Objective Hepatitis D virus (HDV) is a circular RNA virus coinfecting hepatocytes with hepatitis B virus. Chronic hepatitis D results in severe liver disease and an increased risk of liver cancer. Efficient therapeutic approaches against HDV are absent.
Design Here, we combined an RNAi loss-of-function and small molecule screen to uncover host-dependency factors for HDV infection.
Results Functional screening unravelled the hypoxia-inducible factor (HIF)-signalling and insulin-resistance pathways, RNA polymerase II, glycosaminoglycan biosynthesis and the pyrimidine metabolism as virus-hepatocyte dependency networks. Validation studies in primary human hepatocytes identified the carbamoyl-phosphatesynthetase 2, aspartate transcarbamylase and dihydroorotase (CAD) enzyme and estrogen receptor alpha (encoded by ESR1) as key host factors for HDV life cycle. Mechanistic studies revealed that the two host factors are required for viral replication. Inhibition studies using N-(phosphonoacetyl)-L-aspartic acid and fulvestrant, specific CAD and ESR1 inhibitors, respectively, uncovered their impact as antiviral targets.
Conclusion The discovery of HDV host-dependency factors elucidates the pathogenesis of viral disease biology and opens therapeutic strategies for HDV cure.
- antiviral therapy
- hepatitis D
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Contributors TB initiated the study. TB and ERV designed and supervised research. ERV, AW, LB and TB set up, designed and performed the siRNA and small molecule screens. TG and PPa produced the sparfosic acid. ERV, CB, LH, VT-L and EC performed the validation experiments. CSu performed the HDV northern blots. PPe provided human hepatocytes. ERV, AW, CB, LH, EC, MBZ, CSu, CSc, LB and TB analysed the data. ERV, HES and AK performed the bioinformatical analyses of the screens. EV and TB wrote the manuscript. All the authors approved the study.
Funding This work was supported by Inserm, the University of Strasbourg, the European Union (Infect-ERA hepBccc, ERC-2014-AdG-671231-HEPCIR and Horizon 2020 research and innovation programme under grant agreement 667273 - HEPCAR), Agence Nationale de Recherches sur le Sida et les Hépatites Virales (ANRS 15/1099) and the French Cancer Agency (ARC IHU201301187). This work has been published under the framework of the LabEx ANR-10-LAB-28 and benefits from a funding from the state managed by the French National Research Agency as part of the Investments for the Future (Investissements d’Avenir) programme. ERV is the recipient of an ANRS fellowship (ECTZ50121).
Competing interests None declared.
Ethics approval Protocols were approved by the local Ethics Committee of the Strasbourg University Hospitals (CPP) and the Ministry of Higher Education and Research of France (DC 2016 2616).
Provenance and peer review Not commissioned; externally peer reviewed.
Data sharing statement The datasets generated in this study, including the results from both RNAi and small molecule primary screens, are available within online supplementary material files. The rest of the data are available from the corresponding authors on reasonable request.
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