Defense Date

2007

Document Type

Dissertation

Degree Name

Doctor of Philosophy

Department

Medicinal Chemistry

First Advisor

Dr. Umesh R. Desai

Abstract

The occurrence of thrombosis in several pathophysiological conditions creates a huge need for anticoagulation therapy. Thrombin and factor Xa have been prime targets for regulation of clotting through the direct and indirect mechanism of inhibition. This work investigates chemo-enzymatically prepared oligomers of 4-hydroxycinnamic acids (DHPs) as potential anticoagulants. Oligomers were prepared through peroxidase-catalyzed oxidative coupling of 4-hydroxycinnamic acids. The products resulting from this reaction are called CDs, FDs and SDs. Structurally, these sulfated DHPs are unique and do not resemble any of the anticoagulants known in the literature.DHP oligomers were found to increase clotting times at concentrations comparable to heparin. Studies in blood and plasma show that DHPs possess an anticoagulation profile similar to enoxaparin. To understand the mechanism of action of DHPs, we studied the inhibition of thrombin, FXa, FIXa, and FVIIa in the presence and absence of antithrombin. CDs and FDs display a preference for direct inhibition of thrombin and FXa, and exhibit a high level of specificity over FIXa and FVIIa. In the presence of AT, CDs and FDs displayed weaker inhibition of FXa and thrombin suggesting that binding to AT is a competitive side reaction. SDs exhibited potent inhibition of FXa and thrombin in the absence of antithrombin, but was inactive against FIXa and FVIIa representing the best selectivity among the DHPs. For SDs, inhibition of all the pro-coagulant enzymes favored the antithrombin dependent pathway. Binding studies were performed to determine how CDs directly inhibits thrombin. Competitive binding studies suggest that CDs interacts with exosite II and disrupts the catalytic triad of thrombin. These results indicate that the preferred mechanism of CDs action is exosite II mediated allosteric disruption of thrombin. CDs appears to be the first exosite II mediated DTI and this represents a novel mechanism of inhibitor function. The inhibition characteristics of DHPs are unique and radically different in structure from all the current clinically used anticoagulants. To the best of our knowledge this dual mechanism of anticoagulation and unique binding mode has not been described as yet in literature and represents a novel strategy that our laboratory has discovered.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

June 2008

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