Protective immunity against alpha-cobratoxin following a single administration of a genetic vaccine encoding a non-toxic cobratoxin variant.

Venomous snakebites result in almost 125,000 deaths per year worldwide. We present a new paradigm for the development of vaccines to protect against snakebite, using knowledge of the structure and action of specific toxins combined with a gene-based strategy to deliver a toxin gene modified to render it non-toxic while maintaining its three-dimensional structure and hence its ability to function as an immunogen. As a model for this approach, we developed a genetic vaccine to protect against alpha-cobratoxin (CTX), a potent, post-synaptic neurotoxin that is the major toxic component of the venom of Naja kaouthia, the monocellate cobra. To develop the vaccine, substitutions in the CTX cDNA were introduced at two residues critical for binding to the nicotinic acetylcholine receptor (Asp27 to Arg, Arg33 to Gly). The mutated CTX expression cassette was delivered in the context of a replication deficient adenovirus vector (AdmCTX). To assess whether expression of the mutated CTX in vivo leads to the development of protective immunity, BALB/c mice were challenged by IV administration of 2 microg of alpha-cobratoxin protein 21 or 63 days after administration of AdmCTX or Ad- Null (as a control; both, 10(9) particle units). Animals receiving AdmCTX but no alpha-cobratoxin challenge suffered no ill effects, but > or =80% of naive animals or those receiving the AdNull control vector died within 10 min from the alpha-cobratoxin challenge. In contrast, 100% of animals receiving a single dose of AdmCTX 21 or 63 days prior to alpha-cobratoxin challenge survived. The data demonstrates that an adenovirus-based vaccine can be developed to protect against lethal challenge with a potent snake venom. The effectiveness of this approach might serve as a basis to consider the development of a global public health program to protect those at risk for death by snakebite.