Yang Gi-Hyeok, Yoon Sun Ok, Jang Myung Hee, Hong Hyo Jeong
Therapeutic Antibody Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejon, Republic of Korea.
J Microbiol. 2007 Dec;45(6):528-33.
In a previous study we generated an anti-Hepatitis B Virus (HBV) preS1 humanized antibody (HzKR127) that showed in vivo HBV-neutralizing activity in chimpanzees. However, the antigen-binding affinity of the humanized antibody may not be sufficient for clinical use and thus affinity maturation is required for better therapeutic efficacy. In this study, phage display technique was employed to increase the affinity of HzKR127. All six amino acid residues (Glu95-Tyr96-Asp97-Glu98-Ala99-Tyr100) in the heavy (H) chain complementarydetermining region 3 (HCDR3) of HzKR127 were randomized and phage-displayed single chain Fv (scFv) library was constructed. After three rounds of panning, 12 different clones exhibiting higher antigen-binding activity than the wild type ScFv were selected and their antigen-binding specificity for the preS1 confirmed. Subsequently, five ScFv clones were converted to whole IgG and subjected to affinity determination. The results showed that two clones (B3 and A19) exhibited an approximately 6 fold higher affinities than that of HzKR127. The affinity-matured humanized antibodies may be useful in anti-HBV immunotherapy.
在之前的一项研究中，我们制备了一种抗乙型肝炎病毒（HBV）前S1人源化抗体（HzKR127），该抗体在黑猩猩体内显示出HBV中和活性。然而，人源化抗体的抗原结合亲和力可能不足以用于临床，因此需要进行亲和力成熟以获得更好的治疗效果。在本研究中，采用噬菌体展示技术提高HzKR127的亲和力。对HzKR127重链互补决定区3（HCDR3）中的所有六个氨基酸残基（Glu95 - Tyr96 - Asp97 - Glu98 - Ala99 - Tyr100）进行随机化处理，并构建噬菌体展示单链Fv（scFv）文库。经过三轮淘选，选择了12个与野生型ScFv相比具有更高抗原结合活性的不同克隆，并确认了它们对前S1的抗原结合特异性。随后，将五个ScFv克隆转化为完整的IgG并进行亲和力测定。结果表明，两个克隆（B3和A19）的亲和力比HzKR127高约6倍。亲和力成熟的人源化抗体可能在抗HBV免疫治疗中有用。
