Third Generation Antivenomics: Pushing the Limits of the In Vitro Preclinical Assessment of Antivenoms

Abstract

Second generation antivenomics is a translational venomics approach designed to complement in vivo preclinical neutralization assays. It provides qualitative and quantitative information on the set of homologous and heterologous venom proteins presenting antivenom-recognized epitopes and those exhibiting impaired immunoreactivity. In a situation of worrying antivenom shortage in many tropical and sub-tropical regions with high snakebite mortality and morbidity rates, such knowledge has the potential to facilitate the optimal deployment of currently existing antivenoms and to aid in the rational design of novel broad specificity antidotes. The aim of the present work was to expand the analytical capability of the immunoaffinity second-generation antivenomics platform, endowing it with the ability to determine the maximal binding capacity of an antivenom toward the different toxins present in a venom, and to quantify the fraction of venom-specific antibodies present in a given antivenom. The application of this new platform, termed third generation (3G) antivenomics, in the preclinical evaluation of antivenoms is illustrated in this paper for the case of antivenom EchiTAb-Plus-ICP® reactivity towards the toxins of homologous (B. arietans) and heterologous (N. melanoleuca) venoms.

Vipera berus berus Venom from Russia: Venomics, Bioactivities and Preclinical Assessment of Microgen Antivenom

“…Antivenomics is a translational venomics approach designed to complement in vivo preclinical neutralization assays. It provides qualitative and quantitative information on the binding capacity of an antivenom toward the different toxins present in a venom and on the fraction of venom-specific antibodies present in a given antivenom [36]. To achieve locus resolution in antivenomics analysis, a sine qua non condition is to know the composition of the venom proteome [37].…”

“…The immunological reactivity of the therapeutic antivenom manufactured by “Microgen” LTD, Russia, was assessed by third-generation immunoaffinity-based antivenomics for the treatment of V. b. berus envenoming [36], as described for other antivenoms [45,46]. Figure 2 displays the series of antivenomics experiments in which a fixed amount (10 mg) of antivenom was confronted with the increasing amounts of V. b. berus venom.…”

“…There is a lack of information on the preclinical efficacy of another antivenom, Anti-Viper Venom Serum, manufactured by the Federal State Company for Immunobiological Medicines, Microgen (Moscow, Russia), despite it being registered in the Russian Federation for V. berus envenoming. This study was designed to assess the preclinical efficacy of the Russian Anti-Viper Venom antivenom to neutralize three key effects of V. b. berus venom, i.e., lethal, hemorrhagic and phospholipase A 2 activity, by combination of in vivo neutralization assays [35] and in vitro third-generation antivenomics analysis [36,37]. This will ultimately help in assessing the toxin recognition landscape of the antivenom and quantify the fraction of therapeutic antivenom molecules.…”

An analysis of preclinical efficacy testing of antivenoms for sub-Saharan Africa: Inadequate independent scrutiny and poor-quality reporting are barriers to improving snakebite treatment and management

“…For this reason, in recent years, a number of groups have moved towards the use of "rescue" experiments, where venom is often delivered by different routes and treatment is administered after the onset of envenomation [66][67][68][69]. However, it is clear that to use preclinical data to more rationally and precisely model the therapeutic doses required to effect cure, a more comprehensive understanding of the following is needed; i) the relative amount of toxin-neutralising antibodies in antivenom products, such as provided by antivenomic analyses [16], ii) the amount and type of venom toxins circulating in snakebite victims over the time course of an envenoming and in different tissue compartments and iii) the concentrations of antivenom required to bind and inhibit the pathogenicity of venom toxins in that time frame and tissue compartment. Such pharmacodynamic data is routine for the majority of licensed therapeutics that have successfully progressed through normal clinical-trial processes [70].…”

“…The preliminary in vitro approach (e.g. using methods such as ELISA, immunoblotting [ 15 ] or antivenomics [ 16 ]) is primarily to identify antivenoms which are likely to lack efficacy during essential in vivo assays, and are thus crucial to reducing the number of animals required for testing.…”