High-affinity, human antibody-like antibody fragment (single-chain variable fragment) neutralizing the lethal factor (LF) of Bacillus anthracis by inhibiting protective antigen-LF complex formation.
The anthrax lethal toxin (LT) consists of two subunits, the protective antigen (PA) and the lethal factor (LF), and is essential for anthrax pathogenesis. Several recombinant antibodies directed against PA and intended for medical use have been obtained, but none against LF, despite the recommendations of anthrax experts. Here we describe an anti-LF single-chain variable fragment (scFv) that originated from an immunized macaque (Macaca fascicularis) and was obtained by phage display. Panning of the library of 1.8 x 10(8) clones allowed the isolation of 2LF, a high-affinity (equilibrium dissociation constant, 1.02 nM) scFv, which is highly neutralizing in the standardized in vitro assay (50% inhibitory concentration, 1.20 +/- 0.06 nM) and in an in vivo assay. The scFv neutralizes anthrax LT by inhibiting the formation of the LF-PA complex. The genes encoding 2LF are very similar to those of human immunoglobulin germ line genes, sharing substantial (84.2%) identity with their most similar, germinally encoded counterparts; this feature favors medical applications. These results, and others formerly published, demonstrate that our approach can generate antibody fragments suitable for prophylaxis and therapeutics.
Pelat
Thibaut
T
Groupe de Biotechnologie des Anticorps, Département de Biologie des Agents Transmissibles, La Tronche, France.
Hust
Michael
M
Laffly
Emmanuelle
E
Condemine
Florence
F
Bottex
Chantal
C
Vidal
Dominique
D
Lefranc
Marie-Paule
MP
Dübel
Stefan
S
Thullier
Philippe
P
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D000915:Antibody Affinity; D000942:Antigens, Bacterial / Q000008:administration & dosage / Q000276:immunology* / Q000378:metabolism; D001408:Bacillus anthracis; D001427:Bacterial Toxins / Q000008:administration & dosage / Q000276:immunology* / Q000378:metabolism; D019295:Computational Biology; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D008252:Macaca fascicularis; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D019151:Peptide Library; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D017422:Sequence Analysis, DNA
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D011994:Recombinant Proteins; C030325:anthrax toxin