VX-509 attenuates the stemness characteristics of colorectal cancer stem-like cells by regulating the epithelial-mesenchymal transition through Nodal/Smad2/3 signaling

BACKGROUND

Colorectal cancer stem cells (CCSCs) are heterogeneous cells that can self-renew and undergo multidirectional differentiation in colorectal cancer (CRC) patients. CCSCs are generally accepted to be important sources of CRC and are responsible for the progression, metastasis, and therapeutic resistance of CRC. Therefore, targeting this specific subpopulation has been recognized as a promising strategy for overcoming CRC.

AIM

To investigate the effect of VX-509 on CCSCs and elucidate the underlying mechanism.

METHODS

CCSCs were enriched from CRC cell lines by in conditioned serum-free medium. Western blot, Aldefluor, transwell and tumorigenesis assays were performed to verify the phenotypic characteristics of the CCSCs. The anticancer efficacy of VX-509 was assessed in HCT116 CCSCs and HT29 CCSCs by performing cell viability analysis, colony formation, sphere formation, flow cytometry, and western blotting assessments in vitro and tumor growth, immunohistochemistry and immunofluorescence assessments in vivo.

RESULTS

Compared with parental cells, sphere cells derived from HCT116 and HT29 cells presented increased expression of stem cell transcription factors and stem cell markers and were more potent at promoting migration and tumorigenesis, demonstrating that the CRC sphere cells displayed CSC features. VX-509 inhibited the tumor malignant biological behavior of CRC-stem-like cells, as indicated by their proliferation, migration and clonality in vitro, and suppressed the tumor of CCSC-derived xenograft tumors in vivo. Besides, VX-509 suppressed the CSC characteristics of CRC-stem-like cells and inhibited the progression of epithelial-mesenchymal transition (EMT) signaling in vitro. Nodal was identified as the regulatory factor of VX-509 on CRC stem-like cells through analyses of differentially expressed genes and CSC-related database information. VX-509 markedly downregulated the expression of Nodal and its downstream phosphorylated Smad2/3 to inhibit EMT progression. Moreover, VX-509 reversed the dedifferentiation of CCSCs and inhibited the progression of EMT induced by Nodal overexpression.

CONCLUSION

VX-509 prevents the EMT process in CCSCs by inhibiting the transcription and protein expression of Nodal, and inhibits the dedifferentiated self-renewal of CCSCs.

Key Words: Colorectal cancer stem cells, Stemness, VX-509, Epithelial-mesenchymal transition, Nodal

Core Tip: Colorectal cancer (CRC) is a malignant cancer of the digestive tract with high recurrence and metastasis. CRC stem cells (CCSCs) exerted self-renewing and repopulating capacity, which are hard to eliminate and cause recurrence and metastasis of CRC. VX-509 has been reported as a rheumatoid arthritis regular treatment. However, the role of VX-509 in CCSCs is not well studied. In this study, VX-509 inhibited the malignant biological behaviors and stemness of CCSCs. Further experiments suggested that VX-509 downregulated the expression of Nodal to suppress the epithelial-mesenchymal transition and inhibit the stemness of CCSCs, which provided a potent therapy measure for CRC.

• Citation: Yuan Y, Zhang XF, Li YC, Chen HQ, Wen T, Zheng JL, Zhao ZY, Hu QY. VX-509 attenuates the stemness characteristics of colorectal cancer stem-like cells by regulating the epithelial-mesenchymal transition through Nodal/Smad2/3 signaling. World J Stem Cells 2024; 16(2): 207-227
• URL: https://www.wjgnet.com/1948-0210/full/v16/i2/207.htm
• DOI: https://dx.doi.org/10.4252/wjsc.v16.i2.207