(191ck) The Discovery of Enzymatically Depolymerized Heparin Derivatives for the Treatment of Ulcerative Colitis

Heparin is a member of glycosaminoglycan and consists of variably sulfated repeating disaccharide units. Besides its well-known and fully exploited anticoagulant activity, heparin was found to possess many non-anticoagulant activities that were valuable for therapeutic use. Heparan sulfate(HS), very closely related in structure to heparin, is a ubiquitous glycosaminoglycan regulating a wide variety of biological activities including developmental processes, inflammation, coagulation, immunity and tumor metastasis. Based on the review of existing researches, the molecular size and sulfation pattern of heparin/HS both determine their non-anticoagulant efficacy. Accordingly, to maximizing the diversity of heparin structure, strategies of controllable enzymatic depolymerization using three types of heparinases with different substrate specificity and chemically selective modification of heparin molecule were combined. The enzyme kinetics of three heparinases towards different chemically modified heparins were studied, giving instructions to improve the process of enzymatically production of low molecular weight heparin derivatives. Therefore, a library of heparin derivatives with different molecular weights and sulfation patterns was designed and constructed. In spite of a heterogenous mixture of different polysaccharide chains, each species of the heparin library had distinct molecular weight distribution and defined structure, which is the fundamental prerequisite for structrue-activity relationship analysis.

The anti-inflammatory effects of the heparin derivatives were systematically screened using the animal model of DSS induced colitis in mice. The results indicated that an enzymatically depolymerized low molecular weight heparin derivative without anticoagulant activity had better colon-protective effects through the inhibition of cytokine secretion and macrophage recruitment. The degree of desulfation and de-anticoagulation of heparin is extremely important for the treatment of colitis in mice. Furthermore, enzymatically depolymerization of heparin with specific type of heparinase show benefits in ameliorating colitis in mice. The selected heparin derivatives also attenuated intestinal epithelial permeability according to the studies on a normal human colon mucosal epithelial cell line. Several molecular pathways involved in the ulcerative colitis with the treatment of the potent heparin derivative were depicted, and the structure factors of heparin influencing anti-colitis activity were thoroughly analyzed.