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Reversal of the Progression of Fatal Coronavirus Infection in Cats by a Broad-Spectrum Coronavirus Protease Inhibitor.

Kim Y, Liu H, Galasiti Kankanamalage AC, Weerasekara S, Hua DH, Groutas WC, Chang KO, Pedersen NC - PLoS Pathog. (2016)

Bottom Line: The increasing risks of highly virulent coronavirus infections in humans or animals call for effective antiviral drugs, but no such measures are yet available.Experimental FIP is 100% fatal once certain clinical and laboratory signs become apparent.These results indicate that continuous virus replication is required for progression of immune-mediated inflammatory disease of FIP.

View Article: PubMed Central - PubMed

Affiliation: Department of Diagnostic Medicine and Pathobiology, College of Veterinary Medicine, Kansas State University, Manhattan, Kansas, United States of America.

ABSTRACT
Coronaviruses infect animals and humans causing a wide range of diseases. The diversity of coronaviruses in many mammalian species is contributed by relatively high mutation and recombination rates during replication. This dynamic nature of coronaviruses may facilitate cross-species transmission and shifts in tissue or cell tropism in a host, resulting in substantial change in virulence. Feline enteric coronavirus (FECV) causes inapparent or mild enteritis in cats, but a highly fatal disease, called feline infectious peritonitis (FIP), can arise through mutation of FECV to FIP virus (FIPV). The pathogenesis of FIP is intimately associated with immune responses and involves depletion of T cells, features shared by some other coronaviruses like Severe Acute Respiratory Syndrome Coronavirus. The increasing risks of highly virulent coronavirus infections in humans or animals call for effective antiviral drugs, but no such measures are yet available. Previously, we have reported the inhibitors that target 3C-like protease (3CLpro) with broad-spectrum activity against important human and animal coronaviruses. Here, we evaluated the therapeutic efficacy of our 3CLpro inhibitor in laboratory cats with FIP. Experimental FIP is 100% fatal once certain clinical and laboratory signs become apparent. We found that antiviral treatment led to full recovery of cats when treatment was started at a stage of disease that would be otherwise fatal if left untreated. Antiviral treatment was associated with a rapid improvement in fever, ascites, lymphopenia and gross signs of illness and cats returned to normal health within 20 days or less of treatment. Significant reduction in viral titers was also observed in cats. These results indicate that continuous virus replication is required for progression of immune-mediated inflammatory disease of FIP. These findings may provide important insights into devising therapeutic strategies and selection of antiviral compounds for further development for important coronaviruses in animals and humans.

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The chemical structures of 3CLpro inhibitors and their antiviral activity against feline coronavirus in cell culture.The chemical structures of GC376 and NPI64 are shown. The 50% effective concentration (EC50) values of GC376 or NPI64 against FIPV 3CLpro [28, 30] and the 50% cytotoxic concentration (CC50) values of GC376 or NPI64 determined in various cell lines were previously reported [28, 30] and summarized in a table.
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ppat.1005531.g001: The chemical structures of 3CLpro inhibitors and their antiviral activity against feline coronavirus in cell culture.The chemical structures of GC376 and NPI64 are shown. The 50% effective concentration (EC50) values of GC376 or NPI64 against FIPV 3CLpro [28, 30] and the 50% cytotoxic concentration (CC50) values of GC376 or NPI64 determined in various cell lines were previously reported [28, 30] and summarized in a table.

Mentions: GC376 (Fig 1) is a representative compound of the dipeptidyl transition state 3CLpro inhibitors [28–30, 32] whose synthesis was described previously [28]. NPI64 shares homologous structural elements with GC376, except that NPI64 has an additional residue of 1-naththylalanine compared to GC376 in a position that corresponds to the P3 position [30], using the nomenclature of Schechter and Berger [33] (Fig 1). The comparable antiviral activity of GC376 and NPI64 against the replication of feline coronavirus in a cell culture system was previously reported (Fig 1) [28, 30]. However, their PK properties have not been reported. In this study, we investigated the drug plasma concentration changes in healthy specific pathogen free (SPF) cats of 6–9 month age (n = 2 for each compound) following single subcutaneous (s.c.) dose of 10 mg/kg GC376 or 5 mg/kg NPI64. Serial blood samples were then collected and the plasma drug concentrations were measured. Previously, we reported that GC376 is converted into an aldehyde form by the removal of the bisulfite group, and the aldehyde form forms a reversible covalent bond with the nucleophilic cysteine residue of 3CLpro in the x-ray crystallography studies [28]. We also observed the conversion of NPI64 into its aldehyde form in the blood. Therefore the aldehyde forms of GC376 or NPI64 were measured in the plasma samples. Fig 2A shows the plasma drug concentrations over time following single-dose administration of GC376 (red triangles) or NPI64 (black circles). The PK study results indicate that GC376 is rapidly absorbed after s.c. administration and the peak plasma level was reached within 2 hr after injection. The mean plasma drug concentrations remained above the 50% effective concentration (EC50) value of the aldehyde form of GC376 (8 ng/ml) for 18 hrs post injection (Fig 2A, red triangles). The plasma drug concentrations following injection of 5 mg/kg NPI64 stayed above the EC50 value of the aldehyde form of NPI64 (12 ng/ml) for 12 hrs post injection (Fig 2A, black circles). The maximum detected plasma drug concentration following NPI64 administration was substantially lower than that of GC376 by 9.5-fold. This result indicate that GC376 was more easily absorbed than NPI64 via the tested route, even when the lower dose of NPI64 (5 mg/kg), compared to GC376 (10 mg/kg), was taken into account.


Reversal of the Progression of Fatal Coronavirus Infection in Cats by a Broad-Spectrum Coronavirus Protease Inhibitor.

Kim Y, Liu H, Galasiti Kankanamalage AC, Weerasekara S, Hua DH, Groutas WC, Chang KO, Pedersen NC - PLoS Pathog. (2016)

The chemical structures of 3CLpro inhibitors and their antiviral activity against feline coronavirus in cell culture.The chemical structures of GC376 and NPI64 are shown. The 50% effective concentration (EC50) values of GC376 or NPI64 against FIPV 3CLpro [28, 30] and the 50% cytotoxic concentration (CC50) values of GC376 or NPI64 determined in various cell lines were previously reported [28, 30] and summarized in a table.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4814111&req=5

ppat.1005531.g001: The chemical structures of 3CLpro inhibitors and their antiviral activity against feline coronavirus in cell culture.The chemical structures of GC376 and NPI64 are shown. The 50% effective concentration (EC50) values of GC376 or NPI64 against FIPV 3CLpro [28, 30] and the 50% cytotoxic concentration (CC50) values of GC376 or NPI64 determined in various cell lines were previously reported [28, 30] and summarized in a table.
Mentions: GC376 (Fig 1) is a representative compound of the dipeptidyl transition state 3CLpro inhibitors [28–30, 32] whose synthesis was described previously [28]. NPI64 shares homologous structural elements with GC376, except that NPI64 has an additional residue of 1-naththylalanine compared to GC376 in a position that corresponds to the P3 position [30], using the nomenclature of Schechter and Berger [33] (Fig 1). The comparable antiviral activity of GC376 and NPI64 against the replication of feline coronavirus in a cell culture system was previously reported (Fig 1) [28, 30]. However, their PK properties have not been reported. In this study, we investigated the drug plasma concentration changes in healthy specific pathogen free (SPF) cats of 6–9 month age (n = 2 for each compound) following single subcutaneous (s.c.) dose of 10 mg/kg GC376 or 5 mg/kg NPI64. Serial blood samples were then collected and the plasma drug concentrations were measured. Previously, we reported that GC376 is converted into an aldehyde form by the removal of the bisulfite group, and the aldehyde form forms a reversible covalent bond with the nucleophilic cysteine residue of 3CLpro in the x-ray crystallography studies [28]. We also observed the conversion of NPI64 into its aldehyde form in the blood. Therefore the aldehyde forms of GC376 or NPI64 were measured in the plasma samples. Fig 2A shows the plasma drug concentrations over time following single-dose administration of GC376 (red triangles) or NPI64 (black circles). The PK study results indicate that GC376 is rapidly absorbed after s.c. administration and the peak plasma level was reached within 2 hr after injection. The mean plasma drug concentrations remained above the 50% effective concentration (EC50) value of the aldehyde form of GC376 (8 ng/ml) for 18 hrs post injection (Fig 2A, red triangles). The plasma drug concentrations following injection of 5 mg/kg NPI64 stayed above the EC50 value of the aldehyde form of NPI64 (12 ng/ml) for 12 hrs post injection (Fig 2A, black circles). The maximum detected plasma drug concentration following NPI64 administration was substantially lower than that of GC376 by 9.5-fold. This result indicate that GC376 was more easily absorbed than NPI64 via the tested route, even when the lower dose of NPI64 (5 mg/kg), compared to GC376 (10 mg/kg), was taken into account.

Bottom Line: The increasing risks of highly virulent coronavirus infections in humans or animals call for effective antiviral drugs, but no such measures are yet available.Experimental FIP is 100% fatal once certain clinical and laboratory signs become apparent.These results indicate that continuous virus replication is required for progression of immune-mediated inflammatory disease of FIP.

View Article: PubMed Central - PubMed

Affiliation: Department of Diagnostic Medicine and Pathobiology, College of Veterinary Medicine, Kansas State University, Manhattan, Kansas, United States of America.

ABSTRACT
Coronaviruses infect animals and humans causing a wide range of diseases. The diversity of coronaviruses in many mammalian species is contributed by relatively high mutation and recombination rates during replication. This dynamic nature of coronaviruses may facilitate cross-species transmission and shifts in tissue or cell tropism in a host, resulting in substantial change in virulence. Feline enteric coronavirus (FECV) causes inapparent or mild enteritis in cats, but a highly fatal disease, called feline infectious peritonitis (FIP), can arise through mutation of FECV to FIP virus (FIPV). The pathogenesis of FIP is intimately associated with immune responses and involves depletion of T cells, features shared by some other coronaviruses like Severe Acute Respiratory Syndrome Coronavirus. The increasing risks of highly virulent coronavirus infections in humans or animals call for effective antiviral drugs, but no such measures are yet available. Previously, we have reported the inhibitors that target 3C-like protease (3CLpro) with broad-spectrum activity against important human and animal coronaviruses. Here, we evaluated the therapeutic efficacy of our 3CLpro inhibitor in laboratory cats with FIP. Experimental FIP is 100% fatal once certain clinical and laboratory signs become apparent. We found that antiviral treatment led to full recovery of cats when treatment was started at a stage of disease that would be otherwise fatal if left untreated. Antiviral treatment was associated with a rapid improvement in fever, ascites, lymphopenia and gross signs of illness and cats returned to normal health within 20 days or less of treatment. Significant reduction in viral titers was also observed in cats. These results indicate that continuous virus replication is required for progression of immune-mediated inflammatory disease of FIP. These findings may provide important insights into devising therapeutic strategies and selection of antiviral compounds for further development for important coronaviruses in animals and humans.

Show MeSH
Related in: MedlinePlus