Hepatitis E virus (HEV) is an underestimated disease, leading to estimated 20 million infections and up to 70,000 deaths annually. Infections are mostly asymptomatic but can reach mortality rates up to 25% in pregnant women or become chronic in immunocompromised patients. The current therapy options are limited to the unspecific antivirals Ribavirin (RBV) and pegylated Interferon-α (pegIFN-α). RBV leads to viral clearance in only 80% of patients treated, and is, similar to pegIFN-α, contraindicated in the major risk group of pregnant women, emphasizing the importance of new therapy options. In this review, we focus on the urgent need and current efforts in HEV drug development. We provide an overview of the current status of HEV antiviral research. Furthermore, we discuss strategies for drug development and the limitations of the approaches with respect to HEV.

Background and Aims: Being the most common cause of acute viral hepatitis with >20 million cases per year and 70,000 deaths annually, HEV presents a long-neglected and underinvestigated health burden. Although the entry process of viral particles is an attractive target for pharmacological intervention, druggable host factors to restrict HEV entry have not been identified so far. Approach and Results: Here we identify the EGF receptor (EGFR) as a novel host factor for HEV and reveal the significance of EGFR for the HEV entry process. By utilizing RNAi, chemical modulation with Food and Drug Administration–approved drugs, and ectopic expression of EGFR, we revealed that EGFR is critical for HEV infection without affecting HEV RNA replication or assembly of progeny virus. We further unveiled that EGFR itself and its ligand-binding domain, rather than its signaling function, is responsible for the proviral effect. Modulation of EGF expression in HepaRG cells and primary human hepatocytes affected HEV infection. Conclusions: Taken together, our study provides novel insights into the life cycle of HEV and identified EGFR as a possible target for future antiviral strategies against HEV.

Background and Aims: HEV is estimated to be responsible for 70,000 deaths annually, yet therapy options remain limited. In the pursuit of effective antiviral therapies, targeting viral entry holds promise and has proven effective for other viruses. However, the precise mechanisms and host factors required during HEV entry remain unclear. Cellular proteases have emerged as host factors required for viral surface protein activation and productive cell entry by many viruses. Hence, we investigated the functional requirement and therapeutic potential of cellular protease during HEV infection. Approach and Results: Using our established HEV cell culture model and subgenomic HEV replicons, we found that blocking lysosomal cathepsins (CTS) with small molecule inhibitors impedes HEV infection without affecting replication. Most importantly, the pan-cathepsin inhibitor K11777 suppressed HEV infections with an EC₅₀ of ~0.02 nM. Inhibition by K11777, devoid of notable toxicity in hepatoma cells, was also observed in HepaRG and primary human hepatocytes. Furthermore, through time-of-addition and RNAscope experiments, we confirmed that HEV entry is blocked by inhibition of cathepsins. Cathepsin L (CTSL) knockout cells were less permissive to HEV, suggesting that CTSL is critical for HEV infection. Finally, we observed cleavage of the glycosylated ORF2 protein and virus particles by recombinant CTSL. Conclusions: In summary, our study highlights the pivotal role of lysosomal cathepsins, especially CTSL, in the HEV entry process. The profound anti-HEV efficacy of the pan-cathepsin inhibitor K11777, especially with its notable safety profile in primary cells, further underscores its potential as a therapeutic candidate.

Background: Hepatitis C virus (HCV) constitutes a global health problem, while hepatitis E virus (HEV) is the major cause of acute viral hepatitis globally. HCV/HEV co-infections have been poorly characterized, as they are hampered by the lack of robust HEV cell culture systems. This study developed experimental models to study HCV/HEV co-infections and investigate viral interference in cells and humanized mice. Methods: We used state-of-the art human hepatocytes tissue culture models to assess HEV and HCV replication in co- or super-transfection settings. Findings were confirmed by co- and super-infection experiments in human hepatocytes and in vivo in human liver chimeric mice. Results: HEV was inhibited by concurrent HCV replication in human hepatocytes. This exclusion phenotype was linked to the protease activity of HCV. These findings were corroborated by the fact that in HEV on HCV super-infected mice, HEV viral loads were reduced in individual mice. Similarly, HCV on HEV super-infected mice showed reduced HCV viral loads. Conclusion: Direct interference of both viruses with HCV NS3/4A as the determinant was observed. In vivo, we detected reduced replication of both viruses after super-infection in individual mice. These findings provide new insights into the pathogenesis of HCV-HEV co-infections and should contribute to its clinical management in the future.