A yeast-based in vivo bioassay to screen for class I phosphatidylinositol 3-kinase specific inhibitors.



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Fernández Acero, Teresa and Rodríguez Escudero, Isabel and Vicente, Francisca and Monteiro, Maria Cândida and Tormo, José R. and Cantizani, Juan and Molina, María and Cid, Víctor J. (2012) A yeast-based in vivo bioassay to screen for class I phosphatidylinositol 3-kinase specific inhibitors. Journal of biomolecular screening, 17 (8). pp. 1018-1029. ISSN 1552-454X

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The phosphatidylinositol 3-kinase (PI3K) pathway couples receptor-mediated signaling to essential cellular functions by generating the lipid second messenger phosphatidylinositol-3,4,5-trisphosphate. This pathway is implicated in multiple aspects of oncogenesis. A low-cost bioassay that readily measures PI3K inhibition in vivo would serve as a valuable tool for research in this field. Using heterologous expression, we have previously reconstituted the PI3K pathway in the model organism Saccharomyces cerevisiae. On the basis of the fact that the overproduction of PI3K is toxic in yeast, we tested the ability of commercial PI3K inhibitors to rescue cell growth. All compounds tested counteracted the PI3K-induced toxicity. Among them, 15e and PI-103 were the most active. Strategies to raise the intracellular drug concentration, specifically the use of 0.003% sodium dodecyl sulfate and the elimination of the Snq2 detoxification pump, optimized the bioassay by enhancing its sensitivity. The humanized yeast-based assay was then tested on a pilot scale for high-throughput screening (HTS) purposes using a collection of natural products of microbial origin. From 9600 extracts tested, 0.6% led to a recovery of yeast growth reproducibly, selectively, and in a dose-dependent manner. Cumulatively, we show that the developed PI3K inhibition bioassay is robust and applicable to large-scale HTS.

Item Type:Article
Uncontrolled Keywords:Microbiología
Palabras clave (otros idiomas):Microbiology
Subjects:Medical sciences > Pharmacy > Microbiology
ID Code:21854
Deposited On:15 Jun 2015 11:10
Last Modified:30 Jun 2015 08:00

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