N6-Methyladenosine-Mediated Up-Regulation of FZD10 Regulates Liver Cancer Stem Cells' Properties and Lenvatinib Resistance Through WNT/b-Catenin and Hippo Signaling Pathways

BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide, but there is a deficiency of early diagnosis biomarkers and thera-peutic targets. Drug resistance accounts for most HCC-related deaths, yet the mechanisms underlying drug resistance remain poorly understood. METHODS: Expression of Frizzled -10 (FZD10) in liver cancer stem cells (CSCs) was identified by means of RNA sequencing and validated by means of real-time polymerase chain reaction and immunohistochemistry. In vitro and in vivo experiments were used to assess the effect of FZD10 on liver CSC expansion and lenvatinib resistance. RNA sequencing, RNA binding protein immunoprecipitation, and luciferase report assays were applied to explore the mechanism underlying FZD10-mediated liver CSCs expansion and lenvati-nib resistance. RESULTS: Activation of FZD10 in liver CSCs was mediated by METTL3-dependent N6-methyladenosine methyl-ation of FZD10 messenger RNA. Functional studies revealed that FZD10 promotes self-renewal, tumorigenicity, and metas-tasis of liver CSCs via activating b-catenin and YAP1. The FZD10-b-catenin/YAP1 axis is activated in liver CSCs and predicts poor prognosis. Moreover, FZD10-b-catenin/c-Jun axis transcriptionally activates METTL3 expression, forming a pos-itive feedback loop. Importantly, the FZD10/b-catenin/c-Jun/ MEK/ERK axis determines the responses of hepatoma cells to lenvatinib treatment. Analysis of patient cohort, patient-derived tumor organoids, and patient-derived xenografts further suggest that FZD10 might predict lenvatinib clinical benefit in patients with HCC. Furthermore, treatment of lenvatinib-resistant HCC with adeno-associated virus targeting FZD10 or a b-catenin inhibitor restored lenvatinib response. CONCLUSIONS: Elevated FZD10 expression promotes expan-sion of liver CSCs and lenvatinib resistance, indicating that FZD10 expression is a novel prognostic biomarker and thera-peutic target for human HCC.

Automated summary

This study reveals that the expression of Frizzled-10 (FZD10) in liver cancer stem cells is regulated by METTL3-mediated N6-methyladenosine methylation of FZD10 mRNA, and FZD10 promotes self-renewal, tumorigenicity, and metastasis of liver CSCs via activating β-catenin and YAP1. The FZD10-β-catenin/YAP1 axis is activated in liver CSCs and indicates poor prognosis. Furthermore, the FZD10/β-catenin/c-Jun/MEK/ERK axis determines the responses of hepatoma cells to lenvatinib treatment.