Abstract
Activation of the PI3K (phosphoinositide 3-kinase) pathway is a frequent occurrence in human tumors and is thought to promote growth, survival, and resistance to diverse therapies. Here, we report pharmacologic characterization of the pyridopyrimidinone derivative XL765 (SAR245409), a potent and highly selective pan inhibitor of class I PI3Ks (α, β, γ, and δ) with activity against mTOR. Broad kinase selectivity profiling of >130 protein kinases revealed that XL765 is highly selective for class I PI3Ks and mTOR over other kinases. In cellular assays, XL765 inhibits the formation of PIP(3) in the membrane, and inhibits phosphorylation of AKT, p70S6K, and S6 phosphorylation in multiple tumor cell lines with different genetic alterations affecting the PI3K pathway. In a panel of tumor cell lines, XL765 inhibits proliferation with a wide range of potencies, with evidence of an impact of genotype on sensitivity. In mouse xenograft models, oral administration of XL765 results in dose-dependent inhibition of phosphorylation of AKT, p70S6K, and S6 with a duration of action of approximately 24 hours. Repeat dose administration of XL765 results in significant tumor growth inhibition in multiple human xenograft models in nude mice that is associated with antiproliferative, antiangiogenic, and proapoptotic effects.
Highlights
Class I PI3 kinases convert phosphatidylinositol 4,5bisphosphate (PIP2) to phosphatidylinositol 3,4,5-trisphosphate (PIP3) in response to external cell stimuli [1, 2]
A similar assay format was used for DNAPK (DNA protein kinase; purchased from Promega) and VPS34 [PIK3C3; prepared at Exelixis as an N-terminal tagged full-length human fusion protein, which was expressed in insect cells using Baculovirus Expression Vector Systems (BEVS) and affinity purified using glutathione sepharose]
XL765 is a selective inhibitor of class I PI3Ks and of mTOR in biochemical assays
Summary
Class I PI3 kinases convert phosphatidylinositol 4,5bisphosphate (PIP2) to phosphatidylinositol 3,4,5-trisphosphate (PIP3) in response to external cell stimuli [1, 2]. Activation of class IA PI3Ks (phosphoinositide 3-kinases; PI3Ka, -b, and -d) is mediated by receptor tyrosine kinases (RTK). G-protein–coupled hormone receptors are implicated in activation of PI3Kb and class IB PI3K Ras, another important mediator of extracellular stimuli, can promote PI3K activation, and PI3K can mediate cellular transformation by Ras [4]. Downstream effectors of PI3K signaling, such as phosphoinositidedependent kinase-1 (PDK1) and AKT, bind to PIP3 at the cell membrane and are subsequently activated by phosphorylation [1]. PDK1 and AKT activate growth pathways, inhibit apoptotic signaling, and regulate transition through restriction points in the cell cycle via phosphorylation of their respective substrates [1]
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