Journal of Vaccines & Vaccination

Evaluating the potential of an In vitro expression system in generating diffi cult to-express Hc botulinum vaccines

International Conference & Exhibition on Vaccines & Vaccination

22-24 Nov 2011 Philadelphia Airport Marriott, USA

R. Zichel, A. Mimran, E. Diamant, A. Keren, A. Barnea, I. Steinberger- Levy, D. Marcus, A. Turgeman, and S. Reuveny

Scientific Tracks Abstracts: J Vaccines Vaccin

Abstract:

Botulinum neurotoxins produced by the anaerobic bacteria Clostridium botulinum are the most potent biological toxins in nature. Current vaccine strategies focus on the carboxy-terminal fragment of the neurotoxin (Hc) that contains most of the neutralizing epitopes. However, due to the high AT-content of clostridial genes, expression of recombinant Hc proteins in cellular systems is challenging, and was demonstrated to require codon-optimized synthetic gene. We used an alternative strategy in which the native genes encoding Hc proteins of botulinum toxins A, B and E were expressed in a cell-free expression system. Th e unique property of this open system was used to optimize solubility and expression level by introducing various chaperones, directly to the expression reaction. Expression levels of more than 1mg/ml were obtained in the continuous-exchange mode of the cell-free system. Th e purifi ed Hc proteins were detected by anti-homologue toxin sera suggesting that native immunogenic epitopes are presented on the in vitro-expressed proteins. Mice immunized with, alum-absorbed, HcA, HcB or HcE vaccines generated a high serum ELISA-titers against the homologue native toxin complex, which enabled protection against a high-dose toxin challenge (10 3 -10 6 MsLD 50 ). Moreover, immunization with a trivalent HcA, HcB and HcE vaccine protected mice against the corresponding trivalent toxin challenge (10 5 MsLD 50 ). Finally, the anti Hc sera were used to develop an ELISA for specifi c detection of native botulinum A, B or E. Our results together with the latest developments in scalability of the in vitro expression systems off er alternative routes for the preparation of diffi cult to express sub-unit vaccines.