Preparation, biochemical characterization and biological properties of radiolabelled N-alkylated deoxynojirimycins

Abstract

We have reductively alkylated deoxynojirimycin imino sugars using sodium cyanoborohydride to provide an efficient means of generating a series of N-alkylated compounds containing 4–18 carbon side chains. The yields were greater than 90% using a variety of aldehydes of different chain lengths, and after purification were >95% pure using 1H-NMR. Radiolabelled compounds were prepared using sodium cyanoborotriti-ide to selectively label the first carbon atom in the alkyl chain and used in protein-binding and cell- and tissue-uptake experiments. Protein binding was chain-length-dependent with compounds of intermediate chain length (C9—C12), demonstrating an equal distribution between the aqueous and protein-bound phase. The extent of cell uptake also increased proportionally with increased chain length in a time-dependent manner. When administered to mice, the longer alkyl-chain compounds showed reduced absorption from the intestine and a marked deposition of compound in the liver and brain, suggesting that the more hydrophobic compounds were poorly cleared by the major tissues. In tissue-culture cells compounds with 8 or fewer carbon atoms were non-toxic and had CC50 (the concentration at which the number of cells or cell proliferation is reduced by 50%) values greater than 1mM. Compounds with chain lengths above C8 showed a chain-length-dependent increase in cytotoxicity. N-alkylated deoxynojirimycins (C4—C18) were evaluated for their inhibitory effects on ceramide-specific glucosyltransferase and glycoprotein-processing α-glucosidase. Increasing the alkyl chain length had little effect on α-glucosidase activity, but inhibition of ceramide-specific glucosyltransferase increased 10-fold when C4 and C9—C18 compounds were compared. Overall these data provide further definition of the molecular features of alkylated imino sugars that influence tissue selectivity and efficacy for cellular enzyme inhibition.