Abstract A192: Proteomic analysis of epithelial-to-mesenchymal transition induced by EGF in breast cancer cells

Abstract

Abstract Metastasis is responsible for most of the deaths related to cancer. Therefore, the most effective methods to decrease cancer morbidity and mortality consist of early detection, prevention, and treatment of metastasis. Epithelial-to-mesenchymal transition (EMT) is one of the processes involved in cancer progression and metastasis. EMT induces cellular and microenvironmental changes that result in loss of the epithelial phenotype and acquisition of mesenchymal properties, thus promoting cellular invasive and migratory capabilities. EMT can be triggered by extracellular factors, including EGF and TGF-beta. Overexpression of transcription factors such as SNAIL also induce EMT and is correlated to cancer aggressiveness. Here, the breast adenocarcinoma cell line MCF7 was treated with the growth factor EGF (10 ng/mL) to identify proteomic alterations and specific mechanisms involved in the induction of EMT and metastasis. EGF treatment led to EMT, which was validated by molecular, morphologic, and functional experiments. Subcellular proteome enrichment followed by target proteomics was performed to identify changes in protein expression between the subcellular compartments cytoplasm, nucleus, and membrane. Thirty micrograms of each subcellular compartment of cells induced (or not) to EMT were processed to obtain tryptic peptide digest for subsequent MRM analysis. Proteotypic peptides representing relevant proteins for EMT and cancer progression, including proteins identified in our previous studies, were selected to compose panels for MRM analysis. EGF treatment activated the ERK/MAPK signaling pathway, which is involved in cancer progression by deregulating cell growth. Changes in expression of important proteins related to the EMT process were observed after EGF treatment, such as CDH1, CDH2, VIME, SOD2, and TGFB1. Subcellular compartment analysis allowed the identification of the main location of the proteins selected as well as differential regulation between subcellular compartments and potential mechanisms of subcellular translocation resulting from the EGF treatment and EMT induction, such as the regulation of PRDX3. Changes in the cellular protein localization may lead to alterations in the protein function and may thus be correlated with tumor progression. Other proteins related to cancer progression, including cytoskeleton rearrangement and cell cycle control, were also regulated after EGF treatment in specific compartments, such as CDK1, ARPC2, CKAP5, CNN3, and ARHGDIB. Besides that, we could identify the sumoylation as a possible post-translational modification regulated during EMT, which was verified by the expression of SUMO1/2 proteins, mainly in the cytoplasmic compartment. This detailed analysis allowed a better comprehension of the mechanisms of the EMT induction by EGF in a breast cancer cell line and can help to identify the biologic process and proteins relevant to cancer progression and that can potentially serve as targets for metastasis inhibition and/or diagnosis. (Financial support: FAPESP 2015/08693-0, 2011-0947-1, 2016/03809-3; CISBi NAP/USP grant 12.1.17598.1.3 and CTC-CEPID 2013/08135-2.) Citation Format: Camila S. Palma, Mariana L. Grassi, Carolina H. Thomé, Aline Poersch, Guilherme P. Lanfredi, Vitor M. Faça. Proteomic analysis of epithelial-to-mesenchymal transition induced by EGF in breast cancer cells [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A192.