title: Using tandem mass spectrometry in targeted mode to identify activators of class IA PI3K in cancer.
abstract: Phosphatiditylinositide-3-kinase (PI3K) is activated in some cancers by direct mutation, but it is activated more commonly in cancer by mutation of upstream acting receptor tyrosine kinases (TK). At present, there is no systematic method to determine which TK signaling cascades activate PI3K in certain cancers, despite the likely utility of such information to help guide selection of tyrosine kinase inhibitor (TKI) drug strategies for personalized therapy. Here, we present a quantitative liquid chromatography tandem mass spectrometry approach that identifies upstream activators of PI3K both in vitro and in vivo. Using non-small cell lung carcinoma to illustrate this approach, we show a correct identification of the mechanism of PI3K activation in several models, thereby identifying the most appropriate TKI to downregulate PI3K signaling. This approach also determined the molecular mechanisms and adaptors required for PI3K activation following inhibition of the mTOR kinase TORC1. We further validated the approach in breast cancer cells with mutational activation of PIK3CA, where tandem mass spectrometry detected and quantitatively measured the abundance of a helical domain mutant (E545K) of PIK3CA connected to PI3K activation. Overall, our findings establish a mass spectrometric approach to identify functional interactions that govern PI3K regulation in cancer cells. Using this technique to define the pathways that activate PI3K signaling in a given tumor could help inform clinical decision making by helping guide personalized therapeutic strategies for different patients.
authors.affiliation: Beth Israel Deaconess Medical Center, Division of Signal Transduction, Boston, Massachusetts 02115, USA.
authors.forename: Xuemei
authors.initials: X
authors.lastname: Yang
authors.forename: Alexa B
authors.initials: AB
authors.lastname: Turke
authors.forename: Jie
authors.initials: J
authors.lastname: Qi
authors.forename: Youngchul
authors.initials: Y
authors.lastname: Song
authors.forename: Brent N
authors.initials: BN
authors.lastname: Rexer
authors.forename: Todd W
authors.initials: TW
authors.lastname: Miller
authors.forename: Pasi A
authors.initials: PA
authors.lastname: Jänne
authors.forename: Carlos L
authors.initials: CL
authors.lastname: Arteaga
authors.forename: Lewis C
authors.initials: LC
authors.lastname: Cantley
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Engelman
authors.forename: John M
authors.initials: JM
authors.lastname: Asara
mesh_terms: D000818:Animals; D002289:Carcinoma, Non-Small-Cell Lung / Q000201:enzymology* / Q000235:genetics; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid / Q000379:methods; D058534:Class I Phosphatidylinositol 3-Kinases; D058543:Class Ia Phosphatidylinositol 3-Kinase / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology* / Q000235:genetics; D051379:Mice; D008819:Mice, Nude; D009154:Mutation; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000235:genetics; D020794:Receptor Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D020123:Sirolimus / Q000494:pharmacology; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000378:metabolism; D014162:Transfection
chemical_list: D058534:Class I Phosphatidylinositol 3-Kinases; D058543:Class Ia Phosphatidylinositol 3-Kinase; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D034741:RNA, Small Interfering; D020794:Receptor Protein-Tyrosine Kinases; D020123:Sirolimus; D014157:Transcription Factors; C500984:CRTC1 protein, human; C484760:PIK3CA protein, human