Development of a neutralizing monoclonal antibody targeting Bothrops atrox venom metalloproteinases

Snakebites are classified as a neglected tropical disease by the World Health Organization. In South America's tropical rainforests, Bothrops genus, particularly Bothrops atrox, is responsible for most incidents. Severe local effects, such as hemorrhage, are primarily caused by snake venom metalloproteinases (SVMPs), which are not fully neutralized by conventional therapy. Here, we report the production of a neutralizing monoclonal antibody (mAb) against the hemorrhagic activity of B. atrox venom metalloproteinases. mAbs were produced by immunization of BALB/c mice using the B. atrox venom (BaV). The resulting hybridomas were screened by ELISA using BaV as antigen. The selected clone 4H4D11 (mAb-BaSVMP) showed cross-reactivity with other medically important species of Bothrops snakes in Brazil and Peru. Western blot assays revealed that the produced mAb binds to proteins with molecular masses of approximately 50 kDa and 20 kDa in BaV and recognizes native Atroxlysin-III and Atroxlysin-I by ELISA. mAb-BaSVMP did not bind to a cellulose membrane containing the primary sequence of a metalloproteinase, suggesting that it may recognize a conformational epitope. Additionally, mAb-BaSVMP neutralizes the in vivo hemorrhagic activity caused by BaV in mice. These results highlight the potential usefulness of mAb-BaSVMP for developing effective antivenoms for passive immunotherapy against bothropic envenomation.