Ahn Hye-Mi, Ryu Jihye, Song Jin Myeong, Lee Yunhee, Kim Hye-Jin, Ko Dongjoon, Choi Inpyo, Kim Sang Jick, Lee Jung Weon, Kim Semi
Immunotherapy Convergence Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejon, Korea.
Department of Pharmacy, College of Pharmacy, Seoul National University, Seoul, Korea.
Theranostics. 2017 Jan 11;7(3):594-613. doi: 10.7150/thno.15629. eCollection 2017.
The transmembrane four L6 family member 5 (TM4SF5) protein is a novel molecular target for the prevention and treatment of hepatocellular carcinoma. TM4SF5 is highly expressed in liver, colon, esophageal, and pancreatic cancers and is implicated in tumor progression. Here, we screened monoclonal antibodies that specifically bound to the extracellular loop 2 (EC2) of TM4SF5 from a phage-displayed murine antibody (single-chain variable fragment; scFv) library. We constructed and characterized chimeric antibodies, Ab27 and Ab79, of scFv fused with Fc domain of human IgG1. The affinity (K) of Ab27 and Ab79 for soluble EC2 was approximately 9.2 nM and 16.9 nM, respectively, as determined by surface plasmon resonance analysis. Ab27 and Ab79 efficiently bound to native TM4SF5 on the cell surface were internalized into the cancer cells, leading to a decrease in cell surface TM4SF5. Ab27 and Ab79 inhibited the proliferation and invasion of TM4SF5-positive liver and colon cancer cells and reduced FAK and c-Src phosphorylation. Ab27 and Ab79 also enhanced anoikis sensitivity and reduced survivin. Ab27 mediated antibody-dependent cell-mediated cytotoxicity . Ab27 and Ab79 efficiently inhibited tumor growth in a liver cancer xenograft model. These results strongly support the further development of Ab27 as a novel anti-cancer agent in the clinic.
跨膜四区L6家族成员5（TM4SF5）蛋白是预防和治疗肝细胞癌的新型分子靶点。TM4SF5在肝癌、结肠癌、食管癌和胰腺癌中高表达，并与肿瘤进展有关。在此，我们从噬菌体展示的鼠源抗体（单链可变片段；scFv）文库中筛选出特异性结合TM4SF5细胞外环2（EC2）的单克隆抗体。我们构建并鉴定了scFv与人IgG1的Fc结构域融合的嵌合抗体Ab27和Ab79。通过表面等离子体共振分析测定，Ab27和Ab79对可溶性EC2的亲和力（K）分别约为9.2 nM和16.9 nM。Ab27和Ab79有效结合细胞表面的天然TM4SF5，并被内化到癌细胞中，导致细胞表面TM4SF5减少。Ab27和Ab79抑制TM4SF5阳性肝癌和结肠癌细胞的增殖和侵袭，并降低FAK和c-Src磷酸化水平。Ab27和Ab79还增强了失巢凋亡敏感性并降低了生存素水平。Ab27介导抗体依赖的细胞介导的细胞毒性作用。Ab27和Ab79在肝癌异种移植模型中有效抑制肿瘤生长。这些结果有力地支持了将Ab27进一步开发为临床新型抗癌药物。
