Solid- and solution-phase synthesis of heparin and other glycosaminoglycans

Abstract

Described is a modular, general synthetic strategy for the preparation in solution and on a solid support of heparin, heparin-like glycosaminoglycans, glycosaminoglycans and non-natural analogs of each of them. Additionally, the modular strategy provides the basis for the preparation of combinatorial libraries and parallel libraries of defined glycosaminoglycan oligosaccharides. The defined glycosaminoglycan structures may be used in high-throughput screening experiments to identify carbohydrate sequences that regulate a host of recognition and signal-transduction processes. The determination of specific sequences involved in receptor binding holds great promise for the development of molecular tools which will allow modulation of processes underlying viral entry, angiogenesis, kidney diseases and diseases of the central nervous system. Notably, the present invention enables the automated synthesis of glycosaminoglycans in much the same fashion that peptides and oligonucleotides are currently assembled.

1 - 5 . (canceled)  
     
     
         6 . A trisaccharide selected from the group consisting of:  
       
         
           
           
               
               
           
         
         wherein  
         X represents independently for each occurrence hydroxyl, acyloxy, silyloxy, halide, alkylthio, arylthio, alkoxy, aryloxy, or —OC(NH)CCl 3 ;  
         R represents independently for each occurrence H, alkyl, aryl, arylalkyl, heteroarylalkyl, silyl, acyl, alkenyloxycarbonyl, or aralkyloxycarbonyl; and  
         R′ represents independently for each occurrence H, alkyl, aryl, arylalkyl, or heteroarylalkyl.  
       
     
     
         7 . The trisaccharide of  claim 6 , wherein X represents fluoro, bromo, 4-pentenyloxy or —OC(NH)CCl 3 .  
     
     
         8 . The trisaccharide of  claim 6 , wherein R′ represents independently for each occurrence alkyl.  
     
     
         9 . The trisaccharide of  claim 6 , wherein X represents fluoro, bromo, 4-pentenyloxy or —OC(NE)CCl 3 ; and R′ represents independently for each occurrence alkyl.  
     
     
         10 . The trisaccharide of  claim 6 , wherein said trisaccharide is selected from the group consisting of:  
       
         
           
           
               
               
           
         
         wherein  
         X is silyloxy or —OC(NH)CCl 3 ; and  
         R is H or silyloxy.  
       
     
     
         11 . A method of preparing a glycosaminoglycan, comprising the step of: 
 reacting a first mono-, di- or tri-saccharide, comprising an activated anomeric carbon, with a second mono-, di- or tri-saccharide, comprising a hydroxyl or amino group, to form an oligosaccharide, comprising a glycosidic linkage between said anomeric carbon of said first mono-, di- or tri-saccharide and said hydroxyl or amino group of said second mono-, di- or tri-saccharide.    
     
     
         12 . The method of  claim 11 , wherein the first mono-, di- or tri-saccharide is not identical to the second mono-, di- or tri-saccharide.  
     
     
         13 . The method of  claim 11 , wherein neither the first mono-, di- or tri-saccharide nor the second mono-, di- or tri-saccharide is covalently linked to a solid support.  
     
     
         14 . The method of  claim 11 , wherein the first first mono-, di- or tri-saccharide or the second mono-, di- or tri-saccharide is covalently linked to a solid support.  
     
     
         15 . The method of  claim 14 , further comprising the step of: 
 cleaving said covalent linkage between said oligosaccharide and said solid support with an alkene metathesis catalyst and an alkene.    
     
     
         16 . The method of claim  1   1 , further comprising the step of: 
 sulfating a hydroxyl or amino moiety of said oligosaccharide.    
     
     
         17 . The method of  claim 11 , further comprising the step of: 
 removing a hydroxyl or amino protecting group from said oligosaccharide by hydrogenolysis.    
     
     
         18 - 22 . (canceled)