Abstract
The aim of our study was to determine whether the use of cisplatin in the presence echistatin in MDA-MB-231 breast cancer cells leads to a reduction of toxic effects associated with the use of cisplatin. The expression of β1-integrin and insulin-like growth factor 1 receptor (IGF-IR), signaling pathway protein expression: protein kinase B (AKT), mitogen-activated protein kinases (ERK1/ERK2), nuclear factor kappa B (NFκB), and caspase-3 and -9 activity was measured after 24 h of incubation with tested compounds to explain detailed molecular mechanism of induction of apoptosis. The viability of MDA-MB-231 breast cancer cells was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Annexin V-FITC/propidium iodide staining assay was performed to detect the induction of apoptosis. Inhibition DNA biosynthesis was determined by [3H]thymidine incorporation into DNA. The expression of of β1-integrin, IGF-IR, AKT, ERK1/ERK2, NFκB, caspase-3 and -9 was evaluated using Western blot. The results suggest that treatment of MDA-MB-231 breast cancer cells for 24 h cisplatin plus echistatin severely inhibits cell growth and activates apoptosis by upregulation of caspase-3 and -9 expressions. The effect was stronger than treatment cisplatin and echistatin alone. In this study, we have found that cisplatin plus echistatin treatment decreases collagen biosynthesis in MDA-MB-231 breast cancer cells stronger than the individual compounds. The inhibition was found to be dependent on the β1-integrin and IGF receptor activation. A significant reduction of ERK1/ERK2, AKT expression in cancer cells after cisplatin plus echistatin treatment was also found. The cancer cells treated by echistatin, cisplatin, and in particular the combination of both compounds drastically increased expression of NFκB transcription factor. Our results suggest that combined therapy cisplatin plus echistatin is a possible way to improve selectiveness of cisplatin. This mechanism probably is due to downregulation of expression of β1-integrin and IGF-IR receptors, and the signaling pathway proteins induced by these receptors. Our results suggest that therapy cisplatin plus echistatin is a possible way to improve selectiveness of cisplatin.
Highlights
The aim of our study was to determine whether the use of cisplatin in the presence echistatin in MDA-MB-231 breast cancer cells leads to a reduction of toxic effects associated with the use of cisplatin
The results suggest that treatment of MDAMB-231 breast cancer cells for 24 h cisplatin plus echistatin severely inhibits cell growth and activates apoptosis by upregulation of caspase-3 and -9 expressions
Our results suggest that therapy cisplatin plus echistatin is a possible way to improve selectiveness of cisplatin
Summary
The aim of our study was to determine whether the use of cisplatin in the presence echistatin in MDA-MB-231 breast cancer cells leads to a reduction of toxic effects associated with the use of cisplatin. The results suggest that treatment of MDAMB-231 breast cancer cells for 24 h cisplatin plus echistatin severely inhibits cell growth and activates apoptosis by upregulation of caspase-3 and -9 expressions. It is known that echistatin as an inhibitor of b1-integrin receptor contributes to inhibition of collagen biosynthesis and decrease in the expression of FAK (focal adhesion kinase), SOS-protein (son of sevenless protein), and phosphorylated MAP-kinases (mitogen-activated protein kinases), and ERK1 (extracellular signal-regulated kinase 1) and ERK2 (extracellular signal-regulated kinase 2) [18]. Stimulated b1-integrin receptor induces autophosphorylation of nonreceptor protein kinase FAK, which is capable of interacting with adaptor proteins, such as Grb (growth factor receptor bound protein 2), through Src and Shc proteins This interaction allows activating further cascade of signaling pathway through SOS, Ras, and Raf proteins and subsequently, two MAP-kinases: ERK1 and ERK2. The end point of this cascade is induction of transcription factor(s) that regulate(s) gene expression of integrins, proteinases, and many proteins involved in the regulation of cell growth and differentiation [22,23,24]
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