

Vol 11 (9) ◽

Author(s):

Yanwei Luo ◽

Fengxia Liu ◽

Jinqi Ma ◽

Yunfeng Fu ◽

Rong Gui

Keyword(s):

Colon Cancer ◽

Cell Viability ◽

Cancer Cells ◽

Quantitative Polymerase Chain Reaction ◽

Human Colon ◽

Tumor Xenograft ◽

Colon Cancer Cells ◽

Poor Overall Survival ◽

Sw620 Cell

Abstract Foxp1 is a tumor suppressor in colon cancer. However, circFoxp1 derived from Foxp1 is an oncogene. In this study, we aim to investigate the role of circFoxp1 in colon cancer and the regulatory mechanism between circFoxp1 and Foxp1. 78 human colon tumor tissues and the matched paracancerous tissues were collected. Quantitative polymerase chain reaction, immunohistochemistry, quantitative methylation-specific PCR, chromatin immunoprecipitation assay, CCK-8 assay, and Tumor xenograft in nude mice were performed. The expression of circFoxp1 was increased and Foxp1 was reduced in colon cancer tissues, which were associated with a poor overall survival rate of the patients with colon cancer. CircFoxp1 recruited DNMT1 to the promoter of Foxp1, leading to promotor hypermethylation, thereby inhibiting Foxp1 transcription. Interfering circFoxp1 by siRNA in SW620 cells significantly inhibited cell viability, while knockdown Foxp1 expression partially restored SW620 cell viability. In addition, knockdown of circFoxp1 significantly sensitized colon cancer cells to Capecitabine in vitro and vivo through regulating Foxp1. We discovered a novel epigenetic pathway that circFoxp1 regulated Foxp1 in colon cancer cells. CircFoxp1 may regulate DNA methylation and demethylation to coordinate colon cancer cell proliferation and participate in chemotherapy drug responses. Therefore, circFoxp1 may be a potential therapeutic target for colon cancer.

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