Assessment of the Inhibitory Effects of Ficin-hydrolyzed Gelatin Derived from Squid (Uroteuthis duvauceli) on Breast Cancer Cell Lines and Animal Model.

Marine novel natural products have been applied for cancer therapy. Enzyme-digested gelatin hydrolysates have proven to serve as promising sources of potent biologically active peptides. Potential anti-breast cancer properties of the extracted Ficin-digesterd gelatin hydrolysate from Indian squid (Uroteuthis duvauceli) was extensively characterized by cellular and animal models. Gelatin was extracted from squid skin, hydrolyzed by Ficin, and characterized by standard physico-chemical methods. Ficin-digested gelatin hydrolysate was used at various doses of 0-0.1 mg/mL for assessment of MCF-7 and MDA-MB-231 breast cancer cells versus HUVEC normal cells. Cytotoxicity, phase-contrast morphological examination, apoptosis/necrosis, clonal-growth, cell-migration, Matrix-metalloproteinases (MMPs) zymography, and Western blotting were used for cellular assessments. For animal studies, breast tumor-induced BALB/c mice received hydrolyzed gelatin regimen, followed by tumor size/growth and immune-histochemical analyses. Significant inhibition of MCF-7 and MDA-MB-231 with no cytotoxicity on HUVEC cells were detected. Apoptosis was increased in cancer cells, as revealed by elevated ratio of cleaved caspase-3 and PARP. MMP-2 and MMP-9 activities in both cancer cells were diminished. In mice, gelatin hydrolysate prevented weight loss, decreased tumor size, induced p53, and down-regulated Ki67 levels. These findings suggest that Ficin-digested gelatin hydrolysate could be a beneficial candidate for novel breast cancer therapies.

A New, High Yield, Rapid, and Cost-Effective Protocol to Deprotection of Cysteine-Rich Conopeptide, Omega-Conotoxin MVIIA.

Almost all conopeptides purified from Conus venoms are cysteine-rich peptides. Among them, omega-conotoxin MVIIA, FDA approved peptide drug (Prialt(®)), selected as a cysteine-rich model that its protection from oxidation is critical during solid phase synthesis. Deprotection of cysteines is a crucial step after peptide synthesis. The current study aimed to set up a new highly efficient deprotection protocol for omega-conotoxin MVIIA. Deprotection was performed based on mercury acetate with significant major modification. The protocol accomplished based on the best molar ratio of peptide/mercury/2-ME that adjusted to 0.2 mm/3 mm/10 mm (50 µg/1 mg/10 µL). The yield and purity of omega-conotoxin MVIIA obtained at 93 and 95%, respectively. The total time of protocol shortened to 90 min instead of 6-20 h in routine methods. In this study, omega-conotoxin MVIIA was recovered in high yield and in the shortest time. Despite of other known protocols, molar ratio adjusted to minimum amount. In conclusion, this protocol would be suggested to cost-effective deprotection of thiol groups for similar cysteine-rich peptides.