Biology subjects

Froggatt, H. M.

Publications and source records attributed to Froggatt, H. M..

3 recordsLinked to original sources

bioRxiv2020

Drug repurposing screen identifies masitinib as a 3CLpro inhibitor that blocks replication of SARS-CoV-2 in vitro

There is an urgent need for anti-viral agents that treat SARS-CoV-2 infection. The shortest path to clinical use is repurposing of drugs that have an established safety profile in humans. Here, we first screened a library of 1,900 clinically safe drugs for inhibiting replication of OC43, a human beta-coronavirus that causes the common-cold and is a relative of SARS-CoV-2, and identified 108 effective drugs. We further evaluated the top 26 hits and determined their ability to inhibit SARS-CoV-2, as well as other pathogenic RNA viruses. 20 of the 26 drugs significantly inhibited SARS-CoV-2 replication in human lung cells (A549 epithelial cell line), with EC50 values ranging from 0.1 to 8 micromolar. We investigated the mechanism of action for these and found that masitinib, a drug originally developed as a tyrosine-kinase inhibitor for cancer treatment, strongly inhibited the activity of the SARS-CoV-2 main protease 3CLpro. X-ray crystallography revealed that masitinib directly binds to the active site of 3CLpro, thereby blocking its enzymatic activity. Mastinib also inhibited the related viral protease of picornaviruses and blocked picornaviruses replication. Thus, our results show that masitinib has broad anti-viral activity against two distinct beta-coronaviruses and multiple picornaviruses that cause human disease and is a strong candidate for clinical trials to treat SARS-CoV-2 infection.

microbiology↗

bioRxiv2020

Development of a fluorescence based, high-throughput SARS-CoV-2 3CLpro reporter assay

In late 2019 a human coronavirus, now known as SARS-CoV-2, emerged, likely from a zoonotic reservoir. This virus causes COVID-19 disease, has infected millions of people, and has led to hundreds of thousands of deaths across the globe. While the best interventions to control and ultimately stop the pandemic are prophylactic vaccines, antiviral therapeutics are important to limit morbidity and mortality in those already infected. At this time, only one FDA approved anti-SARS-CoV-2 antiviral drug, remdesivir, is available and unfortunately, its efficacy appears to be limited. Thus, the identification of new and efficacious antivirals is of highest importance. In order to facilitate rapid drug discovery, flexible, sensitive, and high-throughput screening methods are required. With respect to drug targets, most attention is focused on either the viral RNA-dependent RNA polymerase or the main viral protease, 3CLpro. 3CLpro is an attractive target for antiviral therapeutics as it is essential for processing newly translated viral proteins, and the viral lifecycle cannot be completed without protease activity. In this work, we present a new assay to identify inhibitors of the SARS-CoV-2 main protease, 3CLpro. Our reporter is based on a GFP-derived protein that only fluoresces after cleavage by 3CLpro. This experimentally optimized reporter assay allows for antiviral drug screening in human cell culture at biosafety level-2 (BSL2) with high-throughput compatible protocols. Using this screening approach in combination with existing drug libraries may lead to the rapid identification of novel antivirals to suppress SARS-CoV-2 replication and spread. IMPORTANCEThe COVID-19 pandemic has already led to more than 400,000 deaths and innumerable changes to daily life worldwide. Along with development of a vaccine, identification of effective antivirals to treat infected patients is of the highest importance. However, rapid drug discovery requires efficient methods to identify novel compounds that can inhibit the virus. In this work, we present a method for identifying inhibitors of the SARS-CoV-2 main protease, 3CLpro. This reporter-based assay allows for antiviral drug screening in human cell culture at biosafety level-2 (BSL2) with high-throughput compatible sample processing and analysis. This assay may help identify novel antivirals to control the COVID-19 pandemic.

microbiology↗

bioRxiv2019

ETV7 represses a subset of interferon-stimulated genes that restrict influenza viruses

The type I interferon (IFN) response is an important component of the innate immune response to viral infection. Precise control of interferon responses is critical; insufficient levels of interferon-stimulated genes (ISGs) can lead to a failure to restrict viral spread while excessive ISG activation can result in interferon-related pathologies. While both positive and negative regulatory factors control the magnitude and duration of IFN signaling, it is also appreciated that a number of ISGs regulate aspects of the interferon response themselves. However, the mechanisms underlying these ISG regulatory networks remain incompletely defined. In this study, we performed a CRISPR activation screen to identify new regulators of the type I IFN response. We identified ETS variant transcription factor 7 (ETV7), a strongly induced ISG, as a protein that acts as a negative regulator of the type I IFN response; however, ETV7 did not uniformly suppress ISG transcription. Instead, ETV7 preferentially targeted a subset of known antiviral ISGs. Further, we showed the subset of ETV7-modulated ISGs was particularly important for IFN-mediated control of some viruses including influenza viruses and SARS-CoV-2. Together, our data assign a function for ETV7 as an IFN response regulator and also identify ETV7 as a therapeutic target to increase innate responses and potentiate the efficacy of interferon-based antiviral therapies. One Sentence SummaryETV7 is an interferon-induced, repressive transcription factor that negatively regulates antiviral interferon-stimulated genes essential for controlling influenza virus and SARS-CoV-2 infections.

microbiology↗