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A secondary assay for ceramide kinase inhibitors based on cell growth inhibition by short-chain ceramides.

Graf, Christine and Rovina, Philipp and Bornancin, Frederic (2009) A secondary assay for ceramide kinase inhibitors based on cell growth inhibition by short-chain ceramides. Analytical Biochemistry, 384 (1). pp. 166-169. ISSN 1096-0309

Abstract

We recently reported that ectopic expression of ceramide kinase (CerK) in various cell lines increases their sensitivity to cell death induced by the exogenous addition of short-chain (e.g., C2) ceramides (Cer). Here we show that this higher sensitivity results from CerK catalytic activity and production of C2-ceramide 1-phosphate (C2-C1P). If CerK activity is inhibited by the potent inhibitor NVP-231, C2-C1P is not produced and viability returns to control levels. The EC(50) of NVP-231 in this assay is in the low nanomolar range, consistent with the IC(50) determined in activity assays in vitro using purified CerK. NVP-995, a structurally related but inactive compound, does not protect against C2-Cer-induced cell death. This assay is robust and easy to implement and scale up, thereby providing a valuable secondary screen assay for CerK inhibitors.

Item Type: Article
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Additional Information: author can archive post-print (ie final draft post-refereeing); Publisher's version/PDF cannot be used
Keywords: Ceramide; Ceramide-1-phosphate; Cell death; Vesicle shedding; NVP-231; NVP-995
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Date Deposited: 14 Dec 2009 13:48
Last Modified: 31 Jan 2013 00:56
URI: https://oak.novartis.com/id/eprint/1331

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