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WSU Access

Date of Award


Degree Type


Degree Name



Pharmaceutical Sciences

First Advisor

Robert J. Kerns


Heparan sulfate mediates many biological processes and plays a role in many diseases, such as cancer, atherosclerosis, inflammation, alzheimers, and viral infection. The structurally similar polysaccharide heparin is a powerful clinical anticoagulant but causes numerous side effects and interacts with proteins that are involved in heparan sulfate-mediated biological processes. Numerous therapeutic applications other than anticoagulation have been proposed for heparin or heparin-like analogs and mimics that selectively bind a particular protein target (heparin-binding protein) and thus mediate a specific biological activity. However, due to the polyanionic nature of heparin and most heparin mimics, the discovery of compounds that selectively target individual heparin-binding proteins has been difficult. Explored here is a new strategy for preparing and identifying novel heparin derivatives that display increased affinity and selectivity for individual heparin-binding proteins. Using heparin as a scaffold for the generation of libraries of novel heparin analogs, differential N-desulfonation and N-deacetylation of heparin was performed to reveal free amine groups for the subsequent introduction of structural diversity. Structurally diverse non-anionic moieties to be incorporated into the heparin structure were identified based on their capacity to interact with protein through hydrogen bonds and/or hydrophobic interactions. Utilizing parallel synthesis, these structurally diverse moieties were incorporated into heparin via chemical coupling reactions, affording a library of heparin analogs. Utilizing competition binding assays and enzyme inhibition assays, the library was screened against various heparin and HS binding proteins (alpha-thrombin, SLPI, MPO, laminin, VEGF, FGF1, and FGF2 for binding assays; alpha-thrombin, elastase, and cathepsin G for enzyme inhibition assays). Unique analogs were identified as 'hit' ligands for alpha-thrombin binding and inhibition of thrombin protease activity. One of those analogs, a N-desulfonated heparin fraction substituted with 3-(4-hydroxyphenyl)propionyl, demonstrated increased binding affinity and selectivity toward alpha-thrombin and displayed direct inhibition of this enzyme. Based on the hit ligand, a focused library of heparin analogs was synthesized. Structure-activity relationship studies revealed that a 4-hydroxyphenyl moiety properly distanced from the saccharide core afforded the heparin analogs with the greatest affinity for thrombin and were the most potent inhibitors of thrombin activity.

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