Advancing Antivenom

Nanobody-Based Recombinant Antivenom Shows Promise for African Snakebites

New research published in Nature suggests that an experimental antivenom derived from nanobodies—small antibody fragments —could offer broad protection against venom from 17 medically important elapid snakes in sub-Saharan Africa. Results indicated that this recombinant approach neutralized lethality in preclinical models and reduced tissue damage, performing better than a conventional plasma-derived antivenom in tested scenarios.

The Burden of Snakebite Envenoming

Snakebite envenoming is an important public health consideration in sub-Saharan Africa, particularly for rural communities. Many envenomings occur each year, affecting health and well-being. Traditional antivenoms, derived from the plasma of immunized animals such as horses, have been effective in saving lives, though they come with certain challenges, including potential adverse reactions, variability in batch quality, and varying degrees of protection against local tissue effects or different snake species.

The study focuses on elapids, a family of venomous snakes including cobras, mambas, and the rinkhals, whose bites cause symptoms ranging from paralysis to severe tissue necrosis. "There is thus an urgent unmet medical need for innovation in snakebite envenoming therapy," the researchers stated, emphasizing the complexity of venoms that vary widely between species.

An Innovative Use of Nanobodies

To address these challenges, scientists turned to nanobodies, single-domain antibody fragments derived from heavy-chain-only antibodies found in camelids such as alpacas and llamas. These nanobodies are about one-tenth the size of conventional antibodies—like comparing a grain of sand to a pebble—potentially allowing for better tissue penetration, stability, and recombinant production in microbes for consistency and lower cost.

The approach leverages phage display technology, where nanobodies are displayed on bacteriophages (viruses that infect bacteria) to screen for those that bind strongly to toxins. This method has shown promise in other fields, but applying it to create a polyvalent antivenom across multiple snake genera represents a notable advance.

Methodology: From Immunization to Antivenom Design

The process involved immunizing an alpaca and a llama with a mixture of venoms from 18 elapid species over several months, followed by constructing three unique phage display libraries from their blood samples. Key toxins were isolated using reversed-phase high-performance liquid chromatography (RP-HPLC) and proteomics, targeting families like three-finger toxins (3FTx, which include neurotoxins and cytotoxins) and phospholipases A2 (PLA2, enzymes that break down cell membranes).

Screening yielded over 100 unique nanobodies, with 15 top candidates evaluated for binding affinity and cross-reactivity using assays like DELFIA and BLI. In vitro neutralization tests followed: patch clamp experiments measured protection against neurotoxins disrupting nerve-muscle communication, while cell viability and enzymatic assays assessed mitigation of cytotoxicity and PLA2 activity.

Eight nanobodies were ultimately combined into an oligoclonal mixture, designed to cover seven toxin families. Structural analyses, including cryo-electron microscopy and modeling, revealed that these nanobodies bind to conserved regions on toxins, explaining their broad neutralizing potential.

Key Findings: Neutralization In Vitro and In Vivo

In vitro results showed that selected nanobodies fully neutralized neurotoxins at equimolar ratios and provided 50-95% protection against cytotoxicity from seven spitting cobra venoms. One nanobody completely inhibited PLA2 enzymatic activity in those venoms.

Preclinical mouse experiments demonstrated encouraging results. The nanobody cocktail prevented lethality from 17 of the 18 venoms in pre-incubation tests and in a rescue model simulating real-world bites for 11 venoms. It also markedly reduced dermonecrosis—skin and tissue destruction—for all tested cytotoxic venoms. Compared to a plasma-derived antivenom, the recombinant version offered superior performance against lethality and morbidity.

Underscoring its potential for continent-wide coverage:

"This antivenom effectively prevented venom-induced lethality in vivo across 17 African elapid snake species," - Ahmadi et al.

Implications and Future Directions

These findings suggest that recombinant nanobody-based antivenoms could improve access to effective treatments in resource-limited settings, potentially reducing the reliance on animal-derived products. By targeting conserved toxin elements, the approach might extend to other regions with similar snake species.

"Collectively, this study demonstrates the feasibility of developing a polyvalent recombinant antivenom," - Ahmadi et al.