Cancer cell cross-talk with the host endothelium plays a crucial role in metastasis, but the underlying mechanisms are still not fully understood. We studied the involvement of protein disulphide isomerase A1 (PDIA1) in human breast cancer cell (MCF-7 and MDA-MB-231) adhesion and transendothelial migration. For comparison, the role of PDIA1 in proliferation, migration, cell cycle and apoptosis was also assessed. Pharmacological inhibitor, bepristat 2a and PDIA1 silencing were used to inhibit

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... A1. Inhibition of PDIA1 by bepristat 2a markedly decreased the adhesion of breast cancer cells to collagen type I, fibronectin and human lung microvascular endothelial cells. Transendothelial migration of breast cancer cells across the endothelial monolayer was also inhibited by bepristat 2a, an effect not associated with changes in ICAM-1 expression or changes in cellular bioenergetics. The silencing of PDIA1 produced less pronounced anti-adhesive effects. However, inhibiting extracellular free thiols by non-penetrating blocker p-chloromercuribenzene sulphonate substantially inhibited adhesion. Using a proteomic approach, we identified that β1 and α2 integrins were the most abundant among all integrins in breast cancer cells as well as in lung microvascular endothelial cells, suggesting that integrins could represent a target for PDIA1. In conclusion, extracellular PDIA1 plays a major role in regulating the adhesion of cancer cells and their transendothelial migration, in addition to regulating cell cycle and caspase 3/7 activation by intracellular PDIA1. PDIA1-dependent regulation of cancer–endothelial cell interactions involves disulphide exchange and most likely integrin activation but is not mediated by the regulation of ICAM-1 expression or changes in cellular bioenergetics in breast cancer or endothelial cells.