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Sphingomyelinase (SMase) is a hydrolase enzyme that is involved in sphingolipid metabolism. SMase is a member of the DNase I superfamily of enzymes and is responsible for the breakdown of sphingomyelin into phosphocholine and ceramide. The activation of SMase has been suggested as a major route for the production of ceramide in response to cellular stress.1 SMases are important in many physiological and pathophysiological processes, including: 1. lysosomal digestion of sphingomyelin, which is important for normal neuronal and vascular function; 2. ceramide-mediated signal transduction, leading to cytokine-induced apoptosis, cellular differentiation, and various immune and inflammatory responses; 3. lipoprotein aggregation within the vessel wall, which is a key event in atherogenesis; and 4. intracellular cholesterol trafficking and metabolism.2,3,4,5 Finding inhibitors to SMase (both acidic and neutral enzymes) could be beneficial in determining agents which reduce SMase activity and ceramide levels leading to attenuation of apoptosis and cellular proliferation, possible anti-depressant effects in depressive disorders, and beneficial clinical effects in acute or chronic neurodegenerative disorders, such as stroke and Alzheimer’s dementia.6,7,8 Synthesis of SMase inhibitors could be beneficial in the investigation and establishment of new therapeutic concepts for several diseases by using SMase as an actual target for new drug design.9 Cayman’s Sphingomyelinase Inhibitor Screening Assay provides a convenient method for screening sphingomyelinase inhibitors. This assay includes a neutral bacterial SMase. Even though bacterial SMase only shares a 20% sequence homology with mammalian neutral SMase, it has been shown to share a common catalytic site as the mammalian SMase and similar inhibition profile.10,11 Cleavage of the sphingomyelin conjugate by SMase results in the release of a ceramide analog containing a free thiol which is detected by the fluorescent SMase Detector. This fluorescence is analyzed with an excitation wavelength of 375-385 nm and an emission wavelength of 510-520 nm.12
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1
Hannun, Y.A., Obeid, L.M. The ceramide-centric universe of lipid-mediated cell regulation: Stress encounters of the lipid kind. The Journal of Biological Chemistry 277(29) 25847-25850 (2002).
2
Kolesnick, R.N. Sphingomyelin and derivatives as cellular signals. Prog Lipid Res 30(1) 1-38 (1991).
3
Tabas, I., Li, Y., Brocia, R.W., et al. Lipoprotein lipase and sphingomyelinase synergistically enhance the association of atherogenic lipoproteins with smooth muscle cells and extracellular matrix: A possible mechanism for low density lipoprotein and lipoprotein(a) retention and macrophage foam cell formation. The Journal of Biological Chemistry 268(27) 20419-20432 (1993).
4
Pörn, M.I., Slotte, J.P. Localization of cholesterol in sphingomyelinase-treated fibroblasts. Biochem J 308 269-274 (1995).
5
Smith, E.L., Schuchman, E.H. The unexpected role of acid sphingomyelinase in cell death and the pathophysiology of common diseases. FASEB J 22 3419-3431 (2008).
6
Gulbins, E., Li, P.L. Physiological and pathophysiological aspects of ceramide. Am J Physiol Regul Integr Comp Physiol 290 R11-R26 (2006).
7
Kolesnick, R. The therapeutic potential of modulating the ceramide/sphingomyelin pathway. J Clin Invest 110(1) 3-8 (2002).
8
Kornhuber, J., Medlin, A., Bleich, S., et al. High activity of acid sphingomyelinase in major depression. J Neural Transm 112 1583-1590 (2005).
9
Wascholowski, V., Giannis, A. Neutral sphingomyelinase as a target for drug design. Drug News Perspect 14(10) 581-590 (2001).
10
Clarke, C.J., Snook, C.F., Tani, M., et al. The extended family of neutral sphingomyelinases. Biochemistry 45(38) 11247-11256 (2006).
11
Luberto, C., Hassler, D.F., Signorelli, P., et al. Inhibition of tumor necrosis factor-induced cell death in MCF7 by a novel inhibitor of neutral sphingomyelinase. J Biol Chem 277(43) 41128-41139 (2002).
12
Hakogi, T., Fujii, S., Morita, M., et al. Synthesis of sphingomyelin sulfur analogue and its behavior toward sphingomyelinase. Bioorg Medicinal Chem Letters 15 2141-2144 (2005).
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