title: Ultrasensitive and absolute quantification of the phosphoinositide 3-kinase/Akt signal transduction pathway by mass spectrometry.
abstract: The phosphoinositide 3-kinase (PI3K)/Akt pathway controls a vast array of normal physiological processes and is frequently aberrantly activated in cancer, thus identifying PI3K/Akt-signaling components as promising drug targets in oncology. However, implementation of rational cancer therapies for this pathway needs robust and simple tools to stratify patients according to PI3K pathway activation and to validate and measure the impact of targeted inhibition on primary cancer tissues. Herein we present a technique for the quantification of the PI3K/Akt-signaling pathway based on the mass spectrometric measurement of PI3K-dependent protein kinase activity in cell lysates. The concept of this application of MS is to exploit enzymatic activity to amplify the signal of the enzyme under study analogous to the PCR used to amplify nucleic acid sequences. We show that this approach allows quantitative analysis of a cell-signaling pathway with high sensitivity, precision of quantification, and specificity. Due to its special analytical capabilities and potential for multiplexing, this approach could contribute significantly to cell-signaling studies and to the development and implementation of personalized cancer therapies.
authors.affiliation: Cell Signaling in Cancer Laboratory, Ludwig Institute for Cancer Research, 91 Riding House Street, London W1W 7BS, United Kingdom. pedro@ludwig.ucl.ac.uk
authors.forename: Pedro R
authors.initials: PR
authors.lastname: Cutillas
authors.forename: Asim
authors.initials: A
authors.lastname: Khwaja
authors.forename: Mariona
authors.initials: M
authors.lastname: Graupera
authors.forename: Wayne
authors.initials: W
authors.lastname: Pearce
authors.forename: Severine
authors.initials: S
authors.lastname: Gharbi
authors.forename: Mike
authors.initials: M
authors.lastname: Waterfield
authors.forename: Bart
authors.initials: B
authors.lastname: Vanhaesebroeck
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000378:metabolism; D007554:Isotopes / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D051379:Mice; D009369:Neoplasms / Q000201:enzymology; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000737:chemistry / Q000378:metabolism*; D012680:Sensitivity and Specificity; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D004791:Enzyme Inhibitors; D007554:Isotopes; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt