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Stabilizing and upregulating Axin with tankyrase inhibitor reverses 5-fluorouracil chemoresistance and proliferation by targeting the WNT/caveolin-1 axis in colorectal cancer cells

Abstract

Chemoresistance is a main obstacle for colorectal cancer treatment. In this study, we evaluated the effects and mechanisms of the WNT/β-catenin signaling pathway on the chemoresistance of SW480 and SW620 colorectal cancer cells. The activity of β-catenin was activated/inhibited by the small molecule compound GSK-3 inhibitor 6-bromo-indirubin-3’-oxime and the tankyrase inhibitor XAV939. The downstream target genes of the WNT/β-catenin signaling pathway were screened using a cDNA microarray and bioinformatics analysis. Apoptosis induced by 5-Fu, cell cycle distribution and expression levels of WNT/β-catenin/TCF12/caveolin-1 and multidrug resistance proteins were examed by flow cytometry and western blot after β-catenin activation/inhibition and caveolin-1 overexpression/interference. The effect and mechanism of XAV939 on proliferation and apoptosis induced by 5-Fu in xenograft tumors of nude mice were evaluated by immunohistochemistry and TUNEL staining. 6-Bromo-indirubin-3’-oxime treatment increased β-catenin expression by regulating GSK-3β phosphorylation, accompanied by upregulation of TCF12, caveolin-1, P-gp, and MRP2 and downregulation of apoptosis induced by 5-Fu. Conversely, XAV939 treatment decreased β-catenin expression by upregulating Axin, accompanied by downregulation of TCF12, Caveolin-1, P-gp, and MRP2 and upregulation of apoptosis induced by 5-Fu. The caveolin-1 gene was identified as an important downstream gene of the WNT/β-catenin signaling pathway. Caveolin-1 overexpression upregulated β-catenin expression, increased P-gp and MRP2 expression and decreased apoptosis induced by 5-Fu; conversely, caveolin-1 interference caused the opposite effects. In addition, in vivo experiments showed that XAV939 treatment reduced β-catenin expression, increased apoptosis induced by 5-Fu and repressed xenograft tumor growth. Our findings suggested that inhibition of WNT/β-catenin/TCF12/caveolin-1 provides a new promising therapeutic strategy for colorectal cancer treatment.

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Fig. 1: Functional annotations of the differentially expressed genes after treatment of SW480 and SW620 cells with BIO/XAV939 based on GenCLiP 2.0 online tool analysis.
Fig. 2: Expression levels of P-gp, MRP2, GSK-3β, p-GSK-3β, β-catenin, TCF-12, Cav-1 and Axin in SW480 and SW620 cells after treatment with BIO, XAV939 and 5-Fu for 24 h.
Fig. 3: Expression levels of TCF-12 and β-catenin in SW480 and SW620 cells after treatment with BIO/XAV939 or transfection with Si-TCF12 for 24 h.
Fig. 4: Expression levels of TCF12, β-catenin, P-gp, MRP2 and Cav-1 in SW480 and SW620 cells after transfection with Cav-1/Si-Cav-1 for 24 h.
Fig. 5: Effect of BIO, XAV939 and Cav-1 on 5-Fu-induced proliferation and apoptosis in SW480 and SW620 cells.
Fig. 6: Effects of BIO, XAV939 and Cav-1 on the cell cycle distribution of SW480 and SW620 cells.
Fig. 7: Effect and mechanism of XAV939 on chemotherapy in nude mice.
Fig. 8: BIO treatment regulate the translational activity of Cav-1 promoter.

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Data availability

Data generated for the current study are available from the corresponding author on reasonable request.

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Funding

This study was sponsored by Guangdong Natural Science Foundation (no.2014A030307007, no.2017A030307005).

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L-KP, LF, and L-JB designed the experiments. LF and L-JH conducted the experiments. L-KP, LF and L-JB participated in the data analysis. L-KP, LF and L-JB drafted and polished the manuscript. All authors read and approved the final manuscript.

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Correspondence to Kunping Liu.

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The authors declare no competing interests.

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All studies involving animals were performed following the National Guides for the Care and Use of Laboratory Animals and approved by the Institutional Animal Care and Use Committee of Guangzhou Medical University.

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Luo, F., Li, J., Liu, J. et al. Stabilizing and upregulating Axin with tankyrase inhibitor reverses 5-fluorouracil chemoresistance and proliferation by targeting the WNT/caveolin-1 axis in colorectal cancer cells. Cancer Gene Ther 29, 1707–1719 (2022). https://doi.org/10.1038/s41417-022-00493-y

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