Change search
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Novel Procedures for Identification and Characterization of Viral Proteases Inhibitors
Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Viral proteases are often considered to be attractive drug targets because of their crucial function in the viral replication machinery. In order to increase our knowledge of these important targets and to contribute to the discovery and development of new antiviral drugs, the proteases from hepatitis C virus (HCV) and human cytomegalovirus (HCMV) have been produced and their interactions with inhibitors and fragments have been characterized, using enzyme inhibition and SPR biosensor based interaction assay.

The structure activity relationships and the resistance profiles of a series of HCV NS3 protease inhibitors based on either P2 proline or phenylglycine residues were analyzed using wild type genotype 1a and the major resistant variants A156T and D168V. The observed susceptibility to substitutions associated with these resistance variants was concluded to depend on the P2 and the P1 residue, and not only on the P2 residue as previously had been suggested. In order to be able to evaluate how the potency of inhibitors is affected by genetic variation, their effect was evaluated on wild type NS3 from genotype 1a, 1b and 3a as well as on the resistant variant R155K from genotype 1a. To enable a comparison of the inhibitory effect on the enzyme variants, the compounds were analyzed under conditions optimized for each variant. VX-950 was found to be the least susceptible compound to resistance and genetic variation. A more detailed analysis showed that the kinetic and mechanistic features of the inhibitors were significantly different for the different genotypes. The reversible non covalent macrocyclic inhibitor ITMN 191 was revealed to have favorable kinetics for all three genotypes. This is an advantage for the design of broad spectrum drugs.

A fragment based procedure for identifying and validating novel scaffolds for inhibitors of HCMV protease was established. It identified fragments that may serve as starting points for the discovery of effective inhibitors against this challenging target.  

The procedures developed for the evaluation and identification of novel HCV NS3 and HCMV protease inhibitors have contributed to a deeper understanding of protease-inhibitor interactions that is expected to have an impact on the design of novel antiviral drugs. 

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. , 50 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1115
Keyword [en]
drug discovery, viral proteases, inhibitors, Hepatitis C, NS3, cytomegalovirus
National Category
Chemical Sciences
Research subject
Biochemistry
Identifiers
URN: urn:nbn:se:uu:diva-215700ISBN: 978-91-554-8855-0 (print)OAI: oai:DiVA.org:uu-215700DiVA: diva2:688134
Public defence
2014-02-28, B42, BMC, Uppsala Universitet, Husargatan 3, 751 23, Uppsala, 13:15 (English)
Opponent
Supervisors
Available from: 2014-02-07 Created: 2014-01-15 Last updated: 2014-02-10
List of papers
1. Structure-activity relationships of HCV NS3 protease inhibitors evaluated on the drug-resistant variants A156T and D168V
Open this publication in new window or tab >>Structure-activity relationships of HCV NS3 protease inhibitors evaluated on the drug-resistant variants A156T and D168V
Show others...
2010 (English)In: Antiviral Therapy, ISSN 1359-6535, E-ISSN 2040-2058, Vol. 15, no 6, 841-852 p.Article in journal (Refereed) Published
Abstract [en]

BACKGROUND: HCV infections are a serious threat to public health. An important drug target is the NS3 protease, for which several inhibitors are in clinical trials. Because of the high mutation rate of the virus, resistance against any HCV-specific drug is likely to become a substantial problem. Structure-activity data for the major resistant variants are therefore needed to guide future designs of protease inhibitors. METHODS: The inhibitory potency of tripeptide NS3 protease inhibitors, with either a P2 proline or phenylglycine, in combination with different P3 and P1-P1' groups, was assessed in enzyme activity assays using the full-length NS3 protein with known resistance-conferring substitutions A156T or D168V. The results obtained from these variants were compared with the inhibition of the wild-type enzyme. Molecular modelling was used to rationalize the biochemical results. RESULTS: Inhibitors combining the P2 proline and P1 (1R,2S)-1-amino-2-vinylcyclopropyl-carboxylic acid (vinylACCA) lost much of their potency on the resistant variants. Exchange of the P2 proline for phenylglycine yielded inhibitors that were equipotent on the wild-type and on the A156T and D168V variants. The same result was obtained from the combination of either the P2 residue with a norvaline or an aromatic scaffold in the P1 position. CONCLUSIONS: The combination of a substituted P2 proline and P1 vinylACCA appears to be the main problem behind the observed resistance. Molecular modelling suggests an enforced change in binding conformation for the P2 proline-based inhibitors, whereas the phenylglycine-based inhibitors retained their wild-type binding conformation in the substituted forms of the enzyme.

National Category
Pharmaceutical Sciences Chemical Sciences
Identifiers
urn:nbn:se:uu:diva-131962 (URN)10.3851/IMP1655 (DOI)000282390300004 ()20834096 (PubMedID)
Available from: 2010-10-12 Created: 2010-10-12 Last updated: 2017-12-12Bibliographically approved
2. Accounting for strain variations and resistance mutations in the characterization of hepatitis C NS3 protease inhibitors
Open this publication in new window or tab >>Accounting for strain variations and resistance mutations in the characterization of hepatitis C NS3 protease inhibitors
Show others...
2014 (English)In: Journal of enzyme inhibition and medicinal chemistry (Print), ISSN 1475-6366, E-ISSN 1475-6374, Vol. 29, no 6, 868-876 p.Article in journal (Refereed) Published
Abstract [en]

Context: Natural strain variation and rapid resistance development makes development of broad spectrum hepatitis C virus (HCV) drugs very challenging and evaluation of inhibitor selectivity and resistance must account for differences in the catalytic properties of enzyme variants.

Objective: To understand how to study selectivity and relationships between efficacy and genotype or resistant mutants for NS3 protease inhibitors.

Materials and methods: The catalytic properties of NS3 protease from genotypes 1a, 1b and 3a, and their sensitivities to four structurally and mechanistically different NS3 protease inhibitors have been analysed under different experimental conditions.

Results: The optimisation of buffer conditions for each protease variant enabled the comparison of their catalytic properties and sensitivities to the inhibitors. All inhibitors were most effective against genotype 1a protease, with VX-950 having the broadest selectivity.

Discussion and conclusion: A new strategy for evaluation of inhibitors relevant for the discovery of broad spectrum HCV drugs was established.

National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-193252 (URN)10.3109/14756366.2013.864651 (DOI)000345518000013 ()24517372 (PubMedID)
Available from: 2013-01-29 Created: 2013-01-29 Last updated: 2017-12-06Bibliographically approved
3. Identification of Weak Points of Hepatitis C Virus NS3 Protease Inhibitors Using Surface Plasmon Resonance Biosensor-Based Interaction Kinetic Analysis and Genetic Variants
Open this publication in new window or tab >>Identification of Weak Points of Hepatitis C Virus NS3 Protease Inhibitors Using Surface Plasmon Resonance Biosensor-Based Interaction Kinetic Analysis and Genetic Variants
Show others...
2014 (English)In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 57, no 5, 1802-1811 p.Article in journal (Other academic) Published
National Category
Natural Sciences
Identifiers
urn:nbn:se:uu:diva-193251 (URN)10.1021/jm401690f (DOI)000333005800012 ()
Available from: 2013-01-29 Created: 2013-01-29 Last updated: 2017-12-06Bibliographically approved
4. Challenges in the discovery of fragment targeting human cytomegalovirus protease
Open this publication in new window or tab >>Challenges in the discovery of fragment targeting human cytomegalovirus protease
Show others...
(English)Manuscript (preprint) (Other academic)
Keyword
CMV, cytomagalovirus, protease, SPR, biosensor, fragments-based drug discovery, screening, inhibitor
National Category
Natural Sciences
Research subject
Biochemistry
Identifiers
urn:nbn:se:uu:diva-215703 (URN)
Available from: 2014-01-15 Created: 2014-01-15 Last updated: 2014-02-10

Open Access in DiVA

fulltext(4847 kB)466 downloads
File information
File name FULLTEXT01.pdfFile size 4847 kBChecksum SHA-512
b89f1a858d4341e3208f86146f53a90039f87e1606b56e456c82259a4ae3cbf47d10501c2d3dbc99b99677cbeecde2b4337cd40ff92a5fc24f91de0e0d971b43
Type fulltextMimetype application/pdf
Buy this publication >>

Search in DiVA

By author/editor
Ehrenberg, Angelica
By organisation
Biochemistry
Chemical Sciences

Search outside of DiVA

GoogleGoogle Scholar
Total: 466 downloads
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

isbn
urn-nbn

Altmetric score

isbn
urn-nbn
Total: 1112 hits
CiteExportLink to record
Permanent link

Direct link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association-8th-edition
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf