Leucurogin, a new recombinant disintegrin cloned from Bothrops leucurus (white-tailed-jararaca) with potent activity upon platelet aggregation and tumor growth

Abstract

Disintegrins and disintegrins-like proteins are able to inhibit platelet aggregation and integrin-mediated cell adhesion. The aim of this study was to produce one disintegrin-like cloned from Bothrops leucurus venom gland and to characterize it regarding biological activity. The recombinant protein was purified by one step procedure involving anion-exchange chromatography (DEAE-cellulose) and presented a molecular mass of 10.4 kDa. The purified protein was able to inhibit platelet aggregation induced by collagen (IC₅₀ = 0.65 μM) and to inhibit growth of Ehrlich tumor implanted in mice by more than 50% after 7 days administration of 10 μg/day. No effects were observed upon adenosine 5'-diphosphate (ADP)-and arachidonic acid (AA)-induced platelet aggregation. The recombinant protein was recognized by an antibody specific for jararhagin one metalloproteinase isolated from Bothrops jararaca venom, and therefore it was named leucurogin. Anti-angiogenesis effect of leucurogin was evaluated by the sponge implant model. After 7 days administration leucurogin inhibited, in a dose dependent way, the vascularization process in the sponge. Leucurogin represents a new biotechnological tool to understand biological processes where disintegrins-like are involved and may help to characterize integrins that can be involved in development and progression of malignant cells.

Anti-angiogenic activities of two recombinant disintegrins derived from the Mohave and Prairie rattlesnakes

“…Vicrostatin (VCN), a chimeric recombinant disintegrin generated as a genetic fusion between the C-terminal tail of a viperid disintegrin (echistatin) and crotalid disintegrin (contortrostatin), exerts a potent inhibitory effect on EC migration and tube formation in a dose-dependent manner by forcing these cells to undergo significant actin cytoskeleton reorganization and by inducing apoptosis of EC (Minea et al, 2012, 2010). Leucorogin, is another new recombinant disintegrin cloned from Bothrops leucurus , that inhibited in a dose-dependent manner, the vascularization process evaluated by the sponge implant model (Higuchi et al, 2011). Acurhagin-C, an ECD disintegrin from Agkistrodon acutus venom, inhibited EC adhesion and spreading on extracellular matrix proteins, followed by damage to cell functions and finally eliciting anoikis (Wang, 2010).…”

“…Hemoglobin was quantified by a colorimetric method as described by Higuchi et al (2011). Matrigel plug was homogenized in 2 mL Drabkin reagent (Sigma–Aldrich, MO, USA) and centrifuged at 10,000 × g for 15 min.…”

Anti-invasive and anti-adhesive activities of a recombinant disintegrin, r-viridistatin 2, derived from the Prairie rattlesnake (Crotalus viridis viridis)

“…Therefore the characterization of integrins antagonists is an interesting subject of study and disintegrins appear as putative candidates to be used as effective tools for cancer therapy (Higuchi et al, 2011). The expression of recombinant versions of interesting disintegrins has become essential, thus facilitating the maintenance of a continuous supply for drug development (Sánchez et al, 2010).…”

The role of platelets in hemostasis and the effects of snake venom toxins on platelet function

“…However, the specific integrin-binding motif was not determined (Marcinkiewicz, 2013). In turn, leucurogin, a recombinant disintegrin-like cloned from B. leucurus, is able to inhibit platelet aggregation induced by collagen, probably through interactions with integrins α 1 β 1 or α 2 β 1 (Higuchi et al, 2011).…”