Impairment of cell adhesion and migration by inhibition of protein disulphide isomerases in three breast cancer cell lines

Abstract

Protein disulphide isomerase A3 (PDIA3) is an ER-resident disulphide isomerase and oxidoreductase with known substrates that include some extracellular matrix proteins. PDIA3 is upregulated in invasive breast cancers and correlates in a mouse orthotopic xenograft model with breast cancer metastasis to bone. However, the underlying cellular mechanisms remain unclear. Here we investigated the function of protein disulphide isomerases in attachment, spreading and migration of three human breast cancer lines representative of luminal (MCF-7) or basal (MDA-MB-231 and HCC1937) tumour phenotypes. Pharmacological inhibition by 16F16 in cells decreased initial cell spreading more effectively than inhibition by PACMA-31. Cells displayed diminished cortical F-actin projections, stress fibres and focal adhesions. Cell migration was reduced in a quantified “scratch wound” assay. To examine whether these effects might result from alterations to secreted proteins in the absence of functional PDIA3, adhesion and migration were quantified in the above cells exposed to media conditioned by wild-type or Pdia3-/- mouse embryo fibroblasts. The conditioned medium of Pdia3-/- fibroblasts was less effective in promoting cell spreading and F-actin organisation or supporting “scratch wound” closure. Similarly, extracellular matrix prepared from HCC1937 cells after 16F16 inhibition was less effective than control ECM to support spreading of untreated HCC1937 cells. Overall, these results advance the concept that protein disulphide isomerases including PDIA3 drive the production of secreted proteins that promote a microenvironment favourable to breast cancer cell adhesion and motility, characteristics that are integral to tumour invasion and metastasis. Inhibition of PDIA3 or related isomerases may have potential for anti-metastatic therapies.

PDIA3/ERp57 promotes a matrix-rich secretome that stimulates fibroblast adhesion through CCN2

“…We have validated certain commercial antibodies for specificity against recombinant PDIA3 protein versus recombinant PDI/PDIA1 ( 37 ). An additional anti-PDIA3 raised in mouse was validated here ( Fig.…”

Role of Differentially Expressed Proteins in Acquired Resistance to Cdk4/6 Inhibitor in Breast Cancer

“…Chemo-resistance is caused by the tumour microenvironment facilitating CD9-mediated crosstalk between mesenchymal stem cells and BC cells via CCL5, CCR5, and CXCR12 [39]. PDIA3 expression in the tumor microenvironment is high, which favors invasion and metastasis [40,41].GLUT1 proteins encoded by SLC2A1 aid glucose transport. Hypoxia inducing factor-1(HIF-1) is activated as breast cancer grows aggressively.…”

Roles of Protein Disulfide Isomerase in Breast Cancer

“…Its regulation of breast cancer cell migration and adhesion is executed by inducing the phosphorylation of tyrosine kinases such as Src as proved by the experiment that treatment with Dasatinib, a protein kinase inhibitor, remarkably reduces AGR3-dependent migration. In addition, PDIA3 promotes pro-migratory phenotypes in either luminal (MCF-7) or basal breast tumor subtypes (MDA-MB-231 and HCC1937), and PDI inhibition led by 16F16 efficiently decreases initial cell spreading [94]. Similar to 16F16, another PDI inhibitor, PACMA31, could block the transendothelial migration of MDA-MB-231 cells and the contraction of collagen that affect the exposition of free thiols on integrin molecules [86].…”

“…16F16 reduces the initial rates of closure and overall scratch closure for all cell lines. Also, a reduction in pro-migratory F-actin structures, including lamellipodia, is observed in the three cell lines in treatment with 16F16 [94]. Their different phenotypic responses between PACMA31 and 16F16 in the breast cancer cells are potentially due to the difference in their primary target in PDIs.…”

“…16F16 is an irreversible PDI inhibitor that impairs cell adhesion and the migration of breast cancer cells MCF-7, MDA-MB-231, and HCC1937 [94]. The spreading and attachment of breast cancer cells are strongly reduced by 16F16 treatment, and the effect of 16F16 is stronger than PACMA31.…”