Wu Chongchao, Chen Wei, Chen Jia, Han Bo, Peng Zhou, Ge Feng, Wei Bo, Liu Mingxian, Zhang Meiying, Qian Chuiwen, Hou Zhibo, Liu Ge, Guo Chaowan, Wang Yifei, Kitazato Kaio, Yu Guoying, Zou Chunbin, Xiong Sheng
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Department of Cellular Biology, Jinan University, Guangzhou 510632, Guangdong, People's Republic of China; Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Centre for Systems Biomedicine, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China; Department of Pharmacy, College of Food and Pharmacy & Medical, Zhejiang Ocean University, Zhoushan 316002, People's Republic of China; Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, People's Republic of China; Division of Molecular Pharmacology of Infectious Agents, Department of Molecular Microbiology and Immunology, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki City, Nagasaki Prefecture 852-8521, Japan; and Department of Medicine, University of Pittsburgh, Pittsburgh, PA 15213, USA Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Department of Cellular Biology, Jinan University, Guangzhou 510632, Guangdong, People's Republic of China; Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Centre for Systems Biomedicine, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China; Department of Pharmacy, College of Food and Pharmacy & Medical, Zhejiang Ocean University, Zhoushan 316002, People's Republic of China; Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, People's Republic of China; Division of Molecular Pharmacology of Infectious Agents, Department of Molecular Microbiology and Immunology, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki City, Nagasaki Prefecture 852-8521, Japan; and Department of Medicine, University of Pittsburgh, Pittsburgh, PA 15213, USA.
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Department of Cellular Biology, Jinan University, Guangzhou 510632, Guangdong, People's Republic of China; Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Centre for Systems Biomedicine, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China; Department of Pharmacy, College of Food and Pharmacy & Medical, Zhejiang Ocean University, Zhoushan 316002, People's Republic of China; Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, People's Republic of China; Division of Molecular Pharmacology of Infectious Agents, Department of Molecular Microbiology and Immunology, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki City, Nagasaki Prefecture 852-8521, Japan; and Department of Medicine, University of Pittsburgh, Pittsburgh, PA 15213, USA.
J Biochem. 2015 Jun;157(6):539-48. doi: 10.1093/jb/mvv013. Epub 2015 Feb 20.
Influenza A virus (IAV) has been raising public health and safety concerns worldwide. Cyanovirin-N (CVN) is a prominent anti-IAV candidate, but both cytotoxicity and immunogenicity have hindered the development of this protein as a viable therapy. In this article, linker-CVN (LCVN) with a flexible and hydrophilic polypeptide at the N-terminus was efficiently produced from the cytoplasm of Escherichia coli at a >15-l scale. PEGylation at the N-terminal α-amine of LCVN was also reformed as 20 kDa PEGylated linkered Cyanovirin-N (PEG20k-LCVN). The 50% effective concentrations of PEG20k-LCVN were 0.43 ± 0.11 µM for influenza A/HK/8/68 (H3N2) and 0.04 ± 0.02 µM for A/Swan/Hokkaido/51/96 (H5N3), dramatically lower than that of the positive control, Ribavirin (2.88 ± 0.66 × 10(3) µM and 1.79 ± 0.62 × 10(3) µM, respectively). A total of 12.5 µM PEG20k-LCVN effectively inactivate the propagation of H3N2 in chicken embryos. About 2.0 mg/kg/day PEG20k-LCVN increased double the survival rate (66.67%, P = 0.0378) of H3N2 infected mice, prolonged the median survival period, downregulated the mRNA level of viral nuclear protein and decreased (attenuated) the pathology lesion in mice lung. A novel PEGylated CVN derivative, PEG20k-LCVN, exhibited potent and strain-dependent anti-IAV activity in nanomolar concentrations in vitro, as well as in micromolar concentration in vivo.
甲型流感病毒（IAV）在全球范围内引发了公众对健康和安全的担忧。氰胍蛋白-N（CVN）是一种重要的抗IAV候选药物，但细胞毒性和免疫原性都阻碍了这种蛋白质成为一种可行的治疗方法。在本文中，在大肠杆菌细胞质中以大于15升的规模高效生产了在N端带有柔性亲水性多肽的连接体-CVN（LCVN）。LCVN的N端α-胺的聚乙二醇化也被改造为20 kDa聚乙二醇化连接体氰胍蛋白-N（PEG20k-LCVN）。PEG20k-LCVN对甲型流感病毒/HK/8/68（H3N2）的50%有效浓度为0.43±0.11μM，对A/天鹅/北海道/51/96（H5N3）的50%有效浓度为0.04±0.02μM，显著低于阳性对照利巴韦林（分别为2.88±0.66×10³μM和1.79±0.62×10³μM）。总共12.5μM的PEG20k-LCVN有效地抑制了H3N2在鸡胚中的繁殖。约2.0mg/kg/天的PEG20k-LCVN使感染H3N2的小鼠存活率提高了一倍（66.67%，P = 0.0378），延长了中位生存期，下调了病毒核蛋白的mRNA水平，并减轻了小鼠肺部的病理损伤。一种新型的聚乙二醇化CVN衍生物PEG20k-LCVN在体外纳摩尔浓度以及体内微摩尔浓度下均表现出强大的、依赖毒株的抗IAV活性。
