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Infection and Immunity, July 2009, p. 2795-2801, Vol. 77, No. 7
0019-9567/09/$08.00+0     doi:10.1128/IAI.01252-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Bivalent Recombinant Vaccine for Botulinum Neurotoxin Types A and B Based on a Polypeptide Comprising Their Effector and Translocation Domains That Is Protective against the Predominant A and B Subtypes {triangledown}

Clifford Shone,1 Heidi Agostini,2 Joanna Clancy,2 Mili Gu,2 Huei-Hsiung Yang,2 Yanfang Chu,2 Virginia Johnson,2 Makie Taal,1 Joanna McGlashan,1 John Brehm,1 and Xiaomi Tong2*

Health Protection Agency, Porton Down, Salisbury, Wilts SP4 0JG, United Kingdom,1 Emergent BioSolutions, Inc., 300 Professional Drive, Gaithersburg, Maryland 208792

Received 13 October 2008/ Returned for modification 15 December 2008/ Accepted 21 April 2009

The botulinum neurotoxins (BoNTs) are a large family of extremely potent, neuroparalytic, dichain proteins which act at the peripheral nervous system. The wide genetic diversity observed with this neurotoxin family poses a significant challenge for the development of an effective botulinum vaccine. The present study describes a vaccine development platform based on protein fragments representing the N-terminal two-thirds of each toxin molecule. These fragments, designated LHN, comprise the light chain and translocation domains of each neurotoxin and are devoid of any neuron-binding activity. Using codon-optimized genes, LHN fragments derived from BoNT serotypes A and B were expressed in Escherichia coli in high yield with >1 g of purified, soluble fragment recoverable from 4.5 liter-scale fermentations. The protective efficacy of LHN/A was significantly enhanced by treatment with formaldehyde, which induced intramolecular cross-linking but virtually no aggregation of the fragment. A single immunization of the modified fragment protected mice from challenge with a 103 50% lethal dose (LD50) of BoNT/A1 with an 50% effective dose (ED50) of 50 ng of the vaccine. In similar experiments, the LHN/A vaccine was shown to protect mice against challenge with BoNT/A subtypes A1, A2, and A3, which is the first demonstration of single-dose protection by a vaccine against the principal toxin subtypes of BoNT/A. The LHN/B vaccine was also highly efficacious, giving an ED50 of ~140 ng to a challenge of 103 LD50 of BoNT/B1. In addition, LHN/B provided single-dose protection in mice against BoNT/B4 (nonproteolytic toxin subtype).

* Corresponding author. Mailing address: Emergent BioSolutions, Inc., 300 Professional Drive, Gaithersburg, MD 20879. Phone: (301) 944-0164. Fax: (301) 590-1252. E-mail: tongx{at}ebsi.com

{triangledown} Published ahead of print on 27 April 2009.

Editor: R. P. Morrison

Infection and Immunity, July 2009, p. 2795-2801, Vol. 77, No. 7
0019-9567/09/$08.00+0     doi:10.1128/IAI.01252-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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