Targeting Ceramide Metabolism with a Potent and Specific Ceramide Kinase Inhibitor

  1. Christine Graf,
  2. Martin Klumpp,
  3. Michael Habig,
  4. Philipp Rovina,
  5. Andreas Billich,
  6. Thomas Baumruker,
  7. Berndt Oberhauser and
  8. Frédéric Bornancin
  1. Novartis Institutes for BioMedical Research, Vienna, Austria (C.G., P.R., A.B., T.B., B.O., F.B.), and Basel, Switzerland (M.K., M.H.)
  1. Address correspondence to:
    Frédéric Bornancin, Current affiliation: Novartis Institutes for BioMedical Research, Forum 1, CH-4056 Basel, Switzerland. E-mail: frederic.bornancin{at}novartis.com

Abstract

Ceramide kinase (CerK) produces the bioactive lipid ceramide-1-phosphate (C1P) and appears as a key enzyme for controlling ceramide levels. In this study, we discovered and characterized adamantane-1-carboxylic acid (2-benzoylamino-benzothiazol-6-yl)amide (NVP-231), a potent, specific, and reversible CerK inhibitor that competitively inhibits binding of ceramide to CerK. NVP-231 is active in the low nanomolar range on purified as well as cellular CerK and abrogates phosphorylation of ceramide, resulting in decreased endogenous C1P levels. When combined with another ceramide metabolizing inhibitor, such as tamoxifen, NVP-231 synergistically increased ceramide levels and reduced cell growth. Therefore, NVP-231 represents a novel and promising compound for controlling ceramide metabolism that may provide insight into CerK physiological function.

Footnotes

  • ABBREVIATIONS: CerK, ceramide kinase; C1P, ceramide-1-phosphate; MOPS, 3-(N-morpholino)propanesulfonic acid; DMSO, dimethyl sulfoxide; TLC, thin-layer chromatography; LC/MS, liquid chromatography/mass spectrometry; BMDM, bone marrow-derived macrophages; 7-AAD, 7-aminoactinomycin D; GlcCer, glucosylceramide; SM, sphingomyelin; M-CSF, macrophage colony stimulating factor; PLA2G4A, cytosolic phospholipase A2 α; NBD, N-[7-(4-nitrobenzo-2-oxa-l,3-diazole)]; NBD-Cer, NBD-C6-ceramide; NVP-231, adamantane-1-carboxylic acid (2-benzoylamino-benzothiazol-6-yl)amide.

  • ↵Graphic The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.

    • Received May 6, 2008.
    • Accepted July 1, 2008.
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  1. Published online before print July 8, 2008, doi: 10.1124/mol.108.048652 Molecular Pharmacology October 2008 vol. 74 no. 4 925-932
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