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Recombinant dimeric small immunoproteins neutralize transmissible gastroenteritis virus infectivity efficiently in vitro and confer passive immunity in vivo

Marco Bestagno^1,

Isabel Sola^2,

¹ International Centre for Genetic Engineering and Biotechnology, AREA Science Park, Padriciano 99, 34012 Trieste, Italy
² Department of Molecular and Cell Biology, Centro Nacional de Biotecnologia, Consejo Superior de Investigaciones Cientificas, Campus Univ. Autonoma Madrid, Darwin 3, Cantoblanco, 28049 Madrid, Spain
³ Fort-Dodge Veterinaria SA, Department of Research and Development, Vall de Bianya, 17813 Girona, Spain

Correspondence
Oscar R. Burrone
burrone{at}icgeb.org;
Luis Enjuanes
L.Enjuanes{at}cnb.uam.es

Small immunoproteins (SIPs) are single-chain molecules comprising the variable regions of an antibody assembled in a single polypeptide (scFv) and joined to the immunoglobulin heavy-chain dimerizing domain. To investigate the potential of these molecules to provide protection against enteric infections when supplied orally, SIPs were generated against Transmissible gastroenteritis virus (TGEV), a highly pathogenic porcine virus. Different variants of TGEV-specific SIPs were created, of and isotypes, by exploiting the dimerizing domains CH4 and CH3 of human and swine origin. Transfected cells secreted these recombinant mini-antibodies efficiently, mainly as dimers stabilized covalently by inter-chain disulphide bridges. The specificity and functionality of the recombinant TGEV-specific SIPs were determined by in vitro binding, neutralization and infection-interference assays. The neutralization indices of the TGEV-specific SIPs were all very similar to that of the original TGEV-specific mAb, thus confirming that the immunological properties have been preserved in the recombinant SIPs. In vivo protection experiments on newborn piglets have, in addition, demonstrated a strong reduction of virus titre in infected tissues of animals treated orally with TGEV-specific SIPs. It has therefore been demonstrated that it is possible to confer passive immunization to newborn pigs by feeding them with recombinant SIPs.

These authors contributed equally to this work.