Small-Molecule Antiviral β- <scp>d</scp> - <i>N</i> ⁴ -Hydroxycytidine Inhibits a Proofreading-Intact Coronavirus with a High Genetic Barrier to Resistance

TLDR

Coronaviruses have emerged from animal reservoirs with epidemic and pandemic potential, yet no approved antivirals exist. The study investigates the potent antiviral activity of the broad‑spectrum ribonucleoside analogue β‑d‑N⁴‑hydroxycytidine (NHC) against two divergent coronaviruses. The authors evaluate NHC’s efficacy using in vitro assays on these two divergent CoVs. NHC remains effective despite viral proofreading, induces only low‑level resistance via deleterious transition mutations, and appears to inhibit coronaviruses through a mutagenic mechanism, supporting its further development.

Abstract

The emergence of coronaviruses (CoVs) into human populations from animal reservoirs has demonstrated their epidemic capability, pandemic potential, and ability to cause severe disease. However, no antivirals have been approved to treat these infections. Here, we demonstrate the potent antiviral activity of a broad-spectrum ribonucleoside analogue, β- d - N 4 -hydroxycytidine (NHC), against two divergent CoVs. Viral proofreading activity does not markedly impact sensitivity to NHC inhibition, suggesting a novel interaction between a nucleoside analogue inhibitor and the CoV replicase. Further, passage in the presence of NHC generates only low-level resistance, likely due to the accumulation of multiple potentially deleterious transition mutations. Together, these data support a mutagenic mechanism of inhibition by NHC and further support the development of NHC for treatment of CoV infections.