Humanization of agonistic anti-human 4-1BB monoclonal antibody using a phage-displayed combinatorial library.
Given the key role 4-1BB plays in the stimulation of T cells, humanization of agonistic anti-human 4-1BB monoclonal antibody (mAb) may have important clinical applications. In this paper, we present the humanization of agonistic anti-human 4-1BB mAb, BBK-4, using a phage display library. We first prepared the combinatorial library by incorporating murine and human alternative at positions representing buried residues that might affect the structural integrity of the antigen binding site. Six humanized single chain Fv (scFv) fragments were selected from the combinatorial library expressing phage-displayed humanized scFv. They were found to retain the epitope specificity of the original mAb but had affinities of lower than 1/10 of the original. In spite of the lower affinity, the humanized scFv coated on the surface expanded human peripheral blood mononuclear cells (PBMCs) in MLR similarly to the original mAb in the presence of anti-CD3 mAb. These results suggest that humanized anti-human 4-1BB scFvs can be used as a valuable reagent for clinical application.
Son
Ji Hee
JH
Department of Biological Sciences and Immunomodulation Research Center, University of Ulsan, Ulsan 680-749, South Korea.
Lee
Unn Hwa
UH
Lee
Jeong Jin
JJ
Kwon
Byungsuk
B
Kwon
Byoung Se
BS
Park
Jeong Woo
JW
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D015703:Antigens, CD; D001483:Base Sequence; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D005434:Flow Cytometry; D015723:Gene Library; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D007371:Interferon-gamma / Q000276:immunology / Q000378:metabolism; D015847:Interleukin-4 / Q000276:immunology / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D019151:Peptide Library; D016133:Polymerase Chain Reaction; D017475:Receptors, Nerve Growth Factor / Q000819:agonists* / Q000276:immunology*; D018124:Receptors, Tumor Necrosis Factor / Q000819:agonists* / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D016415:Sequence Alignment; D020349:Surface Plasmon Resonance; D053261:Tumor Necrosis Factor Receptor Superfamily, Member 9
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D007135:Immunoglobulin Variable Region; D019151:Peptide Library; D017475:Receptors, Nerve Growth Factor; D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins; C505714:TNFRSF9 protein, human; D053261:Tumor Necrosis Factor Receptor Superfamily, Member 9; D015847:Interleukin-4; D007371:Interferon-gamma; D004247:DNA