Hao Lina, Zheng Feng, Xiong Siping, Hu Dan, Lv Heng, Tang Qi, Yang Jin, Feng Zhenqing, Wang Changjun, Zhu Jin
Huadong Medical Institute of Biotechniques, Nanjing 210002, China.
Key Laboratory of Antibody Technique of Ministry of Health, Nanjing Medical University, Nanjing 210029, China.
Int J Mol Sci. 2014 Oct 14;15(10):18496-507. doi: 10.3390/ijms151018496.
The aim of this research is to develop a human/murine chimeric Fab antibody which neutralizes the anthrax toxin, protective antigen (PA). The chimeric Fab was constructed using variable regions of murine anti-PA monoclonal antibody in combination with constant regions of human IgG. The chimeric PA6-Fab was expressed in E. coli. BL21 and evaluated by ELISA and co-immunoprecipitation- mass spectra. The potency of PA6-Fab to neutralize LeTx was examined in J774A.1 cell viability in vitro and in Fisher 344 rats in vivo. The PA6-Fab did not have domain similarity corresponding to the current anti PA mAbs, but specifically bound to anthrax PA at an affinity of 1.76 nM, and was able to neutralize LeTx in vitro and protected 56.9% cells at 20 μg/mL against anthrax LeTx. One hundred μg PA6-Fab could neutralize 300 μg LeTx in vivo. The PA6-Fab has potential as a therapeutic mAb for treatment of anthrax.
本研究的目的是开发一种能中和炭疽毒素保护性抗原（PA）的人/鼠嵌合Fab抗体。该嵌合Fab是利用鼠抗PA单克隆抗体的可变区与人IgG的恒定区构建而成。嵌合PA6-Fab在大肠杆菌BL21中表达，并通过酶联免疫吸附测定（ELISA）和免疫共沉淀-质谱进行评估。在体外J774A.1细胞活力实验以及体内Fisher 344大鼠实验中检测了PA6-Fab中和致死毒素（LeTx）的效力。PA6-Fab与目前的抗PA单克隆抗体没有结构域相似性，但能以1.76 nM的亲和力特异性结合炭疽PA，并能在体外中和LeTx，在20μg/mL时可保护56.9%的细胞免受炭疽LeTx的侵害。100μg PA6-Fab在体内能中和300μg LeTx。PA6-Fab具有作为治疗炭疽的治疗性单克隆抗体的潜力。
