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

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Fernández Acero, Teresa y Rodríguez Escudero, Isabel y Vicente, Francisca y Monteiro, Maria Cândida y Tormo, José R. y Cantizani, Juan y Molina, María y 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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Resumen

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.


Tipo de documento:Artículo
Palabras clave:Microbiología
Palabras clave (otros idiomas):Microbiology
Materias:Ciencias Biomédicas > Farmacia > Microbiología
Código ID:21854
Depositado:15 Jun 2015 11:10
Última Modificación:30 Jun 2015 08:00

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