Identification of potent nanobodies to neutralize the most poisonous polypeptide from scorpion venom

Abstract

Scorpion venom, containing highly toxic, small polypeptides that diffuse rapidly within the patient, causes serious medical problems. Nanobodies, single-domain antigen-binding fragments derived from dromedary heavy-chain antibodies, have a size that closely matches that of scorpion toxins. Therefore these nanobodies might be developed into potent immunotherapeutics to treat scorpion envenoming. Multiple nanobodies of sub-nanomolar affinity to AahII, the most toxic polypeptide within the Androctonus australis hector venom, were isolated from a dromedary immunized with AahII. These nanobodies neutralize the lethal effect of AahII to various extents without clear correlation with the kinetic rate constants kon or koff, or the equilibrium dissociation constant, KD. One particular nanobody, referred to as NbAahII10, which targets a unique epitope on AahII, neutralizes 7 LD50 of this toxin in mice, corresponding to a neutralizing capacity of approx. 37000 LD50 of AahII/mg of nanobody. Such high neutralizing potency has never been reached before by any other monoclonal antibody fragment.

Exploiting Cross-reactivity to Neutralize Two Different Scorpion Venoms with One Single Chain Antibody Fragment

“…Another antibody source has been the dromedary immune system. Recent papers showed the ability of antibody fragments, known as nanobodies, to neutralize the most abundant toxins of the scorpion Androctonus australis Hector (39,40). The whole venom has been neutralized by a bi-specific antibody fragment (41).…”

Effectiveness of the Androctonus Australis Hector Nanobody Nbf12-10 Antivenom to Neutralize Significantly the Toxic Effect and Tissue Damage Provoked by Fraction of Androctonus mauretanicus (Morocco) Scorpion Venom

“…This fraction corresponds to most abundant toxic component, obtained after analytical filtration of Am venom (Figure 1). F3 fraction showed a similar retention time as that of AahG50 (derived from Aah venom) fraction whereas the eluted fractions on either side of this F3 peak display significant divergence in retention time [34,36]. The LD 50 of F3 fraction, estimated as the amount of toxins whereby 50% of the mice (cohorts of 4 mice) survived the injection that corresponds to Results of mice groups challenged with molar ratios of F3: NbF12-10 by i.c.v injection.…”

“…The LD 50 represents as amount of venom or toxic fraction leading to 50 % of mice surviving the injection [34,36]. Survivors were counted after 24h.…”

“…Interestingly, the Nbs share a high sequence identity with human VH of family III [32,33]. From VHH library screenings, several Nbs have been retrieved using phage display technology, from which we further selected NbAahI'F12 and NbAahII10 against AahI' and AahII toxins, respectively [34][35][36]. The corresponding bispecific NbF12-10 format of only 29 kDa yielded a therapeutic compound with a strong protective capacity against the whole Aah venom in mice [35].…”

Multiple interests in structural models of DARC transmembrane protein

“…VHHs are easily cloned from lymphocytes from naive or immunized camelids; they can be expressed in E. coli with a good yield and have an excellent solubility [143]. Moreover, they have proved to be efficient as therapeutic and diagnostic agents [144,145].…”