Human kinome siRNA screening to identify novel targets implicated in fibroblast-mediated triple negative breast cancer cells invasion

Introduction: Triple negative Breast-Cancer (TNBC) represents the breast cancer subtype with the poorest prognosis, which lacks targeted treatment options. Despite the increasing evidence that tumor microenvironment signalling influences the behaviour of surrounding cancer cells, still relatively little is known about which changes in stromal cells influence tumor cells behaviour. Kinases are proteins by which signals are transmitted inside and outside the cells. Alterations in kinases expression could lead to dramatic changes in single cells and ultimately the surrounding microenvironment. Future treatment design for TNBC should provide tools to neutralize not only tumorigenic cells but also the cancer-associated microenvironment cells. The aim of this study is to identify kinases expressed in fibroblasts that able to influence the aggressiveness / invasive potential of TNBC cells. Material and methods: A library of siRNAs targeting human kinase were used in fibroblast , after silencing fibroblast were co-culturing with TNBC cell line (MDA-MB231). After spheroids formation, Matrigel and chemoattractant were added to the well in order to promote invasion. Imaging of wells after 3 and 6 days was performed in order to measure invasion and pictures were analysed with Image-J software. Results and discussion: We were able to identify n=33 kinases that were able to significantly (p<0.05) influence the breast cancer cells invasion profile. The Gene Ontology analysis of those kinases shows that the one of main pathways involved in tumor-stroma cross-talk is angiogenesis. Among selected kinases, TIE1 (tyrosine kinase with immunoglobulin-like and EGF-like domains 1) which silencing was able to reduce invasion by 25%, in an independent analysis was found to be overexpressed in cancer-associated fibroblast (CAF) as compared to normal ones, suggesting that it can be involved in both invasion promotion and in fibroblast transition to CAF. Conclusion: We have identified new kinases that are expressed in normal fibroblasts, which appear to be implicated in breast cancer cells behaviour. These kinases can represent a putative molecular target to treat TNBC, that to date lack of an effective pharmacological. We plan to performed additional studies (include exosome and SILAC analysis ) to further elucidated the role of these Kinases in tumor-stroma cross talk and the influence on invasion.