Elsevier

Biochemical Pharmacology

Volume 60, Issue 2, 15 July 2000, Pages 269-274
Biochemical Pharmacology

Inflammation and immunopharmacology
Effectiveness of batimastat, a synthetic inhibitor of matrix metalloproteinases, in neutralizing local tissue damage induced by BaP1, a hemorrhagic metalloproteinase from the venom of the snake Bothrops asper

https://doi.org/10.1016/S0006-2952(00)00302-6Get rights and content

Abstract

Batimastat (BB-94), a synthetic hydroxamate peptidomimetic matrix metalloproteinase inhibitor, was tested for its ability to inhibit proteolytic and toxic effects induced by BaP1, a 24-kDa hemorrhagic metalloproteinase isolated from the venom of Bothrops asper, the medically most important snake species in Central America and southern Mexico. Batimastat inhibited proteolytic activity on biotinylated casein, with anic50 of 80 nM. In addition, batimastat was effective in inhibiting hemorrhagic, dermonecrotic, and edema-forming activities of this metalloproteinase if incubated with the enzyme prior to the assays. When the inhibitor was administered i.m. at the site of the toxin injection without preincubation, rapidly after metalloproteinase administration, it totally abrogated the hemorrhagic and dermonecrotic effects of BaP1. Inhibition was less effective as the time lapse between toxin and batimastat injection increased, due to the extremely rapid development of BaP1-induced local tissue damage in this experimental model. On the other hand, batimastat was ineffective if administered by the i.p. route immediately after toxin injection. It is concluded that batimastat, and probably other synthetic metalloproteinase inhibitors, may become useful therapeutic tools aimed at the in situ inhibition of venom metalloproteinases, when injected at the site of the bite rapidly after envenomation.

Section snippets

Metalloproteinase and inhibitor

BaP1 was isolated from a venom pool of more than 40 adult specimens of B. asper collected in the Pacific region of Costa Rica. Purification was performed by ion-exchange chromatography on Carboxymethyl Sephadex C-50, gel filtration in Sephacryl S-200, and affinity chromatography on Affi-Gel Blue, as previously described 15, 20. Homogeneity was demonstrated by SDS–PAGE run under reducing conditions [26], using 12% acrylamide gels. Batimastat (also known as BB-94; [4-(N-hydroxyamino)-2R-isobutyl-3

Assays with preincubation of BaP1 and batimastat

Batimastat inhibited, in a concentration-dependent way, the proteolytic activity of BaP1 on biotinylated casein, with an ic50 of 80 nM (Fig. 1). A concentration of 400 nM batimastat totally inhibited caseinolytic activity. Moreover, batimastat was effective in inhibiting the hemorrhagic activity of BaP1 in skin and muscle (Fig. 2). When hemorrhage inhibition was assessed in the skin, the ic50 was 1.95 μM. Batimastat concentrations of 5.71 and 12.6 μM abrogated hemorrhagic activity totally in

Discussion

Venom metalloproteinases play a relevant role in the pathogenesis of local tissue damage characteristic of pit viper envenomations 3, 21. In the case of B. asper venom, metalloproteinases induce hemorrhage 17, 27, myonecrosis 18, 19, dermonecrosis, blister formation [15], and edema [20]. Neutralization of these activities by antivenoms is a difficult task, due to the time lapse between envenomation and antivenom administration in hospitals [9]. The use of synthetic metalloproteinase inhibitors

Acknowledgements

The authors thank British Biotech Pharmaceuticals Ltd. for supplying batimastat, and Javier Núñez, Marcos Méndez, Rodrigo Mora, and Rodrigo Chaves for their collaboration in the laboratory work. This study was supported by the International Foundation for Science (project F/2707–1), by the Consejo Nacional para Investigaciones Cientı́ficas y Tecnológicas (CONICIT) of Costa Rica (project 98–012-F0), and by Vicerrectorı́a de Investigación, Universidad de Costa Rica (project 741–98-202).

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