Son Ji Hee, Lee Unn Hwa, Lee Jeong Jin, Kwon Byungsuk, Kwon Byoung Se, Park Jeong Woo
Department of Biological Sciences and Immunomodulation Research Center, University of Ulsan, Ulsan 680-749, South Korea.
J Immunol Methods. 2004 Mar;286(1-2):187-201. doi: 10.1016/j.jim.2004.01.006.
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.
鉴于4-1BB在T细胞刺激中发挥的关键作用，激动性抗人4-1BB单克隆抗体（mAb）的人源化可能具有重要的临床应用价值。在本文中，我们展示了利用噬菌体展示文库对激动性抗人4-1BB mAb BBK-4进行人源化的过程。我们首先通过在可能影响抗原结合位点结构完整性的埋藏残基位置引入鼠源和人源替代物来制备组合文库。从表达噬菌体展示人源化单链Fv（scFv）的组合文库中筛选出6个人源化单链Fv片段。发现它们保留了原始mAb的表位特异性，但亲和力低于原始mAb的1/10。尽管亲和力较低，但在存在抗CD3 mAb的情况下，包被在表面的人源化scFv在混合淋巴细胞反应（MLR）中与原始mAb类似地促进了人外周血单个核细胞（PBMC）的扩增。这些结果表明，人源化抗人4-1BB scFv可作为一种有价值的临床应用试剂。
