PO-234 Transwell co-culture with human breast adipocytes alters the proteome expression profiles of MCF-7 and MDA-MB-231 human breast cancer cells

Abstract

IntroductionBreast tumours grow within adipose tissue, and stromal adipocytes are likely the first cell type encountered by breast cancer cells as they metastasize. Adipocytes at the invasive edge of human breast tumours exhibit a modified phenotype, and consequently these adipocytes have been named Cancer Associated Adipocytes (CAA). Previous in vitro research has shown that co-culture with CAA promotes breast cancer cell resistance to radiation and some therapeutic agents, as well as breast cancer cell invasiveness both in vitro and in vivo. The aim of the current research was to identify cellular proteins that are differentially regulated in breast cancer cells co-cultured with CAA.Material and methodsThis study collected human breast adipose tissue samples, from which pre-adipocytes were isolated, differentiated into mature adipocytes, and co-cultured with hormone receptor positive (MCF-7) and negative (MDA-MB-231) human breast cancer cells for 3 days in a transwell co-culture system. The proteomes of co-cultured and control breast cancer cells were compared quantitatively using iTRAQ labelling and LC-coupled LTQ-Orbitrap tandem mass spectrometry. Validation of iTRAQ results was performed by Western blotting.Results and discussionsThis study identified and quantified a total of 1126 and 1218 proteins expressed in MCF-7 and MDA-MB-231 cells, respectively. Of these, 85 in MCF-7 and 63 in MDA-MB-231 had a fold change >1.5 between co-culture and control breast cancer cell samples. Overall, MCF-7 cells had more proteins downregulated (n=53) than upregulated (n=32) after co-culture, whereas, more proteins were upregulated (n=51) than downregulated (n=12) in MDA-MB-231 cells. Enrichment analysis revealed an upregulation of proteins involved in metabolic pathways, namely TCA cycle proteins in MCF-7 cells and glycolysis proteins in MDA-MB-231 cells. The metabolic enzyme PGK1 was upregulated in response to co-culture with CAA in both cell lines, and is associated with poorer overall survival and resistance to the chemotherapy agent paclitaxel in breast cancer. Western blotting results validated mass spectrometry ratios for three candidate proteins.ConclusionOverall, this study describes for the first time the local effect CAA have on the expression of proteins in breast cancer cells in vitro, and provides a comprehensive platform for further research investigating local interactions between breast cancer cells and CAA.