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