Highly diastereoselective additions to polyhydroxylated pyrrolidine cyclic imines: Ready elaboration of aza-sugar scaffolds to create diverse carbohydrate-processing enzyme probes

Representative diastereomeric. erythritol and threitol polyhydroxylated pyrrolidine imine scaffolds have been rapidly elaborated to diversely functionalized aza-sugars through highly diastereoselective organometallic (RM) additions (R=Me, Et, allyl, hexenyl. Ph, Bn. pMeO-Bn). The yields for these additions have all been substantially enhanced from previously optimised levels (<58%) for normal additions using, a reverse addition procedure (e.g. R = Ph: 44% normal mode --> 78% reverse mode). The high diastereoselectivities (>98% de for all except R Me) are consistent with additions that are controlled by the configuration of the C-2 centre adjacent to the azomethine imine carbon and the conformation of the pyrrolidine imine. The high potential of this method was demonstrated by concise syntheses of 1-epi- and 2-epi-desacetylanisomycins. In addition, the late stage addition of hydrophobic substituents, which this imine addition methodology allows, enabled the preparation of novel aza-sugars with enhanced inhibitory potential. This was highlighted by the screening of a representative selection of these "hydrophobically-modified" aza-sugars against a diverse panel of 12 non-mammalian and human carbohydrate-processing enzymes. This identified a novel nanomolar alpha-galactosidase inhibitor (IC50 = 250nM) and a novel highly selective glucosylceramide synthase inhibitor (IC50 = 52 muM, no alpha-glucosidase inhibition at 1 mm). Furthermore, analysis of the structure-activity relationships of racemic series of inhibitors allowed some validation of Fleet's mirror-image enzyme active site postulate.