Synthesis of fluorescent and radioactive analogues of two lactosylceramides and glucosylceramide containing β-thioglycosidic bonds that are resistant to enzymatic degradation

Condensation of 2-S-(2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-2-thiopseudourea hydrobromide with 2,3,6-tri-O-benzoyl-4-O-trifluoromethylsulfonyl-β-d-galactopyranosyl-(1 → 1)-(2S,3R,4E)-3-O-benzoyl-2-dichloroacetamido-4-octadecen-1,3-diol afforded S-(2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl)-(1 → 4)-2,3,6-tri-O-benzoyl-4-thio-β-d-glucopyranosyl-(1 → 1)-(2S, 3R,4E)-3-O-benzoyl-2-dichloroacetamido-4-octadecen-1,3-diol in good yield. Removal of the protecting groups, followed by selective N-acylation of the sphingosine amino group with either a fluorescent or a radioactive fatty acid, gave labeled lactosylceramide analogues in good yield. Since these products contained a β-thioglycosidic bond between the two sugar moieties, they were totally resistant to the action of acid lysosomal glycosidases. Likewise, condensation of 2-S-(2,3,4,6-tetra-O-acetyl-β-d-glucopyranosyl)-2-thiopseudourea hydrobromide and 2,3,4,6-tetra-O-acetyl-β-d-galactopyranosyl-(1 → 4)-2,3,6-tri-O-acetyl-1-S-acetyl-1-thio-β-d-glucopyranose with (2R,3R,4E)-3-O-benzoyl-2-dichloroacetamido-1-iodo-4-octadecen-3-ol in methanolic sodium acetate afforded the corresponding β-thioglycosides 14 and 16, respectively, in good yield. These β-thioglycosides were converted into glucosylceramide and lactosylceramide analogues following removal of the protecting groups and by subsequent selective N-acylation using either a fluorescent or radioactive fatty acid N-succinimidyl ester. Whereas the glucosylthioceramides thus obtained proved to be completely undegradable by lysosomal glucocerebrosidase, the lactosylceramides containing the β-thioglycosidic bond between the lactose and the ceramide residues could be degraded by lysosomal GM1-β-galactosidase to give the corresponding glucosylthioceramides. These compounds did not yield to any further enzymatic degradation.