Mechanisms of action of repurposed Ebola virus antivirals – the roles of phospholipidosis and cholesterol homeostasis

Abstract

Cell-based drug repurposing screens have been a common approach to identifying compounds with antiviral properties. For Ebola virus (EBOV), such screens yield unexpectedly high hit rates. We investigated two mechanisms underlying the anti-EBOV activities of repurposed compounds. Phospholipidosis (PLD) is excessive accumulation of cellular lipids that confounds screens for SARS-CoV-2. We performed a meta-analysis of published screens and supplemented these with our own using infectious EBOV at biosafety level-4. A list of nearly 400 hit compounds from seven anti-EBOV screens was compiled. Most (63 %) of these hits were predicted to induce PLD, and their anti-EBOV activities broadly correlated with PLD induction. PLD-inducing compounds did not inhibit infection by several other highly pathogenic viruses, suggesting that PLD was not a confounding factor for screens against Lassa, Crimean-Congo hemorrhagic fever, and Rift Valley fever viruses. Of four cells lines tested, HeLa cells were the least susceptible to PLD induction. In addition to PLD, many of the hit compounds identified disrupt cholesterol homeostasis. Previous research found inhibition of cholesterol synthesis by statins blocked EBOV infection. To understand if compounds inhibiting this mechanism could contribute to high hit rates, we further examined this pathway. We identified multiple additional inhibitors of cholesterol biosynthesis, that also blocked EBOV infection, albeit with varying potency and cytotoxicity across cell lines. EBOV inhibitors that acted through this mechanism were suppressed by the addition of exogenous cholesterol. Our findings help define the effects that contribute to anti-EBOV activities and hence facilitate the selection of lead molecules suitable for subsequent development.