Sphingolipids have been known as lipid second messengers in mammalian cells and cell membranes, and a great deal of attention has been devoted to the studies of the biological process regulated by sphingolipids. Now, it has been well accepted that the sphingolipids play key roles in the cellular signal transmission pathway. Sphingomyelin, which is one of the key sphingolipids, is a ubiquitous constituent in animal tissue and has been known to occur in virtually every cell and in cell membranes. Ceramide and phosphoryl choline, which are the primary catabolites of sphingomyelin, are generated through the action of sphingomyelinase; ceramide is believed to display key roles as a signal transduction factor in cell differentiation and in programmed cell death (apoptosis) derivation (Figure 1). Although the significance of the sphingomyelin pathway, which is initiated by hydrolysis of sphingomyelin by SMase, has been well recognized, none of the tertiary-dimensional structures of these important enzymes have been determined and their hydrolytic mechanism has not been well-defined. It is, therefore, a very attractive challenge to reveal the catalytic mechanism of action of this important enzyme. Strong and selective sphingomyelinase inhibitors would contribute to a better understanding both of the roles of these enzymes and of ceramide in signal transduction. In order to elucidate the detailed catalytic mechanism of SMase, the development of the methods for supplying the sphingomyelin analogues, which competitively act at the catalytic site and strongly inhibit the hydrolytic ability of the enzyme, have strongly been desired. We then designed the substrate analogues 1, 2, 3, 4, 5 and 6 as inhibitor candidates on the basis of our previous results obtained on sphingomyelin analogues. In theses analogues, one of the oxygen atoms of the phosphoester, at which sphingomyelin is hydrolyzed by the enzyme, is replaced by either methylene, ethylene, difluoromethylene, nitrogen, or sulfur group, and the relative configuration of the asymmetric centers is (D)-erythro (3S, 4R) form (Figure 2). We will describe in detail the highly efficient stereocontrolled syntheses of the short chain substrate analogues 1, 2, 4, and 6 and their inhibitory activities toward B. cereus SMase.
