New host target identified to block hepatitis E virus

A new host target for hepatitis E

Hepatitis E virus (HEV) is a major cause of acute viral hepatitis, yet antiviral treatment remains largely limited to the off-label use of ribavirin. Ribavirin can cause substantial toxicity, is contraindicated during pregnancy and does not reliably benefit every patient. Seeking alternative treatments, Ring and colleagues screened compounds that interfere with endocytic and endolysosomal pathways-the intracellular transport system used by HEV during cell entry.

Among the compounds tested, apilimod emerged as the most potent inhibitor. Apilimod targets PIKfyve, a phosphoinositide kinase that regulates late-endosome and lysosome organisation. Apilimod, raloxifene, verapamil, chloroquine and tamoxifen all inhibited HEV infection dose-dependently, but apilimod demonstrated the strongest activity, reaching nanomolar potency in HepG2/C3A cells.

PIKfyve is required for viral entry

The researchers confirmed that the antiviral effect was linked to PIKfyve rather than an unrelated action of apilimod. Silencing PIKfyve using small interfering RNA reduced HEV infection by approximately 40%, while two additional PIKfyve inhibitors, YM201636 and APY0201, also suppressed infection.

The effect was observed against both quasi-enveloped and non-enveloped viral particles and across different HEV strains, indicating that dependence on PIKfyve may be a broadly conserved feature of HEV infection.

Time-of-addition and replicon experiments further identified the affected stage of the viral life cycle. PIKfyve inhibitors remained effective when administered around the early hours after viral inoculation but did not inhibit HEV RNA replication when cell entry was experimentally bypassed. Thus, apilimod primarily acts as an entry inhibitor rather than directly targeting the viral replication machinery.

Disrupting the lysosomal route used by HEV

Mechanistic experiments showed that PIKfyve inhibition enlarged lysosomes, reduced LAMP1- and CD63-positive compartments, and impaired the delivery of incoming HEV genomes to late-endosomal and lysosomal vesicles.

Apilimod also reduced the maturation and activity of cathepsins B and L. These lysosomal proteases are thought to process the HEV capsid and facilitate viral genome release into the cytoplasm. The study therefore connects PIKfyve-controlled vesicular organisation with protease-dependent viral uncoating, providing a coherent mechanistic explanation for the blockade of HEV entry.

Evidence in human hepatocytes and animals

Importantly, apilimod reduced HEV infection in primary human hepatocytes without a corresponding loss of cell viability, although effective concentrations were higher than those required in hepatoma cell lines.

The compound was also tested in rats challenged with ratHEV. Animals received apilimod at 75 mg/kg once daily for 12 days. Treatment significantly altered the trajectory of faecal viral shedding and reduced viral RNA levels in several sampled tissues. However, the reduction in the overall area under the curve for faecal viral burden did not reach conventional statistical significance, highlighting the preliminary nature of the in vivo evidence.

Translational promise-and remaining questions

This study positions PIKfyve as a druggable host dependency factor for HEV and raises the possibility of repurposing apilimod, which has previously undergone clinical testing for other indications. Targeting a host pathway may also impose a higher barrier to viral resistance than therapies directed against a viral protein.

Nevertheless, the findings remain preclinical. RatHEV infection does not fully reproduce human HEV disease, and PIKfyve has important physiological functions in endolysosomal homeostasis. Future studies should establish appropriate therapeutic exposure, treatment timing, tissue-specific toxicity, activity against additional human HEV genotypes and efficacy in chronic or established infection. Clinical development will ultimately require evidence that antiviral concentrations can be achieved safely in patients.