M6A RNA methylation-mediated HNF3\u03b3 reduction renders hepatocellular carcinoma dedifferentiation and sorafenib resistance

Tengfei Zhou,Xijun Liang,Cheng Zhang,Junyu Liu,Xiao Li,Jin Ding,Hongyang Wang,Weiqi Jiang,Liu Hui,Liang Li,Xiuliang Cui,Xin Cheng,Beifang Ning,Daimin Xiang,Wen Sun,Zhuo Cheng,Shichao Li

doi:10.1038/s41392-020-00299-0

Abstract

Hepatocyte nuclear factor 3γ (HNF3γ) is a hepatocyte nuclear factor, but its role and clinical significance in hepatocellular carcinoma (HCC) remain unclear. Herein, we report that HNF3γ expression is downregulated in patient HCC and inversely correlated with HCC malignancy and patient survival. Moreover, our data suggested that the HNF3γ reduction in HCC could be mediated by METTL14-dependent m6A methylation of HNF3γ mRNA. HNF3γ expression was increased during hepatic differentiation and decreased in dedifferentiated HCC cells. Interestingly, HNF3γ delivery promoted differentiation of not only HCC cells but also liver CSCs, which led to suppression of HCC growth. Mechanistic analysis suggested an HNF3γ-centered regulatory network that includes essential liver differentiation-associated transcription factors and functional molecules, which could synergistically facilitate HCC cell differentiation. More importantly, enforced HNF3γ expression sensitized HCC cells to sorafenib-induced growth inhibition and cell apoptosis through transactivation of OATP1B1 and OATP1B3 expression, which are major membrane transporters for sorafenib uptake. Clinical investigation showed that patient-derived HCC xenografts with high HNF3γ expression exhibited a sorafenib response and patients with high HCC HNF3γ levels benefited from sorafenib therapy. Together, these results suggest that HNF3γ plays an essential role in HCC differentiation and may serve as a therapeutic target and predictor of sorafenib benefit in patients.

Highlights

Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies in the world, and ~700,000 new cases are diagnosed annually.[1]
We examined the expression of Hepatocyte nuclear factor 3γ (HNF3γ) in fetal liver, adult liver, and HCC tissues and found that HNF3γ expression was significantly activated in mature normal liver tissues (Supplementary Fig. S4b, c), suggesting that HNF3γ expression might be associated with the differentiation status of hepatocytes
Considering the distinguished advantage of adeno-associated virus (AAV) in gene therapy, we further tested the therapeutic effects of AAV-HNF3γ using patient-derived xenograft (PDX) models, and the results showed that AAV-mediated HNF3γ delivery significantly inhibited the HCC progression (Fig. 6f), suggesting that HNF3γ could be an optimized target in HCC differentiation therapy

Summary

1234567890();,: INTRODUCTION

Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies in the world, and ~700,000 new cases are diagnosed annually.[1]. To identify the m6A reader required in m6A modification-regulated HNF3γ mRNA stability, we certain pro-differentiation conditions, we investigated whether HNF3γ could induce the differentiation of liver CSCs. decreased HNF3γ expression was observed in EpCAM+ or CD133+ HCC cells compared with EpCAM− or CD133− cells, suggesting that HNF3γ expression was further reduced in liver CSCs. Enforced HNF3γ expression suppressed the expression of stemness-associated factors and liver CSC markers in HCC spheres knocked down the m6A readers involved in RNA stability in HCC cells respectively.[18] Our data showed that the interference of (Fig. 5b) as well as HCC cells (Supplementary Fig. S6a, b). PDXs with high HNF3γ levels displayed significant growth inhibition upon sorafenib treatment (Fig. 8e), while PDXs with low HNF3γ expression did not (Fig. 8f) These data suggest that HNF3γ expression is correlated with sorafenib response and could be a novel predictor of sorafenib benefit for HCC patients

DISCUSSION

MATERIALS AND METHODS

Findings

13 AUTHOR CONTRIBUTIONS