ABSTRACT
A series of pyrazolone compounds as possible SARS-CoV 3CL protease inhibitors were designed, synthesized, and evaluated by in vitro protease assay using fluorogenic substrate peptide in which several showed potent inhibition against the 3CL protease. Interestingly, one of the inhibitors was also active against 3C protease from coxsackievirus B3. These inhibitors could be potentially developed into anti-coronaviral and anti-picornaviral agents.
Subject(s)
Antiviral Agents/chemical synthesis , Protease Inhibitors/chemical synthesis , Pyrazolones/chemistry , Viral Proteins/antagonists & inhibitors , 3C Viral Proteases , Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Binding Sites , Catalytic Domain , Computer Simulation , Coronavirus 3C Proteases , Cysteine Endopeptidases/metabolism , Enterovirus B, Human/enzymology , Humans , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , Pyrazolones/chemical synthesis , Pyrazolones/pharmacology , Structure-Activity Relationship , Viral Proteins/metabolismABSTRACT
A series of 2-(benzylthio)-6-oxo-4-phenyl-1,6-dihydropyrimidine as SARS-CoV 3CL protease inhibitors were developed and their potency was evaluated by in vitro protease inhibitory assays. Two candidates had encouraging results for the development of new anti-SARS compounds.
Subject(s)
Pyrimidines/chemical synthesis , Pyrimidines/pharmacology , Viral Proteins/antagonists & inhibitors , Computer Simulation , Coronavirus 3C Proteases , Cysteine Endopeptidases , Inhibitory Concentration 50 , Models, Molecular , Protease Inhibitors/chemical synthesis , Protease Inhibitors/pharmacology , Protein Binding , Severe acute respiratory syndrome-related coronavirus/drug effects , Severe acute respiratory syndrome-related coronavirus/enzymology , Structure-Activity RelationshipABSTRACT
Picornaviruses (PV) and coronaviruses (CoV) are positive-stranded RNA viruses which infect millions of people worldwide each year, resulting in a wide range of clinical outcomes. As reported in this study, using high throughput screening against approximately 6800 small molecules, we have identified several novel inhibitors of SARS-CoV 3CL(pro) with IC(50) of low microM. Interestingly, one of them equally inhibited both 3C(pro) and 3CL(pro) from PV and CoV, respectively. Using computer modeling, the structural features of these compounds as individual and common protease inhibitors were elucidated to enhance our knowledge for developing anti-viral agents against PV and CoV.
Subject(s)
Coronavirus/drug effects , Coronavirus/enzymology , Peptide Hydrolases/metabolism , Picornaviridae/drug effects , Picornaviridae/enzymology , Protease Inhibitors/chemistry , Protease Inhibitors/pharmacology , Amino Acid Sequence , Computer Simulation , Conserved Sequence , Drug Evaluation, Preclinical , Models, Molecular , Molecular Sequence Data , Peptide Hydrolases/chemistry , Protein Binding , Protein Structure, Tertiary , Sequence Alignment , Structure-Activity RelationshipABSTRACT
Octaprenyl diphosphate synthase (OPPs) and undecaprenyl diphosphate synthases (UPPs) catalyze consecutive condensation reactions of farnesyl diphosphate (FPP) with 5 and 8 isopentenyl diphosphate (IPP) to generate C(40) and C(55) products with trans- and cis-double bonds, respectively. In this study, we used IPP analogue, 3-bromo-3-butenyl diphosphate (Br-IPP), in conjunction with radiolabeled FPP, to probe the reaction mechanisms of the two prenyltransferases. Using this alternative substrate with electron-withdrawing bromo group at the C3 position to slow down the condensation step, trapping of farnesol in the OPPs reaction from radiolabeled FPP under basic condition was observed, consistent with a sequential mechanism. In contrast, UPPs reaction yielded no farnesyl carbocation intermediate under the same condition with radiolabeled FPP and Br-IPP, indicating a concerted mechanism. Our data demonstrate the different reaction mechanisms for cis- and tran-prenyltransferases although they share the same substrates.
Subject(s)
Alkyl and Aryl Transferases/chemistry , Hemiterpenes/chemistry , Organophosphorus Compounds/chemistry , Transferases/chemistry , Chromatography, Thin Layer , Diphosphates/chemistry , Organophosphates , Polyisoprenyl Phosphates/chemistry , Sesquiterpenes/chemistry , Substrate SpecificityABSTRACT
Human enterovirus (EV) belongs to the picornavirus family, which consists of over 200 medically relevant viruses. A peptidomimetic inhibitor AG7088 was developed to inhibit the 3C protease of rhinovirus (a member of the family), a chymotrypsin-like protease required for viral replication, by forming a covalent bond with the active site Cys residue. In this study, we have prepared the recombinant 3C protease from EV71 (TW/2231/98), a particular strain which causes severe outbreaks in Asia, and developed inhibitors against the protease and the viral replication. For inhibitor design, the P3 group of AG7088, which is not interacting with the rhinovirus protease, was replaced with a series of cinnamoyl derivatives directly linked to P2 group through an amide bond to simplify the synthesis. While the replacement caused decreased potency, the activity can be largely improved by substituting the alpha,beta-unsaturated ester with an aldehyde at the P1' position. The best inhibitor 10b showed EC(50) of 18 nM without apparent toxicity (CC(50)>25 microM). Our study provides potent inhibitors of the EV71 3C protease as anti-EV71 agents and facilitates the combinatorial synthesis of derivatives for further improving the inhibitory activity.
Subject(s)
Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Enterovirus A, Human/drug effects , Enterovirus A, Human/enzymology , Viral Proteins/antagonists & inhibitors , 3C Viral Proteases , Amino Acid Sequence , Binding Sites , Cell Line, Tumor , Computer Simulation , Cysteine Endopeptidases/chemistry , Drug Design , Humans , Models, Molecular , Molecular Structure , Structure-Activity Relationship , Substrate Specificity , Viral Proteins/chemistryABSTRACT
Helicobacter pylori colonizes the human gastric epithelium and causes diseases such as gastritis, peptic ulcers, and stomach cancer. Undecaprenyl pyrophosphate synthase (UPPS), which catalyzes consecutive condensation reactions of farnesyl pyrophosphate with eight isopentenyl pyrophosphate to form lipid carrier for bacterial peptidoglycan biosynthesis, represents a potential target for developing new antibiotics. In this study, we solved the crystal structure of H. pylori UPPS and performed virtual screening of inhibitors from a library of 58,635 compounds. Two hits were found to exhibit differential activities against Helicobacter pylori and Escherichia coli UPPS, giving the possibility of developing antibiotics specially targeting pathogenic H. pylori without killing the intestinal E. coli.
Subject(s)
Alkyl and Aryl Transferases/antagonists & inhibitors , Alkyl and Aryl Transferases/ultrastructure , Enzyme Inhibitors/chemistry , Escherichia coli/enzymology , Helicobacter pylori/enzymology , Models, Chemical , Models, Molecular , Binding Sites , Computer Simulation , Enzyme Activation , Enzyme Stability , Protein Binding , Protein ConformationABSTRACT
Acylcyclohexanedione derivatives have been designed, synthesized, and evaluated for in vitro inhibition activity against the enzyme 4-hydroxyphenylpyruvate dioxygenase (4-HPPD). The biological data demonstrated that 7 is a potent inhibitor of 4-HPPD with an IC(50) value of 40 nM. After metabolism, compound 7 has the potential to become a potent inhibitor of a second enzyme, GA(20) 3beta-hydroxylase.
