To combat emerging coronaviruses, developing safe and efficient platforms to evaluate viral
protease activities and the efficacy of
protease inhibitors is a high priority. Here, we exploit a biosafety level 2 (BSL-2) chimeric Sindbis virus system to evaluate
protease activities and the efficacy of inhibitors directed against the
papain-like
protease (PLpro) of severe acute respiratory syndrome coronavirus (SARS-CoV), a biosafety level 3 (BSL-3) pathogen. We engineered Sindbis virus to coexpress PLpro and a substrate, murine
interferon-stimulated gene 15 (ISG15), and found that PLpro mediates removal of ISG15 (deISGylation) from cellular
proteins. Mutation of the catalytic
cysteine residue of PLpro or addition of a PLpro inhibitor blocked deISGylation in virus-infected cells. Thus, deISGylation is a marker of PLpro activity.
Infection of alpha/
beta interferon receptor knockout (IFNAR(-/-)) mice with these chimeric viruses revealed that PLpro deISGylation activity removed ISG15-mediated protection during
viral infection. Importantly, administration of a PLpro inhibitor protected these mice from lethal
infection, demonstrating the efficacy of a coronavirus
protease inhibitor in a mouse model. However, this PLpro inhibitor was not sufficient to protect the mice from lethal
infection with SARS-CoV MA15, suggesting that further optimization of the delivery and stability of PLpro inhibitors is needed. We extended the chimeric-virus platform to evaluate the
papain-like
protease/deISGylating activity of Middle East respiratory syndrome coronavirus (MERS-CoV) to provide a small-animal model to evaluate PLpro inhibitors of this recently emerged pathogen. This platform has the potential to be universally adaptable to other viral and cellular
enzymes that have deISGylating activities. Importance: Evaluating
viral protease inhibitors in a small-animal model is a critical step in the path toward
antiviral drug development. We modified a biosafety level 2 chimeric virus system to facilitate evaluation of inhibitors directed against highly pathogenic coronaviruses. We used this system to demonstrate the in vivo efficacy of an inhibitor of the
papain-like
protease of severe acute respiratory syndrome coronavirus. Furthermore, we demonstrate that the chimeric-virus system can be adapted to study the
proteases of emerging human pathogens, such as Middle East respiratory syndrome coronavirus. This system provides an important tool to rapidly assess the efficacy of
protease inhibitors targeting existing and emerging human pathogens, as well as other
enzymes capable of removing ISG15 from cellular
proteins.