Broad-spectrum in vitro antiviral activity of ODBG-P-RVn: an orally-available, lipid-modified monophosphate prodrug of remdesivir parent nucleoside (GS-441524)

Abstract

The intravenous administration of remdesivir for COVID-19 confines its utility to hospitalized patients. We evaluated the broad-spectrum antiviral activity of ODBG-P-RVn, an orally available, lipid-modified monophosphate prodrug of the remdesivir parent nucleoside (GS-441524) against viruses that cause diseases of human public health concern, including SARS-CoV-2. ODBG-P-RVn showed 20-fold greater antiviral activity than GS-441524 and had near-equivalent activity to remdesivir in primary-like human small airway epithelial cells. Our results warrant investigation of ODBG-P-RVn efficacy in vivo.

Figure 1.

Comparison of antiviral activities of RVn, RDV, and ODBG-P-RVn in African green monkey (Vero-E6), human hepatoma (Huh7), and human bronchioalveolar carcinoma (NCI-H358) cell lines using reporter-based, image-based, and cytopathic effect (CPE) assays. Representative dose-response inhibition of viral replication and induction of cellular cytotoxicity by RVn (blue shapes), RDV (black shapes), and ODBG-P-RVn (red shapes). A) Direct measurement of reporter fluorescence intensity by recombinant Ebola virus (EBOV) expressing ZsGreen protein in Vero-E6 (left panel) and Huh7 (middle panel) cells, and recombinant Nipah virus (NiV) expressing ZsGreen protein in NCI-H358 (right panel) cells. B) Image-based counting of reporter fluorescence-positive cells infected with recombinant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) expressing mNeonGreen protein (Vero-E6 and Huh7) and recombinant respiratory syncytial virus (RSV) expressing eGFP (NCI-H358). Infected cells treated with DMSO were considered as 100% fluorescence intensity signal and 100% fluorescence-positive cell counts. C) Compound-based inhibition of CPE induced by yellow fever virus (YFV) in Vero-E6 and Huh7 cells and by Hendra virus (HeV) in NCI-H358 cells determined by measuring cellular ATP levels (CellTiterGlo 2.0). ATP levels in uninfected cells treated with DMSO were considered 100% CPE inhibition. D) Compound cytotoxicity/cell viability measured by CellTiterGlo 2.0 assay. Dose-response curves were fitted to the mean value of experiments performed in biological triplicate for each concentration in the 8-point, 3-fold dilution series using a 4-parameter non-linear logistic regression curve with variable slope. Data points and error bars indicate the mean value and standard deviation of 3 biological replicates; each colored shape/line in the legend represents an independent experiment performed in biological triplicate. RVn and RDV used in this study was obtained from MedChemExpress (Monmouth Junction, NJ USA).

Figure 2.

Comparison of cell type-dependent antiviral activities of RVn, RDV, and ODBG-P-RVn in primary-like hTERT-immortalized microvascular endothelial (TIME) cells and small airway epithelial cells (HSAEC1-KT). A) Representative dose-response inhibition of recombinant EBOV, NiV, and Marburg virus (MARV) expressing ZsGreen protein in TIME cells. B) Yield reduction of infectious EBOV-ZsG (left panel) and NiV-ZsG (middle panel) by RDV and ODBG-P-RVn. Compound cytotoxicity/cell viability (right panel) in TIME cells measured via CellTiterGlo 2.0 assay. C) Representative dose-response inhibition of recombinant EBOV, NiV, and MARV expressing ZsGreen protein in HSAEC1-KT cells. D) Reduction of infectious yield of EBOV-ZsG (left panel) and NiV-ZsG (middle panel) by RDV and ODBG-P-RVn in HSAEC1-KT cells. Compound cytotoxicity/cell viability (right panel) in HSAEC1-KT cells measured via CellTiterGlo 2.0 assay. Dose-response curves were fitted to the mean value of experiments performed in biological triplicate for each concentration in the 8-point, 3-fold dilution series using a 4-parameter non-linear logistic regression curve with variable slope. Data points and error bars indicate the mean value and standard deviation of 3 or 4 biological replicates; each colored shape/line in the legend represents an independent experiment performed in biological triplicate. Infectious yield reduction assays were conducted once with biological quadruplicates.