title: Short heparin sequences spaced by glycol-split uronate residues are antagonists of fibroblast growth factor 2 and angiogenesis inhibitors.
abstract: Fibroblast Growth Factor-2 (FGF2) is a major inducer of neovascularization (angiogenesis). Heparin activates FGF2 by favoring formation of ternary complexes with its cellular receptors (FGFRs). Controlled 2-O-desulfation followed by exhaustive periodate oxidation/borohydride reduction has been used to generate sulfation gaps within the prevalent heparin sequences, building-up arrays of pentasulfated trisaccharides (PST, consisting of a 2-O-sulfated iduronic acid flanked by two N,6-disulfated glucosamines) spaced by reduced, glycol-split uronic acid (sU) residues. The structure of the prevalent sequences of the novel heparin derivative has been confirmed by mono- and two-dimensional NMR analysis. NMR spin-lattice relaxation times (T2) and nuclear Overhauser effects suggest that the sU residues act as flexible joints between the PST sequences and cause a marked distortion of the chain conformation of heparin required for formation of ternary complexes. Since the splitting reaction also occurs at the level of the essential glucuronic acid residue of the active site for antithrombin, the heparin derivative has no anticoagulant activity. However, it fully retains the FGF2-binding ability of the original heparin, as shown by its capacity to protect FGF2 from trypsin cleavage and to prevent the formation of heparan sulfate proteoglycan (HSPG)/FGF2/FGFR1 ternary complexes. However, when compared to heparin it showed a reduced capacity to induce FGF2 dimerization and to favor the interaction of [125I]FGF2 with FGFR1 in HSPG-deficient, FGFR1-transfected CHO cells. Accordingly, it was more effective than heparin in inhibiting the mitogenic activity exerted by FGF2 in cultured endothelial cells. Finally, it inhibited angiogenesis in a chick embrio chorioallantoic membrane (CAM) assay in which heparin is inactive.
authors.affiliation: G. Ronzoni Institute for Chemical and Biochemical Research, 20133 Milan, Italy.
authors.forename: Benito
authors.initials: B
authors.lastname: Casu
authors.forename: Marco
authors.initials: M
authors.lastname: Guerrini
authors.forename: Annamaria
authors.initials: A
authors.lastname: Naggi
authors.forename: Marta
authors.initials: M
authors.lastname: Perez
authors.forename: Giangiacomo
authors.initials: G
authors.lastname: Torri
authors.forename: Domenico
authors.initials: D
authors.lastname: Ribatti
authors.forename: Paolo
authors.initials: P
authors.lastname: Carminati
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Giannini
authors.forename: Sergio
authors.initials: S
authors.lastname: Penco
authors.forename: Claudio
authors.initials: C
authors.lastname: Pisano
authors.forename: Mirella
authors.initials: M
authors.lastname: Belleri
authors.forename: Marco
authors.initials: M
authors.lastname: Rusnati
authors.forename: Marco
authors.initials: M
authors.lastname: Presta
mesh_terms: D000482:Allantois / Q000098:blood supply / Q000502:physiology; D020533:Angiogenesis Inhibitors / Q000737:chemistry* / Q000502:physiology; D000818:Animals; D016466:CHO Cells; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D002460:Cell Line; D002642:Chick Embryo; D002823:Chorion / Q000098:blood supply / Q000502:physiology; D006224:Cricetinae; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology; D019855:Ethylene Glycol / Q000737:chemistry*; D005333:Fetus; D016222:Fibroblast Growth Factor 2 / Q000037:antagonists & inhibitors* / Q000502:physiology; D006025:Glycosaminoglycans / Q000737:chemistry; D006131:Growth Inhibitors / Q000502:physiology; D006493:Heparin / Q000031:analogs & derivatives* / Q000737:chemistry*; D006801:Humans; D007067:Iduronic Acid / Q000737:chemistry*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D013552:Swine; D014312:Trisaccharides / Q000737:chemistry
chemical_list: D020533:Angiogenesis Inhibitors; D006025:Glycosaminoglycans; D006131:Growth Inhibitors; D014312:Trisaccharides; D016222:Fibroblast Growth Factor 2; D007067:Iduronic Acid; D006493:Heparin; D019855:Ethylene Glycol