Using Tandem Mass Spectrometry in Targeted Mode to Identify Activators of Class IA PI3K in Cancer
2011
Phosphatiditylinositide-3-kinase (PI3K) is activated in some cancers by direct mutation, but it is activated 7 more commonly in cancer by mutation of upstream acting receptor tyrosine kinases (TK). At present, there is no 8 systematic method to determine which TK signaling cascades activate PI3K in certain cancers, despite the likely 9 utility of such information to help guide selection of tyrosine kinase inhibitor (TKI) drug strategies for 10 personalized therapy. Here, we present a quantitative tandem mass spectrometry (LC/MS/MS) approach that 11 identifies upstream activators of PI3K both in vitro and in vivo. Using non–small cell lung carcinoma to illustrate 12 this approach, we show a correct identification of the mechanism of PI3K activation in several models, thereby 13 identifying the most appropriate TKI to downregulate PI3K signaling. This approach also determined the 14 molecular mechanisms and adaptors required for PI3K activation following inhibition of the mTOR kinase 15 TORC1. We further validated the approach in breast cancer cells with mutational activation of PIK3CA, where 16 tandem mass spectrometry detected and quantitatively measured the abundance of a helical domain mutant 17 (E545K) of PIK3CA connected to PI3K activation. Overall, our findings establish a mass spectrometric approach 18 to identify functional interactions that govern PI3K regulation in cancer cells. Using this technique to define the 19 pathways which activate PI3K signaling in a given tumor could help inform clinical decision making by helping 20 guide personalized therapeutic strategies for different patients. Cancer Res; 71(00); 1–11. � 2011 AACR.
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