Saccharomyces cerevisiae as a platform for assessing sphingolipid lipid kinase inhibitors

Date
2018-04-19Author
Kharel, Yugesh
Agah, Sayeh
Huang, Tao
Mendelson, Anna J.
Eletu, Oluwafunmilayo T.
Barkey-Bircannl, Peter
Gesualdil, James
Smith, Jeffrey S.
Santos, Webster L.
Lynch, Kevin R.
Metadata
Show full item recordAbstract
Successful medicinal chemistry campaigns to discover and optimize sphingosine kinase inhibitors require a robust assay for screening chemical libraries and for determining rank order potencies. Existing assays for these enzymes are laborious, expensive and/or low throughput. The toxicity of excessive levels of phosphorylated sphingoid bases for the budding yeast, Saccharomyces cerevisiae, affords an assay wherein inhibitors added to the culture media rescue growth in a dose-dependent fashion. Herein, we describe our adaptation of a simple, inexpensive, and high throughput assay for assessing inhibitors of sphingosine kinase types 1 and 2 as well as ceramide kinase and for testing enzymatic activity of sphingosine kinase type 2 mutants. The assay was validated using recombinant enzymes and generally agrees with the rank order of potencies of existing inhibitors.