Nitazoxanide is a potent inhibitor of human seasonal coronaviruses acting at postentry level: effect on viral spike glycoprotein

Coronaviridae is recognized as one of the most rapidly evolving virus family as a consequence of the high genomic nucleotide substitution rates and recombination. The family comprises a large number of enveloped, positive-sense single-stranded RNA viruses, causing an array of diseases of varying severity in animals and humans. To date, seven human coronaviruses (HCoV) have been identified, namely HCoV-229E, HCoV-NL63, HCoV-OC43 and HCoV-HKU1, which are globally circulating in the human population (seasonal HCoV, sHCoV), and the highly pathogenic SARS-CoV, MERS-CoV and SARS-CoV-2. Seasonal HCoV are estimated to contribute to 15-30% of common cold cases in humans; although diseases are generally self-limiting, sHCoV can sometimes cause severe lower respiratory infections, as well as enteric and neurological diseases. No specific treatment is presently available for sHCoV infections. Herein we show that the anti-infective drug nitazoxanide has a potent antiviral activity against three human endemic coronaviruses, the Alpha-coronaviruses HCoV-229E and HCoV-NL63, and the Beta-coronavirus HCoV-OC43 in cell culture with IC50 ranging between 0.05 and 0.15 g/ml, and high selectivity indexes. We found that nitazoxanide does not affect HCoV adsorption, entry or uncoating, but acts at postentry level and interferes with the spike glycoprotein maturation, hampering its terminal glycosylation at an endoglycosidase H-sensitive stage. Altogether the results indicate that nitazoxanide, due to its broad-spectrum anti-coronavirus activity, may represent a readily available useful tool in the treatment of seasonal coronavirus infections.

The oral drug nitazoxanide restricts SARS-CoV-2 infection and attenuates disease pathogenesis in Syrian hamsters

A well-tolerated and cost-effective oral drug that blocks SARS-CoV-2 growth and dissemination would be a major advance in the global effort to reduce COVID-19 morbidity and mortality. Here, we show that the oral FDA-approved drug nitazoxanide (NTZ) significantly inhibits SARS-CoV-2 viral replication and infection in different primate and human cell models including stem cell-derived human alveolar epithelial type 2 cells. Furthermore, NTZ synergizes with remdesivir, and it broadly inhibits growth of SARS-CoV-2 variants B.1.351 (beta), P.1 (gamma), and B.1617.2 (delta) and viral syncytia formation driven by their spike proteins. Strikingly, oral NTZ treatment of Syrian hamsters significantly inhibits SARS-CoV-2-driven weight loss, inflammation, and viral dissemination and syncytia formation in the lungs. These studies show that NTZ is a novel host-directed therapeutic that broadly inhibits SARS-CoV-2 dissemination and pathogenesis in human and hamster physiological models, which supports further testing and optimization of NTZ-based therapy for SARS-CoV-2 infection alone and in combination with antiviral drugs.

Impairment of SARS-CoV-2 spike glycoprotein maturation and fusion activity by the broad-spectrum anti-infective drug nitazoxanide

SARS-CoV-2, the causative agent of COVID-19, has caused an unprecedented global health crisis. The SARS-CoV-2 spike, a surface-anchored trimeric class-I fusion-glycoprotein essential for viral entry, represents a key target for developing vaccines and therapeutics capable of blocking virus invasion. The emergence of SARS-CoV-2 spike-variants that facilitate virus spread and may affect vaccine efficacy highlights the need to identify novel antiviral strategies for COVID-19 therapy. Here we demonstrate that nitazoxanide, an antiprotozoal agent with recognized broad-spectrum antiviral activity, interferes with SARS-CoV-2 spike biogenesis, hampering its maturation at an endoglycosidase H-sensitive stage. Engineering multiple SARS-CoV-2 variant-pseudoviruses and utilizing quantitative cell-cell fusion assays, we show that nitazoxanide-induced spike modifications hinder progeny virion infectivity as well as spike-driven pulmonary cell-cell fusion, a critical feature of COVID-19 pathology. Nitazoxanide, being equally effective against the ancestral SARS-CoV-2 Wuhan-spike and different emerging variants, including the Delta variant of concern, may represent a useful tool in the fight against COVID-19 infections.

Early treatment with nitazoxanide prevents worsening of mild and moderate COVID-19 and subsequent hospitalization

BackgroundThere is an urgent need for treatments to prevent the progression to severe COVID-19 and hospitalization. MethodsA randomized double-blind placebo-controlled clinical trial in 36 centers in the U.S. and Puerto Rico investigated the safety and effectiveness of oral nitazoxanide 600 mg twice daily for 5 days in outpatients with symptoms of mild or moderate COVID-19 enrolled within 72 hours of symptom onset. Key objectives were reduction of duration of symptoms (primary) and progression to severe illness (key secondary). Results1,092 subjects were enrolled, and 379 with laboratory-confirmed SARS-CoV-2 infection were analyzed. Overall, times to sustained clinical recovery were similar for the two arms. Nitazoxanide treatment was associated with an 85% reduction in the progression to severe COVID-19 (1/184, [0.5%] vs. 7/195, [3.6%], p=0.07) and 82% reduction in the rate of hospitalization, emergency room visit or death (1/184 [0.5%] vs. 6/195 [3.1%], p=0.12). In subjects with mild illness at baseline, treatment was also associated with a 3.1-day reduction in median time to sustained clinical recovery and a 5.2-day reduction in time to return to usual health. Nitazoxanide was safe and well tolerated. ConclusionsTreatment of mild or moderate COVID-19 with a five-day course of oral nitazoxanide was safe and well tolerated and was associated with an 85% reduction in the progression to severe illness and a 3- to 5-day reduction of the duration of mild illness.