Gene expression-phenotype association study reveals the dual role of TNF- α/TNFR1 signaling axis in confined breast cancer cell migration

Abstract While enhanced tumor cell migration is a key process in the tumor disemination, mechanistic insights into causal relationships between tumor cells and mechanical confinement are still limited. Here we combine the use of microfluidic platforms to characterize cell migration with genetic tools to systematically unravel the global signaling landscape associated with the migratory phenotype of breast cancer (BC) cells. The migration capacity of seven BC cell lines was evaluated in 3D microfluidic devices and their migration capacity was associated with their molecular signature. The gene expression of 715 genes was correlated with the migratory phenotype. TNF-α was found to be one of the most important upstream regulators of the signaling networks in which these 715 genes participate. Furthermore, the expression of the main TNF-α receptor, TNFR1, was strongly associated with the migration capacity, but also with a triple-negative, mesenchymal phenotype. In order to assess the impact of TNF-α/TNFR1 signaling axis on the BC migration capacity, either rhTNF-α was administrated or TNFR1,was silenced by siRNA, in four phenotypically diverse BC cell lines. In each experimental setting, the cell migration capacity was evaluated in microfluidic devices, while the molecular effects triggered by the treatment were monitored by RT-qPCR. According to our results, TNF-α stimulates the confined migration of triple negative, mesenchymal-like BC cells that are also characterized by high TNFR1 expression, but inhibits the migration of epithelial-like cells with low TNFR1 expression. Downstream of TNF/TNFR1 signaling, transcriptional regulation of NFKB seems to be dominant in driving cell migration in confined spaces.