Synthesis of Uronic Acid 1‐Azasugars as Putative Inhibitors of α‐Iduronidase, β‐Glucuronidase and Heparanase**

Abstract

1-Azasugar analogues of l-iduronic acid (l-IdoA) and d-glucuronic acid (d-GlcA) and their corresponding enantiomers have been synthesized as potential pharmacological chaperones for mucopolysaccharidosis I (MPS I), a lysosomal storage disease caused by mutations in the gene encoding α-iduronidase (IDUA). The compounds were efficiently synthesized in nine or ten steps from d-or l-arabinose, and the structures were confirmed by X-ray crystallographic analysis of key intermedi-ates. All compounds were inactive against IDUA, although l-IdoA-configured 8 moderately inhibited β-glucuronidase (β-GLU). The d-GlcA-configured 9 was a potent inhibitor of β-GLU and a moderate inhibitor of the endo-β-glucuronidase heparanase. Co-crystallization of 9 with heparanase revealed that the endocyclic nitrogen of 9 forms close interactions with both the catalytic acid and catalytic nucleophile.

Molecular Basis for Inhibition of Heparanases and β-Glucuronidases by Siastatin B

“…The costructure of 9 and HPSE has hydrogen bonds with the 4 eq -OH and Tyr391 and Asp62, as opposed to the hydrogen bond with water seen for 8 . The geminal diol 9 has the same hydrogen bonding network as the structure containing 11 , reported by Doherty et al, and additional H-bond between the 3 ax -OH with the nucleophile (Glu343), similar to what was observed in the active site of AcGH79. The 3 ax -OH in both structures is also within H-bonding distance of Asp62, an additional interaction that is not seen in the AcGH79-geminal diol structures.…”

“…As well, the hydroxyl on the β-face forms a hydrogen bond with Asp62 and the backbone nitrogen of Thr97, while the hydroxyl on the α-face is positioned to form hydrogen bonds with the catalytic nucleophile Glu343 and Asp62. The hydroxyl at the 4-position of inhibitor 3 lacks the hydrogen bond with Trp391 which is seen for gluco -configured inhibitors. , However, as in the costructure of 4 and BpHEP, the axial 4-hydroxyl forms a hydrogen bond with water present in the crystal structure.…”

“…The hydroxyl at the 4-position of inhibitor 3 lacks the hydrogen bond with Trp391 which is seen for gluco-configured inhibitors. 20,29 However, as in the costructure of 4 and BpHEP, the axial 4-hydroxyl forms a hydrogen bond with water present in the crystal structure. Synthesis of Siastatin B Breakdown Products.…”

“…The 2hydroxyl is within hydrogen bonding distance of the catalytic nucleophile and an active site asparagine in all three of these structures, thereby mimicking the interactions seen for natural substrates 28 and other inhibitors. 20,29 The 4-hydroxyl of 4 has galacto-stereochemistry in contrast to the gluco-configuration of the natural enzyme substrate; however, this position also appears to have productive binding to the active site. The 4hydroxyl of 4 forms hydrogen bond interactions with the enzyme active site, for both AcGH79 and EcGusB, see Figure 2AC.…”

“…Surprisingly, siastatin B also inhibits both β-glucuronidases and human heparanase, , which cleave β-glucuronic acid residues but not sialic acid or N -acetylated sugars. Although siastatin B resembles uronic acid-configured iminosugars [such as the β-glucuronidase inhibitors uronic-deoxynojirimycin ( 2 ), , uronic-noeurostegine ( 3 ), and uronic-isofagamine , ], it bears two major modifications that one would expect to interfere with β-glucuronidase activity. The first of these is the presence of the acetamido group at the 2-position.…”