Generation of Specific Deoxynojirimycin-type Inhibitors of the Non-lysosomal Glucosylceramidase

Abstract

The existence of a non-lysosomal glucosylceramidase in human cells has been documented (van Weely, S., Brandsma, M., Strijland, A., Tager, J. M., and Aerts, J. M. F. G. (1993) Biochim. Biophys. Acta 1181, 55-62). Hypothetically, the activity of this enzyme, which is localized near the cell surface, may influence ceramidemediated signaling processes. To obtain insight in the physiological importance of the non-lysosomal glucosylceramidase, the availability of specific inhibitors would be helpful. Here we report on the generation of hydrophobic deoxynojirimycin (DNM) derivatives that potently inhibit the enzyme. The inhibitors were designed on the basis of the known features of the non-lysosomal glucosylceramidase and consist of a DNM moiety, an N-alkyl spacer, and a large hydrophobic group that promotes insertion in membranes. In particular, N-(5-adamantane-1-yl-methoxy)pentyl)-DNM is a very powerful inhibitor of the non-lysosomal glucosylceramidase at nanomolar concentrations. At such concentrations, the lysosomal glucocerebrosidase and ␣-glucosidase, the glucosylceramide synthase, and the N-linked glycantrimming ␣-glucosidases of the endoplasmic reticulum are not affected.

Lysosomal glycosphingolipid catabolism by acid ceramidase: formation of glycosphingoid bases during deficiency of glycosidases

“…Fibroblasts contain very little nonlysosomal b-glucosidase GBA2. We nevertheless checked the effect of selective inhibition of GBA2 with low nanomolar AMP-DNM [17]. No changes in the formation 13 C 5 -GlcSph were detected (Fig.…”

Klotho-related Protein Is a Novel Cytosolic Neutral β-Glycosylceramidase

“…Alternatively, GlcCer was found to be accumulated in the testes, brain, and liver, causing male infertility possibly because of abnormal sperm (8). GBA2 was likely to be the same enzyme that was previously described as a nonlysosomal CBE-insensitive glucosylceramidase (11), which was extremely sensitive to inhibition by hydrophobic deoxynojirimycin analogues (12).…”

Biochemistry of glycosphingolipid storage disorders: implications for therapeutic intervention

“…Our research group discovered the occurrence of non-lysosomal glycosylceramide catabolism several years ago (Van Weely et al 1993b). The enzyme activity is not affected in Gaucher patients and shows a different inhibitor specificity, all pointing to a product from a distinct gene (Overkleeft et al 1998). It remains unclear to what extent the non-lysosomal glucosylceramidase plays a role in the pathophysiology of Gaucher cells.…”

“…However, it is also conceivable that excessive extra-lysosomal glucosylceramide catabolism results in excessive production of cytosolic ceramide that may act as a signalling molecule and cause abnormal cell behaviour (see figure 4). It is of interest to note that the hydrophobic deoxynojirimycins are also potent inhibitors of the non-lysosomal glucosylceramidase (Overkleeft et al 1998). Current approaches of substrate deprivation are consequently also likely to inhibit the non-lysosomal catabolism of glucosylceramide.…”

“…Overkleeft and co-workers, in their search for inhibitors of glucosidases, have serendipitously developed a more potent inhibitor of glucosylceramide synthase. Adamantane-pentyl-deoxynojirimycin was found to inhibit glycosphingolipid biosynthesis at low nanomolar concentrations (Overkleeft et al 1998) and was able to prevent globotriaosylceramide accumulation in a Fabry knockout mouse model without overt side effects (D. Copeland, personal communication). The first clinical study of the use of NB-DNJ to treat a glycosphingolipid storage disorder has been reported recently .…”