Modulation of the Heparanase-inhibiting Activity of Heparin through Selective Desulfation, Graded N-Acetylation, and Glycol Splitting

Abstract

Heparanase is an endo-␤-glucuronidase that cleaves heparan sulfate (HS) chains of heparan sulfate proteoglycans on cell surfaces and in the extracellular matrix (ECM). Heparanase, overexpressed by most cancer cells, facilitates extravasation of blood-borne tumor cells and causes release of growth factors sequestered by HS chains, thus accelerating tumor growth and metastasis. Inhibition of heparanase with HS mimics is a promising target for a novel strategy in cancer therapy. In this study, in vitro inhibition of recombinant heparanase was determined for heparin derivatives differing in degrees of 2-O-and 6-O-sulfation, N-acetylation, and glycol splitting of nonsulfated uronic acid residues. The contemporaneous presence of sulfate groups at O-2 of IdoA and at O-6 of GlcN was found to be non-essential for effective inhibition of heparanase activity provided that one of the two positions retains a high degree of sulfation. N-Desulfation/ N-acetylation involved a marked decrease in the inhibitory activity for degrees of N-acetylation higher than 50%, suggesting that at least one NSO 3 group per disaccharide unit is involved in interaction with the enzyme. On the other hand, glycol splitting of preexisting or of both preexisting and chemically generated nonsulfated uronic acids dramatically increased the heparanase-inhibiting activity irrespective of the degree of N-acetylation. Indeed N-acetylated heparins in their glycol-split forms inhibited heparanase as effectively as the corresponding N-sulfated derivatives. Whereas heparin and N-acetylheparins containing unmodified D-glucuronic acid residues inhibited heparanase by acting, at least in part, as substrates, their glycol-split derivatives were no more susceptible to cleavage by heparanase. Glycol-split N-acetylheparins did not release basic fibroblast growth factor from ECM and failed to stimulate its mitogenic activity. The combination of high inhibition of heparanase and low release/potentiation of ECM-bound growth factor indicates that N-acetylated, glycol-split heparins are potential antiangiogenic and antimetastatic agents that are more effective than their counterparts with unmodified backbones.

Heparanase induces a differential loss of heparan sulphate domains in overt diabetic nephropathy

“…Since these drugs can also inhibit heparanase activity [46,47], it is possible that their ameliorating effect on proteinuria is due to inhibition of heparanase. A study by Levidiotis et al [48] showed that inhibition of heparanase with the specific heparanase inhibitor PI-88 reduced proteinuria in passive Heymann nephritis.…”

Old and new applications of non-anticoagulant heparin

“…depolymerization, and preserves both the pattern and degree of sulfation [98,99]. Periodate can selectively oxidize adjacent carbons bearing unsubstituted hydroxyl and /or amino groups [100], leading, in heparins, to the splitting of C(2)-C(3) bonds of un-sulfated uronic acids including the glucuronic acid, within the ATBR, essential for high anticoagulant activity [101].…”

Investigating Glycol-Split-Heparin-Derived Inhibitors of Heparanase: A Study of Synthetic Trisaccharides

“…The methyl groups also mimics the anomeric carbon of the next unit in a polysaccharide chain. In the framework of our whole project, fully protected 1,6-anhydro containing trisaccharides will be used as synthetic intermediates and, in passing, Several studies have been carried out to understand the structure-activity relationship of heparin for heparanase inhibition, including variation in the degree of 2-O-sulfation, 6-O-sulfation, N-sulfation, N-acetylation, glycol-split uronic acid [13,14]. These studies led to conclude that O-2-sulfation of IdoA and O-6 sulfation of GlcN were not essential for effective inhibition, and that one N-sulfate group per disaccharide units was in interaction with the enzyme [7].…”