Abstract 4805: Distinctive phospho-proteome signatures in non-small cell lung carcinoma tumors

Non-small cell lung carcinoma (NSCLC) is a major, lethal cancer worldwide. Various molecular and cellular abnormalities are found in NSCLC, and an established theme is the alteration of protein phosphotyrosine (pY), as exemplified by elevated epidermal growth factor receptor (EGFR) activity recognized in a subset of NSCLC. In order to gain a broad perspective of pY-mediated signaling networks in NSCLC, mass spectrometry was used to analyze pY-containing peptides affinity purified from protease-digested xenograft tumors. Seventeen tumors representing the major NSCLC subtypes adenocarcinoma (ADC) and squamous cell carcinoma (SCC) were characterized. The major classes of pY proteins detected were cytoskeletal, protein kinases, and adhesion proteins. Pathways and cellular functions enriched in the data set included focal adhesion, adherens junction, and ErbB signaling. Five receptor tyrosine kinases (RTKs) were detected, with EGFR and EPHA2 most prevalent. Src-family kinases (YES, LYN, FRK and LCK) and focal adhesion kinase (FAK) comprised the majority of non-receptor tyrosine kinases measured. Analysis of individual pY sites on 14 tyrosine kinases indicates their activation by kinase domain activation loop phosphorylation, but also revealed additional pY sites suggestive of regulation by trafficking/endocytosis and inter- and intra-molecular protein-protein interactions involving, for example, SH2 and SH3 domains, and extra-kinase domain regions. Analysis of tumors based on protein-pY suggested subtypes distinguished by their activated tyrosine kinase networks. Genetic analysis of 3 tumors that displayed EGFR-associated phospho-proteome signatures confirmed that they carried either EGFR activating mutations (in two ADCs) or increased gene copy number (in one SCC). Western blot analysis of protein pY confirmed different patterns of cellular protein pY among the tumor phospho-types in agreement with the phospho-proteomic analysis. Quantification of kinase and substrate pY sites, according to their integrated extracted ion currents, revealed a set of phosphorylations that constituted a molecular signature that differentiated between the ADC and SCC subtypes. In summary, quantitative phospho-proteomic profiling of NSCLC tumors is feasible and may have utility in the categorization of individual tumors according to their networks of activated tyrosine kinases and substrates. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4805. doi:1538-7445.AM2012-4805