Protease activated receptor 2 signaling promotes self-renewal and metastasis in colorectal cancer through β-catenin and periostin

Polymethoxylated flavones (PMFs) from citrus fruits are reported to present anticancer potential. However, there is a lack of information regarding their effect on cancer stem cell (CSC) populations, which has been recognized as responsible for tumor initiation, relapse, and chemoresistance. In this study, we evaluated the effect of an orange peel extract (OPE) and its main PMFs, namely, nobiletin, sinensetin, tangeretin, and scutellarein tetramethylether in targeting cell proliferation and stemness using a 3D cell model of colorectal cancer composed of HT29 cell spheroids cultured for 7 days in stirred conditions. Soft agar assay, ALDH1 activity, and relative quantitative gene expression analysis of specific biomarkers were carried out to characterize the stemness, self-renewal, and mesenchymal features of HT29 cell spheroids. Then, the impact of OPE and PMFs in reducing cell proliferation and modulating cancer stemness and self-renewal was assessed. Results showed that, when compared with monolayer cultures, HT29 cell spheroids presented higher ALDH1 activity (81.97% ± 5.27% compared to 63.55% ± 17.49% for 2D), upregulation of CD44, PROM1, SOX9, and SNAI1 genes (1.83 ± 0.34, 2.54 ± 0.51, 2.03 ± 0.15, and 6.12 ± 1.59 times) and high self-renewal capability (352 ± 55 colonies compared to 253 ± 42 for 2D). Incubation with OPE (1 mg/mL) significantly inhibited cell proliferation and modulated cancer stemness and self-renewal ability: colony formation, ALDH1 activity, and the expression of cancer stemness biomarkers PROM1 and LGR5 were significantly reduced (0.66 ± 0.15 and 0.51 ± 0.14 times, respectively). Among all PMFs, tangeretin was the most efficient in targeting the CSC population by decreasing colony formation and the expression of PROM1 and LGR5. Scutellarein tetramethylether was shown to modulate markers of mesenchymal/metastatic transition (increasing CDH1 and reducing ZEB1 and SNAI1) and nobiletin was capable of downregulating PROM1 and SNAI1 expression. Importantly, all PMFs and OPE were shown to synergistically interact with 5-fluorouracil, improving the antiproliferative response of this drug.

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Erratum: Corrigendum: Adenylate kinase hCINAP determines self-renewal of colorectal cancer stem cells by facilitating LDHA phosphorylation

Nature Communications ◽

10.1038/ncomms16000 ◽

2017 ◽

Vol 8 (1) ◽

Author(s):

Yapeng Ji ◽

Chuanzhen Yang ◽

Zefang Tang ◽

Yongfeng Yang ◽

Yonglu Tian ◽

...

Keyword(s):

Colorectal Cancer ◽

Stem Cells ◽

Cancer Stem Cells ◽

Adenylate Kinase ◽

Self Renewal ◽

Colorectal Cancer Stem Cells

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A novel BMI-1 inhibitor QW24 for the treatment of stem-like colorectal cancer

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-019-1392-8 ◽

2019 ◽

Vol 38 (1) ◽

Author(s):

Jinhua Wang ◽

Yajing Xing ◽

Yingying Wang ◽

Yundong He ◽

Liting Wang ◽

...

Keyword(s):

Colorectal Cancer ◽

Cell Proliferation ◽

Liver Metastasis ◽

Tumor Growth ◽

Cell Lines ◽

Xenograft Model ◽

Proliferation Assay ◽

Self Renewal ◽

Metastasis Model

Abstract Background Cancer-initiating cell (CIC), a functionally homogeneous stem-like cell population, is resonsible for driving the tumor maintenance and metastasis, and is a source of chemotherapy and radiation-therapy resistance within tumors. Targeting CICs self-renewal has been proposed as a therapeutic goal and an effective approach to control tumor growth. BMI-1, a critical regulator of self-renewal in the maintenance of CICs, is identified as a potential target for colorectal cancer therapy. Methods Colorectal cancer stem-like cell lines HCT116 and HT29 were used for screening more than 500 synthetic compounds by sulforhodamine B (SRB) cell proliferation assay. The candidate compound was studied in vitro by SRB cell proliferation assay, western blotting, cell colony formation assay, quantitative real-time PCR, flow cytometry analysis, and transwell migration assay. Sphere formation assay and limiting dilution analysis (LDA) were performed for measuring the effect of compound on stemness properties. In vivo subcutaneous tumor growth xenograft model and liver metastasis model were performed to test the efficacy of the compound treatment. Student’s t test was applied for statistical analysis. Results We report the development and characterization of a small molecule inhibitor QW24 against BMI-1. QW24 potently down-regulates BMI-1 protein level through autophagy-lysosome degradation pathway without affecting the BMI-1 mRNA level. Moreover, QW24 significantly inhibits the self-renewal of colorectal CICs in stem-like colorectal cancer cell lines, resulting in the abrogation of their proliferation and metastasis. Notably, QW24 significantly suppresses the colorectal tumor growth without obvious toxicity in the subcutaneous xenograft model, as well as decreases the tumor metastasis and increases mice survival in the liver metastasis model. Moreover, QW24 exerts a better efficiency than the previously reported BMI-1 inhibitor PTC-209. Conclusions Our preclinical data show that QW24 exerts potent anti-tumor activity by down-regulating BMI-1 and abrogating colorectal CICs self-renewal without obvious toxicity in vivo, suggesting that QW24 could potentially be used as an effective therapeutic agent for clinical colorectal cancer treatment.

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Chemotherapy activates cancer-associated fibroblasts to maintain colorectal cancer-initiating cells by IL-17A

Journal of Experimental Medicine ◽

10.1084/jem.20131195 ◽

2013 ◽

Vol 210 (13) ◽

pp. 2851-2872 ◽

Cited By ~ 165

Author(s):

Fiorenza Lotti ◽

Awad M. Jarrar ◽

Rish K. Pai ◽

Masahiro Hitomi ◽

Justin Lathia ◽

...

Keyword(s):

Colorectal Cancer ◽

Tumor Microenvironment ◽

Cellular Heterogeneity ◽

Cancer Therapies ◽

Cancer Associated Fibroblasts ◽

Self Renewal ◽

Cytokines And Chemokines ◽

Response To Chemotherapy ◽

Cytotoxic Treatment

Many solid cancers display cellular hierarchies with self-renewing, tumorigenic stemlike cells, or cancer-initiating cells (CICs) at the apex. Whereas CICs often exhibit relative resistance to conventional cancer therapies, they also receive critical maintenance cues from supportive stromal elements that also respond to cytotoxic therapies. To interrogate the interplay between chemotherapy and CICs, we investigated cellular heterogeneity in human colorectal cancers. Colorectal CICs were resistant to conventional chemotherapy in cell-autonomous assays, but CIC chemoresistance was also increased by cancer-associated fibroblasts (CAFs). Comparative analysis of matched colorectal cancer specimens from patients before and after cytotoxic treatment revealed a significant increase in CAFs. Chemotherapy-treated human CAFs promoted CIC self-renewal and in vivo tumor growth associated with increased secretion of specific cytokines and chemokines, including interleukin-17A (IL-17A). Exogenous IL-17A increased CIC self-renewal and invasion, and targeting IL-17A signaling impaired CIC growth. Notably, IL-17A was overexpressed by colorectal CAFs in response to chemotherapy with expression validated directly in patient-derived specimens without culture. These data suggest that chemotherapy induces remodeling of the tumor microenvironment to support the tumor cellular hierarchy through secreted factors. Incorporating simultaneous disruption of CIC mechanisms and interplay with the tumor microenvironment could optimize therapeutic targeting of cancer.

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Overexpression of PER3 Inhibits Self-Renewal Capability and Chemoresistance of Colorectal Cancer Stem-Like Cells via Inhibition of Notch and β-Catenin Signaling

Oncology Research Featuring Preclinical and Clinical Cancer Therapeutics ◽

10.3727/096504016x14772331883976 ◽

2017 ◽

Vol 25 (5) ◽

pp. 709-719 ◽

Cited By ~ 13

Author(s):

Feng Zhang ◽

Hong Sun ◽

Sai Zhang ◽

Xin Yang ◽

Guogang Zhang ◽

...

Keyword(s):

Colorectal Cancer ◽

Self Renewal

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RNAi mediated silencing of Nanog expression suppresses the growth of human colorectal cancer stem cells

10.21203/rs.3.rs-22167/v1 ◽

2020 ◽

Author(s):

Chen Zhang ◽

Yuanyuan Zhao ◽

Yongjing Yang ◽

Chunlian Zhong ◽

Tianju Ji ◽

...

Keyword(s):

Colorectal Cancer ◽

Stem Cells ◽

Cancer Stem Cells ◽

Small Interfering Rna ◽

Embryonic Stem ◽

Self Renewal ◽

Nanog Expression ◽

Hct 116 ◽

Hct 116 Cells ◽

Interfering Rna

Abstract Background: Colorectal cancer (CRC) is the third most common cancer in the world known for its poor recurrence-free prognosis. Previous studies have shown that it is closely linked with cancer stem cells (CSCs), which have self-renewal potential and the capacity to differentiate into diverse populations. Nanog is an important transcription factor that functions to maintain the self-renewal and proliferation of embryonic stem cells; however, many recent studies have shown that Nanog is also highly expressed in many cancer stem cells.Methods: To investigate whether Nanog plays a crucial role in maintaining the stemness of colorectal CSCs (CCSCs), RNA interference was used to downregulate Nanog expression in the CRC stem cell line, EpCAM+CD44+HCT-116. We examined the anti-tumor function of Nanog in vitro and in vivo, using small interfering RNA.Results: Our results revealed that the Nanog mRNA expression level in CCSCs was higher than that in HCT-116 cells. We found that the depletion of Nanog inhibited proliferation and promoted apoptosis in EpCAM+CD44+HCT-116 cells. In addition, the invasive ability of EpCAM+CD44+HCT-116 cells was markedly restricted when Nanog was silenced by small interfering RNA. Furthermore, we found that the silencing of Nanog decreased tumor size and weight and improved the survival rate of tumor-bearing mice.Conclusions: In conclusion, these findings collectively demonstrate that Nanog, which is highly expressed in CRC stem cells, is a key factor in the development of tumor growth, and it may serve as a potential marker of prognosis and a novel and effective therapeutic target for the treatment of CRC.

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SKP1 promotes YAP-mediated colorectal cancer stemness via suppressing RASSF1

10.21203/rs.3.rs-29004/v3 ◽

2020 ◽

Author(s):

Cong Tian ◽

Tingyuan Lang ◽

Jiangfeng Qiu ◽

Kun Han ◽

Lei Zhou ◽

...

Keyword(s):

Colorectal Cancer ◽

Stem Cells ◽

Protein Level ◽

Drug Target ◽

Target Genes ◽

Xenograft Model ◽

Luciferase Reporter ◽

Hippo Signaling ◽

Self Renewal ◽

Underlying Mechanisms

Abstract Background: Cancer stem cells (CSCs) have been recognized as an important drug target, however, the underlying mechanisms have not been fully understood. SKP1 is a traditional drug target for cancer therapy, while, whether SKP1 promotes colorectal cancer (CRC) stem cells (CRC-SCs) and the underlying mechanisms have remained elusive. Methods: Human CRC cell lines and primary human CRC cells were used in this study. Gene manipulation was performed by lentivirus system. The mRNA and protein levels of target genes were examined by qRT-PCR and western blot. The sphere-forming and in vitro migration capacities were determined by sphere formation and transwell assay. The self-renewal was determined by limiting dilution assay. The tumorigenicity and metastasis of cancer cells were examined by xenograft model. The promoter activity was examined by luciferase reporter assay. Nuclear run-on and Chromatin immunoprecipitation-PCR (ChIP-PCR) assay were employed to examine the transcription and protein-DNA interaction. Co-immunoprecipitation assay was used to test protein-protein interaction. The relationship between gene expression and survival was analyzed by Kaplan-meier analysis. The correlation between two genes was analyzed by Spearman analysis. Data are represented as mean ± s.d. and the significance was determined by Student’s t-test.Results: SKP1 was upregulated in CRC-SCs and predicted poor prognosis of colon cancer patients. Overexpression of SKP1 promoted the stemness of CRC cells reflected by increased sphere-forming, migration and self-renewal capacities as well as the expression of CSCs markers. In contrast, SKP1 depletion produced the opposite effects. SKP1 strengthened YAP activity and knockdown of YAP abolished the effect of SKP1 on the stemness of CRC cells. SKP1 suppressed RASSF1 at both mRNA and protein level. Overexpression of RASSF1 abolished the effect of SKP1 on YAP activity and CRC stemness. Conclusion: Our results demonstrated that SKP1 suppresses RASSF1 at both mRNA and protein level, attenuates Hippo signaling, activates YAP, and thereby promoting the stemness of CRC cells.

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