DEVELOPMENT OF A NEW GENERATION OF ANTIVENOMIC APPROACH BY MASS SPECTROMETRY

Introduction
Snakebite causes the death of about 150,000 people/year. Envenomations are classically treated by injecting anti-venomous sera. However, these treatments can induce immunological reactions which can present several adverse effects to the patient. Venom compositions strongly differ from species to species, gender and habitat and providing antivenoms targeting a specific venom is then already a challenge. Quantitatively evaluating the efficacy of any antivenom is primordial to improve the production of effective sera and to determine the nature of the toxins indeed bound by the antibodies (Igs). Therefore, mass spectrometry (MS) is important to performed it, ant it is then called ¿antivenomics¿. We plan to exploit the huge potential of magnetic beads and of mass spectrometry to speed up the antivenoms efficacy.
Methods
Shotgun proteomics was performed with 10¿g Echis ocellatus and Dendroaspis polylepis venoms, which were reduced/alkylated/digested with a mixture of Trypsin/GluC/Chymotrypsin, then analysed using a Q-Exactive¿ Plus Mass Spectrometer and the protein identification performed by PeaksStudio X+ using Uniprot/transcriptomes databases. Next step consists in grafting the antivenom antibodies to magnetic beads and incubating with the crude venoms. Comparative MS analysis of the toxins remaining in suspension (not recognized by Ig) and, more relevantly, those remaining on the beads (recognized by Ig) allows the effectiveness and selectivity of the antivenoms studied to be determined.
Preliminary data (results)
In Echis ocellatus venom, 82.8% were identified as toxins and 17.2% as non-toxins while for Dendroaspis polylepis venom we obtained 26.2% for toxins, 39.9% for non-toxins and 33.9% for cellular components. The most expressed group of toxins were metalloproteinases for Echis ocellatus venom and three-finger toxins for Dendroaspis polylepis venom. These findings will be important in the efficacy evaluation and targeting the main toxins for the next step of the development the antivenomic
approach. Snake venoms of the genus Echis and/or Dendroaspis are responsible for a large proportion of snake envenomations in sub-Saharan Africa.Please explain why your abstract is innovative for mass spectrometry?The optimization of the antivenomics can increase the sensibility of the antivenoms targeting the most important toxins responsible for the symptoms in the envenoming and also improve the treatment response.
Keyword :Antivenomic, MALDI, Mass spectrometry, Venom.