Abstract 1320: Molecular imaging of EMT reveals crosstalk of multiple pathways and provides novel sites for therapeutic intervention of EMT

Abstract

Abstract Ionizing radiation therapy have been also suggested to result in increased incidence of local or distant metastasis which is, at least in part, mediated by TGFß signaling. TGFß is a known potentiator of epithelial to mesenchymal transition (EMT), a complex process by which a tumor cell bolsters its metastatic potential by transmogrifying into a highly motile and invasive cell that is ready to take on the challenge of distant travel and resettlement in a new milieu. Therefore it is important, in order to increase efficacy of ionizing radiation, a combination therapy along with inhibitors of EMT pathway, be utilized in the treatment of cancers. To date there is no clinically relevant EMT inhibitor and therefore to assist in the process of identifying a small molecular weight inhibitor, we developed three distinct non-invasive and quantitative molecular imaging reporters for monitoring EMT. These were based on changes in expression profile of E-cadherin and N-cadherin and complementation of luciferase based TGFβR kinase (BTR) activity. In the lung cancer EMT model studies using TGFβ treated A549 cells we demonstrated a time and dose dependent increase in activity of luciferase reporter driven by N-cadherin promoter and a decrease in the activity of a luciferase reporter driven by E-cadherin promoter. These changes were validated by phenotypic alteration of A549 cells and by western blotting for signaling intermediates of EMT pathway. BTR (bioluminescent TGFβR reporter), was a gain of function sensor of SMAD2/3 phosphorylation wherein inhibition of TGFβR signaling leads to increased reporter activity. Such reporters are extremely useful in high throughput screening (HTS) as cytotoxic agents are eliminated early in the screening process. Treatment of A549 cells stably expressing BTR with SB431542 (TGFβR specific inhibitor) lead to a more than 20 fold increase in BTR activity. All three reporters were adapted for HTS assay and were used to screen a library of known kinase inhibitors. Analyses and validation of hits suggested that the loss of epithelial or gain of mesenchymal phenotypes are independent processes. Further it also highlighted complex cross talks between EMT and different pathways involving mTor, Src, p38 MAPK, Akt, JNK, PI3-K, PKA, PKC, GSK-3β, PDGFRK and EGFR. Efficacy of several lead compounds (Erlotinib, SP600125, SB203580, and SB431542) in combination with radiotherapy is being accessed to increase radiation sensitivity in epithelial cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1320.