A Bowman–Birk inhibitor induces apoptosis in human breast adenocarcinoma through mitochondrial impairment and oxidative damage following proteasome 20S inhibition

Abstract

Proteasome inhibitors are emerging as a new class of chemopreventive agents and have gained huge importance as potential pharmacological tools in breast cancer treatment. Improved understanding of the role played by proteases and their specific inhibitors in humans offers novel and challenging opportunities for preventive and therapeutic intervention. In this study, we demonstrated that the Bowman–Birk protease inhibitor from Vigna unguiculata seeds, named black-eyed pea trypsin/chymotrypsin Inhibitor (BTCI), potently suppresses human breast adenocarcinoma cell viability by inhibiting the activity of proteasome 20S. BTCI induced a negative growth effect against a panel of breast cancer cells, with a concomitant cytostatic effect at the G2/M phase of the cell cycle and an increase in apoptosis, as observed by an augmented number of cells at the sub-G1 phase and annexin V-fluorescin isothiocyanate (FITC)/propidium iodide (PI) staining. In contrast, BTCI exhibited no cytotoxic effect on normal mammary epithelial cells. Moreover, the increased levels of intracellular reactive oxygen species (ROS) and changes in the mitochondrial membrane potential in cells treated with BTCI indicated mitochondrial damage as a crucial cellular event responsible for the apoptotic process. The higher activity of caspase in tumoral cells treated with BTCI in comparison with untreated cells suggests that BTCI induces apoptosis in a caspase-dependent manner. BTCI affected NF-kB target gene expression in both non invasive and invasive breast cancer cell lines, with the effect highly pronounced in the invasive cells. An increased expression of interleukin-8 (IL-8) in both cell lines was also observed. Taken together, these results suggest that BTCI promotes apoptosis through ROS-induced mitochondrial damage following proteasome inhibition. These findings highlight the pharmacological potential and benefit of BTCI in breast cancer treatment.

Plant Protease Inhibitors in Therapeutics-Focus on Cancer Therapy

“…Recently, Souza et al (2014) reported the effects of a BTCI on proteasome 20S in MCF-7 breast cancer cells and the catalytic activity of the purified 20S proteasome from horse erythrocytes, as well as the structural analysis of the BTCI-20S proteasome complex. Furthermore, Mehdad et al (2016) stated that BTCI significantly decreased human breast adenocarcinoma cell viability by inhibiting the activity of proteasome 20S, with an associated cytostatic effect at the G2/M phase of the cell cycle and an increase in apoptosis.…”

Oligomerization affects the kinetics and thermodynamics of the interaction of a Bowman-Birk inhibitor with proteases

“…It presents rich biotechnological potential as an anticarcinogenic agent, as a guanylin-enhancing natriuresis and renal functional response in rats, as a bradykinin-potentiating molecule, and as an insecticide, inhibiting the growth of larvae with agricultural importance [7][8][9][10][11][12].…”

Chemoprevention by means of soy proteins and peptides – current status and future approaches: a review

“…Moreover, BBI inhibits AGS gastric adenocarcinoma proliferation through the inhibition of VEGF, MMP‐2 and MMP‐9 production (Fereidunian et al ., ). Seed BBI induces apoptosis through intracellular reactive oxygen species‐induced mitochondrial damage following suppression of proteasome 20S activity in breast cancer cells (Mehdad et al ., ). In animal models, BBI also inhibits the growth of colon cancer in rats, prevents chemical‐carcinogen‐induced lung cancer in A/J mice, and prevents prostate neoplasms in rats and TRAP transgenic rats (Hernández‐Ledesma & Hsieh, ).…”