Studies on synthesis of inhibitors of sulfotransferases in mucopolysaccharide biosynthesis and application of the inhibitors to substrate deprivation therapy for mucopolysaccharidoses

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gem-Diamine 1-N-iminosugars related to L-iduronic acid and D-glucosamine were investigated to inhibit iduronate 2-O-sulfotransferase (2-O-ST) and N-deacetylase/N-sulfotransferase (NDST) for the GAG-chain biosynthesis in Hunter and Sanfilipo syndrome, respectively. Within the three years, about 60 compounds were synthesized and evaluated as inhibitors of 2-O-ST and NDST using an in vitro enzyme assay. Two iminosugars containing guanidine groups acted as potent inhibitors of 2-O-ST. Two inhibitors were subsequently tested in vivo as inhibitors of glycosaminoglycan biosynthesis using (i) biotinylated FGF-2 (ii) total ^<35>SO_4-labeling of the glycosaminoglycan chains and (iii) ^3H-GlcN-labeling of the glycosaminoglycan chains followed by disaccharide analysis. However, none of the active in vitro compounds were inhibitors of GAG biosynthesis in vivo. Both compounds were also found non-toxic to CHO-K1 cells. The lack of activity most likely reflects poor uptake by cells due to the positively charged guanidine moiety. These biological studies may require introduction of substituents and/or removal blocking groups to increase the permeability of the compounds to cell membranes.This is the first evidence that gem-diamine 1-N-iminosugars act as inhibitors of an enzyme involved in heparan sulfate synthesis. This suggests that the combination of substrate deprivation therapy and enzyme replacement therapy would also be anticipated.