Protein Disulphide Isomerase A1 Is Involved in the Regulation of Breast Cancer Cell Adhesion and Transmigration via Lung Microvascular Endothelial Cells.

Joanna Suraj-Prazmowska,Joanna Banach,Marta Stojak,Joanna Wietrzyk,Kamila Wojnar-Lason,Ivars Kalviņš,Martyna Stachowicz-Suhs,Joanna Rossowska,Stefan Chlopicki,Anna Kurpinska,Magdalena Milczarek,Patrycja Kaczara

doi:10.3390/cancers12102850

Abstract

Simple SummaryMetastasis is one of the most devastating aspect of cancer progression and involves biochemical and physical interactions between cancer cells and surrounding microenvironment. In particular, cancer cells adhesion to endothelium and their subsequent transendothelial migration represent important steps in the metastatic process in target organs. In this study we characterized the functional role of Protein Disulfide Isomerase A1 (PDIA1) in breast cancer cells adhesion and transendothelial migration. We identified the full repertoire of protein disulfide isomerases in endothelial cells as well as in breast cancer cells. We provided insight into the mechanisms involved in cancer-endothelial cells interactions and suggested that PDIA1 regulates the adhesion and transendothelial migration of breast cancer cells by disulphide exchange involving most likely the activation of integrins. Our results suggest that the inhibition of extracellular PDIA1 or other PDIs represents an interesting target for anti-metastatic treatment. 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 PDIA1. 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.

Highlights

IntroductionThe major cause of cancer patient death [1], is a complex phenomenon
Cancer cell metastasis, the major cause of cancer patient death [1], is a complex phenomenon.It involves cancer cell invasion at the primary site, survival and arrest in the bloodstream, and extravasation at a distant site
We explored whether the effects of protein disulphide isomerase A1 (PDIA1) inhibition on cancer–endothelial cell interactions could be attributed to disulphide re-arrangement of integrins known to mediate adhesion of platelets and leukocytes to endothelium [7,26,27,28], to cellular bioenergetics in breast cancer or endothelial cells, an important target for anti-adhesive mechanisms [29], or to alterations in ICAM-1 expression involved in this interaction [30]

Summary

Introduction

The major cause of cancer patient death [1], is a complex phenomenon. It involves cancer cell invasion at the primary site, survival and arrest in the bloodstream, and extravasation at a distant site. Organ-specific colonisation is a key feature of most metastatic cancer cells, and breast cancer cells tend to infiltrate the bone, brain, liver and lungs [2,3,4,5]. The mechanisms that promote the adhesion of breast cancer cells to organ-specific endothelial layers remain unclear. Better understanding of the specific molecular mechanisms used by cancer cells to activate dynamic interactions with endothelial cells is instrumental to identify promising novel molecular targets for anti-metastatic therapy

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