Passive immunization of pigs with bispecific llama single-domain antibody fragments against foot-and-mouth disease and porcine immunoglobulin

Foot-and-mouth disease (FMD) is a contagious viral disease of cloven-hoofed animals that occasionally causes outbreaks in Europe. We aim to develop an immunotherapy that confers rapid protection against FMD in outbreak situations. For this purpose, we previously isolated llama single-domain antibody fragments (VHHs) binding to FMDV or porcine immunoglobulin (pIg). The pig binding VHHs can be genetically fused to other VHHs, resulting in so-called VHH2s. As compared to non-plg binding VHHs such VHH2s have a 100-fold increased serum half-life which is essential for effective immunotherapy. We have now produced three bispecific VHH2s by fusion of three FMDV binding VHHs (clones M3, M8 and M23) to a pig binding VHH (VI-4). The resulting yeast-produced VHH2s bound FMDV and pIg with high affinity (K-D about 1 nM) and neutralized FMDV in vitro as efficiently as their monovalent counterparts. To evaluate their therapeutic potential all three VHH2s were intramuscularly injected into pigs that were challenge infected with FMDV 24 h later. Administration of one of these VHH2s (M23ggsVI-4) reduced the viremia significantly (P = 0.0034) and reduced viral shedding almost significantly (P = 0.11). However, it did not prevent development of clinical signs or transmission of FMDV. These results suggest that immunotherapy using bispecific VHH2s binding to FMDV and pig is possible in principle, but should be improved by increasing VHH2 dosage or using more potent VHH2s. (C) 2008 Elsevier B.V. All rights reserved.