Abstract 1026: Correlation of complex signaling pathway interactions with cell migration in a combined phosphorylation-cell migration assay

Abstract The acquisition of the ability of a cell to migrate, a process critical to tumor progression, is dependent on the activation of key cell signaling pathways. Screening assays that simultaneously quantify cell migration and the activity of the underlying signaling kinases may facilitate more efficient drug development. We report here an assay that allows simultaneous assessment of cell migration and pathway activity. The assay was performed in a commercially available microtiter plate and used near-infrared detection to achieve high sensitivity and reproducible results. The quality of the assay as determined by Z-factor analysis was 0.7 for migration and 0.58 for detection of phosphorylation. To model the drug screening process, we examined the effects of inhibitors of IGF-1R, PDGFR, and EGFR on cell migration and phosphorylation of AKT and ERK. An IGF-1R inhibitor strongly decreased both cell migration and phosphorylation of AKT and ERK. A PDGFR inhibitor showed a similar effect, but to a lesser degree. Inhibitors of PI-3 kinase/AKT and MEK also decreased cell migration but had surprising effects on the phosphorylation of AKT and ERK. Inhibition of MEK reduced phosphorylation of ERK to undetectable levels, yet did not completely abolish cell migration. Instead, AKT phosphorylation increased significantly. Likewise, inhibition of AKT decreased but did not abolish migration and increased ERK phosphorylation. The combination assay effectively assessed both migration and pathway activity as well as pathway crosstalk, a feature that should be useful for drug development. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1026. doi:10.1158/1538-7445.AM2011-1026