Potent small molecule inhibitors against the 3C protease of foot-and-mouth disease virus.

Foot-and-mouth disease (FMD) is one of the most devastating diseases of livestock which can cause significant economic losses, especially when introduced to FMD-free countries. FMD virus (FMDV) belongs to the family and is antigenically heterogeneous with seven established serotypes. The prevailing preventive and control strategies are limited to restriction of animal movement and elimination of infected or exposed animals, which can be potentially combined with vaccination.

Potent 3CLpro inhibitors effective against SARS-CoV-2 and MERS-CoV in animal models by therapeutic treatment.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Middle East respiratory syndrome coronavirus (MERS-CoV) are zoonotic betacoronaviruses that continue to have a significant impact on public health. Timely development and introduction of vaccines and antivirals against SARS-CoV-2 into the clinic have substantially mitigated the burden of COVID-19. However, a limited or lacking therapeutic arsenal for SARS-CoV-2 and MERS-CoV infections, respectively, calls for an expanded and diversified portfolio of antivirals against these coronavirus infections.

The smallest functional antibody fragment: Ultralong CDR H3 antibody knob regions potently neutralize SARS-CoV-2.

Cows produce antibodies with a disulfide-bonded antigen-binding domain embedded within ultralong heavy chain third complementarity determining regions. This "knob" domain is analogous to natural cysteine-rich peptides such as knottins in that it is small and stable but can accommodate diverse loops and disulfide bonding patterns. We immunized cattle with SARS-CoV-2 spike and found ultralong CDR H3 antibodies that could neutralize several viral variants at picomolar IC potencies in vitro and could protect from disease in vivo.

Intestinal colonization with Escherichia fergusonii enhances infectivity of GII.12 human norovirus in gnotobiotic pigs.

The role of gut microbiota [especially, histo-blood group antigen (HBGA)-expressing bacteria] in influencing human norovirus (HuNoV) infections is unclear. We investigated if infectivity of GII.12 HuNoV in gnotobiotic (Gn) pigs is altered by intestinal colonization with Escherichia fergusonii known to express HBGA A and H on their cell surface.

Roles of host proteases in the entry of SARS-CoV-2.

The spike protein (S) of SARS-CoV-2 is responsible for viral attachment and entry, thus a major factor for host susceptibility, tissue tropism, virulence and pathogenicity. The S is divided with S1 and S2 region, and the S1 contains the receptor-binding domain (RBD), while the S2 contains the hydrophobic fusion domain for the entry into the host cell. Numerous host proteases have been implicated in the activation of SARS-CoV-2 S through various cleavage sites.

Structure-guided design of direct-acting antivirals that exploit the gem-dimethyl effect and potently inhibit 3CL proteases of severe acute respiratory syndrome Coronavirus-2 (SARS-CoV-2) and middle east respiratory syndrome coronavirus (MERS-CoV).

The high morbidity and mortality associated with SARS-CoV-2 infection, the etiological agent of COVID-19, has had a major impact on global public health. Significant progress has been made in the development of an array of vaccines and biologics, however, the emergence of SARS-CoV-2 variants and breakthrough infections are an ongoing major concern. Furthermore, there is an existing paucity of small-molecule host and virus-directed therapeutics and prophylactics that can be used to counter the spread of SARS-CoV-2, and any emerging and re-emerging coronaviruses.

Broad-Spectrum Cyclopropane-Based Inhibitors of Coronavirus 3C-like Proteases: Biochemical, Structural, and Virological Studies.

The advent of SARS-CoV-2, the causative agent of COVID-19, and its worldwide impact on global health, have provided the impetus for the development of effective countermeasures that can be deployed against the virus, including vaccines, monoclonal antibodies, and direct-acting antivirals (DAAs). Despite these efforts, the current paucity of DAAs has created an urgent need for the creation of an enhanced and diversified portfolio of broadly acting agents with different mechanisms of action that can effectively abrogate viral infection. SARS-CoV-2 3C-like protease (3CL), an enzyme essential for viral replication, is a validated target for the discovery of SARS-CoV-2 therapeutics.

SARS-CoV-2 Omicron (B.1.1.529) Infection of Wild White-Tailed Deer in New York City.

There is mounting evidence of SARS-CoV-2 spillover from humans into many domestic, companion, and wild animal species. Research indicates that humans have infected white-tailed deer, and that deer-to-deer transmission has occurred, indicating that deer could be a wildlife reservoir and a source of novel SARS-CoV-2 variants. We examined the hypothesis that the Omicron variant is actively and asymptomatically infecting the free-ranging deer of New York City.

Potent Protease Inhibitors of Highly Pathogenic Lagoviruses: Rabbit Hemorrhagic Disease Virus and European Brown Hare Syndrome Virus.

Rabbit hemorrhagic disease (RHD) and European brown hare syndrome (EBHS) are highly contagious diseases caused by lagoviruses in the family. These infectious diseases are associated with high mortality and a serious threat to domesticated and wild rabbits and hares, including endangered species such as riparian brush rabbits (). In the United States (U.

Effects of Spike Mutations in SARS-CoV-2 Variants of Concern on Human or Animal ACE2-Mediated Virus Entry and Neutralization.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a zoonotic agent capable of infecting humans and a wide range of animal species. Over the duration of the pandemic, mutations in the SARS-CoV-2 spike (S) protein have arisen, culminating in the spread of several variants of concern (VOCs) with various degrees of altered virulence, transmissibility, and neutralizing antibody escape. In this study, we used pseudoviruses that express specific SARS-CoV-2 S protein substitutions and cell lines that express angiotensin-converting enzyme 2 (ACE2) from nine different animal species to gain insights into the effects of VOC mutations on viral entry and antibody neutralization capability.

Structure-Guided Design of Potent Spirocyclic Inhibitors of Severe Acute Respiratory Syndrome Coronavirus-2 3C-like Protease.

The worldwide impact of the ongoing COVID-19 pandemic on public health has made imperative the discovery and development of direct-acting antivirals aimed at targeting viral and/or host targets. SARS-CoV-2 3C-like protease (3CL) has emerged as a validated target for the discovery of SARS-CoV-2 therapeutics because of the pivotal role it plays in viral replication. We describe herein the structure-guided design of highly potent inhibitors of SARS-CoV-2 3CL that incorporate in their structure novel spirocyclic design elements aimed at optimizing potency by accessing new chemical space.

Structure-Guided Design of Potent Inhibitors of SARS-CoV-2 3CL Protease: Structural, Biochemical, and Cell-Based Studies.

The COVID-19 pandemic is having a major impact on public health worldwide, and there is an urgent need for the creation of an armamentarium of effective therapeutics, including vaccines, biologics, and small-molecule therapeutics, to combat SARS-CoV-2 and emerging variants. Inspection of the virus life cycle reveals multiple viral- and host-based choke points that can be exploited to combat the virus. SARS-CoV-2 3C-like protease (3CLpro), an enzyme essential for viral replication, is an attractive target for therapeutic intervention, and the design of inhibitors of the protease may lead to the emergence of effective SARS-CoV-2-specific antivirals.

Effects of Spike Mutations in SARS-CoV-2 Variants of Concern on Human or Animal ACE2-Mediated Virus Entry and Neutralization.

Unlabelled: SARS-CoV-2 is a zoonotic agent capable of infecting humans and a wide range of animal species. Over the duration of the pandemic, mutations in the SARS-CoV-2 Spike protein (S) have arisen in circulating viral populations, culminating in the spread of several variants of concern (VOC) with varying degrees of altered virulence, transmissibility, and neutralizing antibody escape. In this study, we employed lentivirus-based pseudotyped viruses that express specific SARS-CoV-2 S protein substitutions and cell lines that stably express ACE2 from nine different animal species to gain insights into the effects of VOC mutations on viral entry and antibody neutralization capability.

Structure-Guided Design of Conformationally Constrained Cyclohexane Inhibitors of Severe Acute Respiratory Syndrome Coronavirus-2 3CL Protease.

A series of nondeuterated and deuterated dipeptidyl aldehyde and masked aldehyde inhibitors that incorporate in their structure a conformationally constrained cyclohexane moiety was synthesized and found to potently inhibit severe acute respiratory syndrome coronavirus-2 3CL protease in biochemical and cell-based assays. Several of the inhibitors were also found to be nanomolar inhibitors of Middle East respiratory syndrome coronavirus 3CL protease. The corresponding latent aldehyde bisulfite adducts were found to be equipotent to the precursor aldehydes.

Postinfection treatment with a protease inhibitor increases survival of mice with a fatal SARS-CoV-2 infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection continues to be a serious global public health threat. The 3C-like protease (3CLpro) is a virus protease encoded by SARS-CoV-2, which is essential for virus replication. We have previously reported a series of small-molecule 3CLpro inhibitors effective for inhibiting replication of human coronaviruses including SARS-CoV-2 in cell culture and in animal models.

Structure-Guided Optimization of Dipeptidyl Inhibitors of Norovirus 3CL Protease.

Acute gastroenteritis caused by noroviruses has a major impact on public health worldwide in terms of morbidity, mortality, and economic burden. The disease impacts most severely immunocompromised patients, the elderly, and children. The current lack of approved vaccines and small-molecule therapeutics for the treatment and prophylaxis of norovirus infections underscores the need for the development of norovirus-specific drugs.

3C-like protease inhibitors block coronavirus replication in vitro and improve survival in MERS-CoV-infected mice.

Pathogenic coronaviruses are a major threat to global public health, as exemplified by severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and the newly emerged SARS-CoV-2, the causative agent of coronavirus disease 2019 (COVID-19). We describe herein the structure-guided optimization of a series of inhibitors of the coronavirus 3C-like protease (3CLpro), an enzyme essential for viral replication. The optimized compounds were effective against several human coronaviruses including MERS-CoV, SARS-CoV, and SARS-CoV-2 in an enzyme assay and in cell-based assays using Huh-7 and Vero E6 cell lines.

Caspase-mediated cleavage of nucleocapsid protein of a protease-independent porcine epidemic diarrhea virus strain.

Porcine epidemic diarrhea virus (PEDV) infection in neonatal piglets can cause up to 100% mortality, resulting in significant economic loss in the swine industry. Like other coronaviruses, PEDV N protein is a nucleocapsid protein and abundantly presents at all stages of infection. Previously, we reported that the N protein of trypsin-independent PEDV 8aa is cleaved during virus replication.

Characterization of amino acid substitutions in feline coronavirus 3C-like protease from a cat with feline infectious peritonitis treated with a protease inhibitor.

Feline infectious peritonitis (FIP) is a highly fatal disease caused by a virulent feline coronavirus in domestic and wild cats. We have previously reported the synthesis of potent coronavirus 3C-like protease (3CLpro) inhibitors and the efficacy of a protease inhibitor, GC376, in client-owned cats with FIP. In this study, we studied the effect of the amino acid changes in 3CLpro of feline coronavirus from a feline patient who received antiviral treatment for prolonged duration.

Proteases facilitate the endosomal escape of porcine epidemic diarrhea virus during entry into host cells.

Exogenous and endogenous proteases play important roles in porcine epidemic diarrhea virus (PEDV) entry and replication. The roles of proteases in the viral endosomal escape and replication using trypsin (KD) or elastase (AA)-adapted US PEDV strains were studied. While PEDV KD and AA require different exogenous protease for efficient replication in cells, PEDV KD was more dependent on the protease than PEDV AA.

Putative structural rearrangements associated with the interaction of macrocyclic inhibitors with norovirus 3CL protease.

Human noroviruses are the primary cause of outbreaks of acute gastroenteritis worldwide. The problem is further compounded by the current lack of norovirus-specific antivirals or vaccines. Noroviruses have a single-stranded, positive sense 7 to 8 kb RNA genome which encodes a polyprotein precursor that is processed by a virus-encoded 3C-like cysteine protease (NV 3CLpro) to generate at least six mature nonstructural proteins.

Antiviral Drug Discovery: Norovirus Proteases and Development of Inhibitors.

Proteases are a major enzyme group playing important roles in a wide variety of biological processes in life forms ranging from viruses to mammalians. The aberrant activity of proteases can lead to various diseases; consequently, host proteases have been the focus of intense investigation as potential therapeutic targets. A wide range of viruses encode proteases which play an essential role in viral replication and, therefore, constitute attractive targets for the development of antiviral therapeutics.

Protease inhibitors broadly effective against feline, ferret and mink coronaviruses.

Ferret and mink coronaviruses typically cause catarrhal diarrhea in ferrets and minks, respectively. In recent years, however, systemic fatal coronavirus infection has emerged in ferrets, which resembles feline infectious peritonitis (FIP) in cats. FIP is a highly fatal systemic disease caused by a virulent feline coronavirus infection in cats.