Drug combination therapy for emerging viral diseases.

Effective therapeutics to combat emerging viral infections are an unmet need. Historically, treatments for chronic viral infections with single drugs have not been successful, as exemplified by human immunodeficiency virus (HIV) and hepatitis C virus (HCV) infections. Combination therapy for these diseases has led to improved clinical outcomes with dramatic reductions in viral load, morbidity, and mortality. Drug combinations can enhance therapeutic efficacy through additive, and ideally synergistic, effects for emerging and re-emerging viruses, such as influenza, severe acute respiratory syndrome-coronavirus (SARS-CoV), Middle East respiratory syndrome (MERS)-CoV, Ebola, Zika, and SARS-coronavirus 2 (CoV-2). Although novel drug development through traditional pipelines remains a priority, in the interim, effective synergistic drug candidates could be rapidly identified by drug-repurposing screens, facilitating accelerated paths to clinical testing and potential emergency use authorizations.

Inhibiting SARS-CoV-2 infection with lysosomal alkalizers

Understanding the SARS-CoV-2 virus? routes of infection, virus?host?protein interactions, and mechanisms of virus-induced cytopathic effects will greatly aid in the discovery and design of new therapeutics to treat COVID-19. Chloroquine and hydroxychloroquine, extensively explored as clinical agents for COVID-19, have multiple cellular effects including alkalizing lysosomes and blocking autophagy as well as exhibiting dose-limiting toxicities in patients. To identify an alternative lysosome-based drug repurposing opportunity we evaluated additional lysosomotropic compounds . We found that six of these compounds blocked the cytopathic effect of SARS-CoV-2 in Vero E6 cells with half-maximal effective concentration (EC50) values ranging from 2.0 to 13 ?M and selectivity indices (SIs;SI = CC50/EC50) ranging from 1.5- to >10-fold. We demonstrate how the compounds (1) blocked lysosome functioning and autophagy, (2) prevented pseudotyped particle entry, (3) increased lysosomal pH, and (4) that ROC-325 reduced viral titers in the EpiAirway 3D tissue model. Consistent with these findings, the siRNA knockdown of ATP6V0D1 blocked the HCoV-NL63 cytopathic effect in LLC-MK2 cells. Moreover, an analysis of SARS-CoV-2 infected Vero E6 cell lysate revealed significant dysregulation of autophagy and lysosomal function, suggesting a contribution of the lysosome to the life cycle of SARS-CoV-2. Our findings support targeting the lysosome to combat SARS-CoV-2 infections and inhibitors of lysosomal function could become an important component of drug combination therapies aimed at improving treatment and outcomes for COVID-19.

The SARS-CoV-2 Cytopathic Effect Is Blocked by Lysosome Alkalizing Small Molecules.

Understanding the SARS-CoV-2 virus' pathways of infection, virus-host-protein interactions, and mechanisms of virus-induced cytopathic effects will greatly aid in the discovery and design of new therapeutics to treat COVID-19. Chloroquine and hydroxychloroquine, extensively explored as clinical agents for COVID-19, have multiple cellular effects including alkalizing lysosomes and blocking autophagy as well as exhibiting dose-limiting toxicities in patients. Therefore, we evaluated additional lysosomotropic compounds to identify an alternative lysosome-based drug repurposing opportunity. We found that six of these compounds blocked the cytopathic effect of SARS-CoV-2 in Vero E6 cells with half-maximal effective concentration (EC50) values ranging from 2.0 to 13 µM and selectivity indices (SIs; SI = CC50/EC50) ranging from 1.5- to >10-fold. The compounds (1) blocked lysosome functioning and autophagy, (2) prevented pseudotyped particle entry, (3) increased lysosomal pH, and (4) reduced (ROC-325) viral titers in the EpiAirway 3D tissue model. Consistent with these findings, the siRNA knockdown of ATP6V0D1 blocked the HCoV-NL63 cytopathic effect in LLC-MK2 cells. Moreover, an analysis of SARS-CoV-2 infected Vero E6 cell lysate revealed significant dysregulation of autophagy and lysosomal function, suggesting a contribution of the lysosome to the life cycle of SARS-CoV-2. Our findings suggest the lysosome as a potential host cell target to combat SARS-CoV-2 infections and inhibitors of lysosomal function could become an important component of drug combination therapies aimed at improving treatment and outcomes for COVID-19.