Towards better antivenoms: navigating the road to new types of snakebite envenoming therapies

Abstract Snakebite envenoming is a significant global health challenge, and for over a century, traditional plasma-derived antivenoms from hyperimmunized animals have been the primary treatment against this infliction. However, these antivenoms have several inherent limitations, including the risk of causing adverse reactions when administered to patients, batch-to-batch variation, and high production costs. To address these issues and improve treatment outcomes, the development of new types of antivenoms is crucial. During this development, key aspects such as improved clinical efficacy, enhanced safety profiles, and greater affordability should be in focus. To achieve these goals, modern biotechnological methods can be applied to the discovery and development of therapeutic agents that can neutralize medically important toxins from multiple snake species. This review highlights some of these agents, including monoclonal antibodies, nanobodies, and selected small molecules, that can achieve broad toxin neutralization, have favorable safety profiles, and can be produced on a large scale with standardized manufacturing processes. Considering the inherent strengths and limitations related to the pharmacokinetics of these different agents, a combination of them might be beneficial in the development of new types of antivenom products with improved therapeutic properties. While the implementation of new therapies requires time, it is foreseeable that the application of biotechnological advancements represents a promising trajectory toward the development of improved therapies for snakebite envenoming. As research and development continue to advance, these new products could emerge as the mainstay treatment in the future.

Determination of low tetanus or diphtheria antitoxin titers in sera by a toxin neutralization assay and a modified toxin-binding inhibition test

A method for the screening of tetanus and diphtheria antibodies in serum using anatoxin (inactivated toxin) instead of toxin was developed as an alternative to the in vivo toxin neutralization assay based on the toxin-binding inhibition test (TOBI test). In this study, the serum titers (values between 1.0 and 19.5 IU) measured by a modified TOBI test (Modi-TOBI test) and toxin neutralization assays were correlated (P < 0.0001). Titers of tetanus or diphtheria antibodies were evaluated in serum samples from guinea pigs immunized with tetanus toxoid, diphtheria-tetanus or triple vaccine. For the Modi-TOBI test, after blocking the microtiter plates, standard tetanus or diphtheria antitoxin and different concentrations of guinea pig sera were incubated with the respective anatoxin. Twelve hours later, these samples were transferred to a plate previously coated with tetanus or diphtheria antitoxin to bind the remaining anatoxin. The anatoxin was then detected using a peroxidase-labeled tetanus or diphtheria antitoxin. Serum titers were calculated using a linear regression plot of the results for the corresponding standard antitoxin. For the toxin neutralization assay, L+/10/50 doses of either toxin combined with different concentrations of serum samples were inoculated into mice for anti-tetanus detection, or in guinea pigs for anti-diphtheria detection. Both assays were suitable for determining wide ranges of antitoxin levels. The linear regression plots showed high correlation coefficients for tetanus (r² = 0.95, P < 0.0001) and for diphtheria (r² = 0.93, P < 0.0001) between the in vitro and the in vivo assays. The standardized method is appropriate for evaluating titers of neutralizing antibodies, thus permitting the in vitro control of serum antitoxin levels.