Redirection of the reaction between activated protein C and a serpin to the substrate pathway

Andrey A. Komissarov, Peter A. Andreasen, Paul J. Declerck, Yuichi Kamikubo, Aiwu Zhou, Andras Gruber

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Background: Activated protein C (APC) reduces mortality in severe sepsis. Protecting APC in the circulatory system from inactivation by serine protease inhibitors (serpins) could improve its therapeutic efficiency. Significantly elevated levels of a serpin plasminogen activator inhibitor 1 (PAI-1) correlate with a lethal outcome in severe sepsis and disseminated intravascular coagulation. Intermolecular mechanisms were employed to redirect the reaction between APC and PAI-1 from the inhibitory to the substrate pathway, which results in the catalytic neutralization of the serpin. Methods: The effects of anti-PAI-1 monoclonal antibodies (mAbs) and vitronectin, as well as their fragments, on the kinetics and stoichiometry of the reaction between PAI-1 and APC were studied using SDS PAGE and fluorescence spectroscopy. Results: MAbs with epitopes at α-helix F redirected 70-80% of the reaction between PAI-1 and APC, to the substrate pathway. Vitronectin and its SMB domain did not affect the stoichiometry of acyl-enzyme formation, but enhanced the effect of mAbs. While vitronectin induced a more than two-fold increase in the rate of the reaction between PAI-1 and APC, neither mAbs (mAb fragments), nor SMB domain of vitronectin affected it. Conclusions: Ligands interacting with α-helix F of PAI-1 demonstrated a potential for the protection of APC from inactivation by PAI-1. Since the mechanism of proteinase/serpin interaction is universal, a similar design and approach could be employed for enhancing the inactivation of other serpins in order to preserve APC activity in the circulation. Rational pharmacological targeting of the inhibitors of APC could have therapeutic utility.

Original languageEnglish (US)
Pages (from-to)397-404
Number of pages8
JournalThrombosis Research
Volume122
Issue number3
DOIs
StatePublished - 2008

Fingerprint

Serine Proteinase Inhibitors
Plasminogen Activator Inhibitor 1
Protein C
Vitronectin
Monoclonal Antibodies
Sepsis
Immunoglobulin Fragments
Protein C Inhibitor
Disseminated Intravascular Coagulation
Fluorescence Spectrometry
Cardiovascular System
Epitopes
Polyacrylamide Gel Electrophoresis
Peptide Hydrolases
Pharmacology
Ligands
Mortality
Enzymes
Therapeutics

Keywords

  • Activated protein C
  • Disseminated intravascular coagulation
  • Monoclonal antibodies
  • Plasminogen activator inhibitor 1
  • Sepsis
  • Vitronectin

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine
  • Hematology

Cite this

Redirection of the reaction between activated protein C and a serpin to the substrate pathway. / Komissarov, Andrey A.; Andreasen, Peter A.; Declerck, Paul J.; Kamikubo, Yuichi; Zhou, Aiwu; Gruber, Andras.

In: Thrombosis Research, Vol. 122, No. 3, 2008, p. 397-404.

Research output: Contribution to journalArticle

Komissarov, Andrey A. ; Andreasen, Peter A. ; Declerck, Paul J. ; Kamikubo, Yuichi ; Zhou, Aiwu ; Gruber, Andras. / Redirection of the reaction between activated protein C and a serpin to the substrate pathway. In: Thrombosis Research. 2008 ; Vol. 122, No. 3. pp. 397-404.
@article{9b5a157517654f038a6fd28cc26645d2,
title = "Redirection of the reaction between activated protein C and a serpin to the substrate pathway",
abstract = "Background: Activated protein C (APC) reduces mortality in severe sepsis. Protecting APC in the circulatory system from inactivation by serine protease inhibitors (serpins) could improve its therapeutic efficiency. Significantly elevated levels of a serpin plasminogen activator inhibitor 1 (PAI-1) correlate with a lethal outcome in severe sepsis and disseminated intravascular coagulation. Intermolecular mechanisms were employed to redirect the reaction between APC and PAI-1 from the inhibitory to the substrate pathway, which results in the catalytic neutralization of the serpin. Methods: The effects of anti-PAI-1 monoclonal antibodies (mAbs) and vitronectin, as well as their fragments, on the kinetics and stoichiometry of the reaction between PAI-1 and APC were studied using SDS PAGE and fluorescence spectroscopy. Results: MAbs with epitopes at α-helix F redirected 70-80{\%} of the reaction between PAI-1 and APC, to the substrate pathway. Vitronectin and its SMB domain did not affect the stoichiometry of acyl-enzyme formation, but enhanced the effect of mAbs. While vitronectin induced a more than two-fold increase in the rate of the reaction between PAI-1 and APC, neither mAbs (mAb fragments), nor SMB domain of vitronectin affected it. Conclusions: Ligands interacting with α-helix F of PAI-1 demonstrated a potential for the protection of APC from inactivation by PAI-1. Since the mechanism of proteinase/serpin interaction is universal, a similar design and approach could be employed for enhancing the inactivation of other serpins in order to preserve APC activity in the circulation. Rational pharmacological targeting of the inhibitors of APC could have therapeutic utility.",
keywords = "Activated protein C, Disseminated intravascular coagulation, Monoclonal antibodies, Plasminogen activator inhibitor 1, Sepsis, Vitronectin",
author = "Komissarov, {Andrey A.} and Andreasen, {Peter A.} and Declerck, {Paul J.} and Yuichi Kamikubo and Aiwu Zhou and Andras Gruber",
year = "2008",
doi = "10.1016/j.thromres.2007.10.012",
language = "English (US)",
volume = "122",
pages = "397--404",
journal = "Thrombosis Research",
issn = "0049-3848",
publisher = "Elsevier Limited",
number = "3",

}

TY - JOUR

T1 - Redirection of the reaction between activated protein C and a serpin to the substrate pathway

AU - Komissarov, Andrey A.

AU - Andreasen, Peter A.

AU - Declerck, Paul J.

AU - Kamikubo, Yuichi

AU - Zhou, Aiwu

AU - Gruber, Andras

PY - 2008

Y1 - 2008

N2 - Background: Activated protein C (APC) reduces mortality in severe sepsis. Protecting APC in the circulatory system from inactivation by serine protease inhibitors (serpins) could improve its therapeutic efficiency. Significantly elevated levels of a serpin plasminogen activator inhibitor 1 (PAI-1) correlate with a lethal outcome in severe sepsis and disseminated intravascular coagulation. Intermolecular mechanisms were employed to redirect the reaction between APC and PAI-1 from the inhibitory to the substrate pathway, which results in the catalytic neutralization of the serpin. Methods: The effects of anti-PAI-1 monoclonal antibodies (mAbs) and vitronectin, as well as their fragments, on the kinetics and stoichiometry of the reaction between PAI-1 and APC were studied using SDS PAGE and fluorescence spectroscopy. Results: MAbs with epitopes at α-helix F redirected 70-80% of the reaction between PAI-1 and APC, to the substrate pathway. Vitronectin and its SMB domain did not affect the stoichiometry of acyl-enzyme formation, but enhanced the effect of mAbs. While vitronectin induced a more than two-fold increase in the rate of the reaction between PAI-1 and APC, neither mAbs (mAb fragments), nor SMB domain of vitronectin affected it. Conclusions: Ligands interacting with α-helix F of PAI-1 demonstrated a potential for the protection of APC from inactivation by PAI-1. Since the mechanism of proteinase/serpin interaction is universal, a similar design and approach could be employed for enhancing the inactivation of other serpins in order to preserve APC activity in the circulation. Rational pharmacological targeting of the inhibitors of APC could have therapeutic utility.

AB - Background: Activated protein C (APC) reduces mortality in severe sepsis. Protecting APC in the circulatory system from inactivation by serine protease inhibitors (serpins) could improve its therapeutic efficiency. Significantly elevated levels of a serpin plasminogen activator inhibitor 1 (PAI-1) correlate with a lethal outcome in severe sepsis and disseminated intravascular coagulation. Intermolecular mechanisms were employed to redirect the reaction between APC and PAI-1 from the inhibitory to the substrate pathway, which results in the catalytic neutralization of the serpin. Methods: The effects of anti-PAI-1 monoclonal antibodies (mAbs) and vitronectin, as well as their fragments, on the kinetics and stoichiometry of the reaction between PAI-1 and APC were studied using SDS PAGE and fluorescence spectroscopy. Results: MAbs with epitopes at α-helix F redirected 70-80% of the reaction between PAI-1 and APC, to the substrate pathway. Vitronectin and its SMB domain did not affect the stoichiometry of acyl-enzyme formation, but enhanced the effect of mAbs. While vitronectin induced a more than two-fold increase in the rate of the reaction between PAI-1 and APC, neither mAbs (mAb fragments), nor SMB domain of vitronectin affected it. Conclusions: Ligands interacting with α-helix F of PAI-1 demonstrated a potential for the protection of APC from inactivation by PAI-1. Since the mechanism of proteinase/serpin interaction is universal, a similar design and approach could be employed for enhancing the inactivation of other serpins in order to preserve APC activity in the circulation. Rational pharmacological targeting of the inhibitors of APC could have therapeutic utility.

KW - Activated protein C

KW - Disseminated intravascular coagulation

KW - Monoclonal antibodies

KW - Plasminogen activator inhibitor 1

KW - Sepsis

KW - Vitronectin

UR - http://www.scopus.com/inward/record.url?scp=45849148895&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=45849148895&partnerID=8YFLogxK

U2 - 10.1016/j.thromres.2007.10.012

DO - 10.1016/j.thromres.2007.10.012

M3 - Article

VL - 122

SP - 397

EP - 404

JO - Thrombosis Research

JF - Thrombosis Research

SN - 0049-3848

IS - 3

ER -