Please wait a minute...

中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志
中国生物工程杂志  2020, Vol. 40 Issue (10): 24-34    DOI: 10.13523/j.cb.2006058
综述     
纳米抗体在传染病的预防、诊断和治疗中的应用 *
梅雅贤,王玥,罗文新()
厦门大学公共卫生学院 国家传染病诊断试剂与疫苗工程技术研究中心 厦门 361102
Application of Nano-antibody in the Prevention, Diagnosis and Treatment of Infectious Diseases
MEI Ya-xian,WANG Yue,LUO Wen-xin()
State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics & National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen 361102, China
 全文: PDF(5988 KB)   HTML
摘要：

传染病是一种由致病性微生物引起,能够影响人类身体健康甚至引发严重社会危机的传播性疾病。近年来,新冠、埃博拉等传染病的恶性暴发促使人们寻找更为高效便捷的防治手段以遏制疾病的进程。抗体在传染病防治中的应用引起了广泛关注,palivizumab是目前唯一被批准应用于呼吸道合胞病毒在免疫力低下人群的预防的单克隆抗体。纳米抗体(nano-antibody, Nb)是目前已知的能与抗原稳定结合的最小功能性单域抗体,具有稳定性高、亲水性强、易于表达和改造等优势。独特的分子特性使其在病毒、细菌、寄生虫等引发的传染病的预防、诊断和治疗中展现出良好的应用前景,相关研究显示纳米抗体对艾滋、流感、新型冠状病毒等都有很好的治疗效果。重点叙述纳米抗体的结构特点及其在传染性疾病中的研究进展。
关键词： 传染性疾病;  纳米抗体;  预防;  诊断;  治疗    
Abstract:

Infectious disease mainly refer to transmissible diseases caused by pathogenic microorganisms. It can affect human health and even cause serious social crisis. In recent years, the outbreaks of infectious diseases like COVID-19 and ebola have prompted people to seek more efficient and convenient means to control and prevent infectious diseases. As an effective way currently, antibody has drawn people’s attention. However, palivizumab, for the prevention and treatment of respiratory syncytial virus, is the only one monoclonal antibody approved for infectious diseases. Nano-antibody (Nb) is the smallest known functional antibody capable of stably binding to antigens. Nb has the advantages of high stability, strong hydrophilicity, easy production through microbial systems, easy modification, etc. Due to its unique molecular properties, Nb has shown promising application prospects in the prevention, diagnosis and treatment of infectious diseases caused by viruses, bacteria, parasites, etc. Related studies have shown that Nb has good therapeutic effects on AIDS, influenza, novel coronavirus, etc. This paper focuses on the structural characteristics of Nb and its research progress in infectious diseases.
Key words: Infectious diseases    Nano-antibody    Prevention    Diagnosis    Treatment
收稿日期: 2020-06-30 出版日期: 2020-11-10
ZTFLH:  Q819  
基金资助: * 国家自然科学基金(31870925)
通讯作者: 罗文新     E-mail: wxluo@xmu.edu.cn
服务　　
把本文推荐给朋友
加入引用管理器
E-mail Alert
RSS
作者相关文章　　
梅雅贤
王玥
罗文新

引用本文:

梅雅贤,王玥,罗文新. 纳米抗体在传染病的预防、诊断和治疗中的应用 *[J]. 中国生物工程杂志, 2020, 40(10): 24-34.

MEI Ya-xian,WANG Yue,LUO Wen-xin. Application of Nano-antibody in the Prevention, Diagnosis and Treatment of Infectious Diseases. China Biotechnology, 2020, 40(10): 24-34.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.2006058    或    https://manu60.magtech.com.cn/biotech/CN/Y2020/V40/I10/24

图1  传统单克隆抗体、重链抗体、纳米抗体结构示意图
传统单克隆抗体 重链抗体 纳米抗体 纳米抗体优势 参考文献
分子量 150kDa 28kDa 15kDa 组织穿透性更高
半衰期更短
肾脏清除率更高
CDR3区氨基酸数目 8~15(VH) 8~15(VH) 3~28 抗体库多样性
识别隐藏表位		 [11]
可变区二硫键 - - CDR3与CDR1或FR2间存在二硫键 稳定性较强 [31]
序列同源性 - - >80%(与VH相比) 同源性高 [31]
FR2氨基酸差异 Val,Gly,Leu,Trp(VH) Val,Gly,Leu,Trp(VH) Phe或Tyr,Glu,Arg,Gly 亲水性强
组成方式 两条重链,两条轻链通过二硫键连接组成 两条重链通过二硫键连接组成,缺乏轻链和重链CH1区 由重链可变区(单蛋白域)组成 易于与其它蛋白分子融合,进行蛋白工程改造
表1  抗体特点对比
图2  VH和VHH结构示意图
作者 国家 抗体来源 靶向位点 发表时间 研究进展
Dong等[60] 美国 天然库和合成库 S蛋白 2020-05-22 构建的双特异性纳米抗体显示了有效的S/ACE2结合阻断作用;构建了人源化纳米抗体降低免疫原性
Wrapp 等[62] 美国 SARS-CoV-1和
MERS-CoV S蛋白的免疫库		 靶向SARS-CoV-1
RBD区的纳米抗体,与SARS-CoV-2 RBD区发生交叉反应		 2020-05-28 构建的一株二价Nb-Fc融合蛋白中和带有SARS-CoV-2 S蛋白假病毒,IC50值为0.2μg/ml
Beroni集团和天津大学[61] 澳大利亚,
中国		 - S蛋白,核衣壳蛋白 2020-05-08 靶向S蛋白的纳米抗体计划用于抗病毒药物,靶向核衣壳蛋白的纳米抗体计划用于诊断试验的标记物
Huo等[63] 英国 天然库(随机诱变提升亲和力) SARS-CoV-2 RBD
(H11-D4 和H11-H4两株纳米抗体)		 2020-07-13 Nb-Fc融合蛋白对SARS-CoV-2具有病毒中和活性; 与单克隆抗体CR3022具有协同中和作用
Walter等[64] 瑞士 合成库 SARS-CoV-2 RBD 2020-04-16
Nieto等[65] 智利 免疫库 SARS-CoV-2 RBD 2020-06-09 纳米抗体株与SARS-CoV-2 RBD亲和力达KD~295±84pM
Li等[66] 中国 合成库 SARS-CoV-2 RBD 2020-06-09 与SARS-CoV-2 RBD亲和力达KD~1.0nmol/L; 与SARS-CoV-2假病毒中和 IC50为0.40μg/ml
Custódio等[67] 德国 合成库 SARS-CoV-2 RBD(其中一株Sb23竞争结合ACE2结合位点) 2020-06-23 Sb23中和带有SARS-CoV-2 S蛋白假病毒,IC50为0.6μg/ml
Hanke等[68] 瑞典 免疫库 SARS-CoV-2 RBD 2020-06-02 纳米抗体株Ty1 与RBD亲和力为KD 5-50nmol/L,与SARS-CoV-2假病毒颗粒中和IC50值为0.77μg/ml
表2  SARS-CoV-2特异性纳米抗体
[1] Sanaei M, Setayesh N, Sepehrizadeh Z, et al. Nanobodies in human infections: prevention, detection, and treatment. Immunological Investigations, 2019: 1-22.
doi: 10.1080/08820139.2020.1817934 pmid: 32954867
[2] Statistics N B O. National data.[2020-06-03]. http://data.stats.gov.cn/search.htm?s=%E4%BC%A0%E6%9F%93%E7%97%85.
doi: 10.3390/data4010020 pmid: 30956970
[3] Worldmeter. Covid-19 coronavirus pandemic. [2020-06-03]. https://www.worldometers.info/coronavirus/.
[4] 百度. 疫情实时大数据报告. [2020-06-03]. https://voice.baidu.com/act/newpneumonia/newpneumonia#tab4.
[5] Hey A. History and practice: Antibodies in infectious diseases. Microbiology Spectrum, 2015, 3(2): AID-0026-2014.
doi: 10.1128/microbiolspec.MDNA3-0058-2014 pmid: 26104699
[6] Steeland S, Vandenbroucke R E, Libert C. Nanobodies as therapeutics: Big opportunities for small antibodies. Drug Discovery Today, 2016,21(7):1076-1113.
doi: 10.1016/j.drudis.2016.04.003 pmid: 27080147
[7] Hassanzadeh-Ghassabeh G, Devoogdt N, De Pauw P, et al. Nanobodies and their potential applications. Nanomedicine, 2013,8(6):1013-1026.
doi: 10.2217/nnm.13.86 pmid: 23730699
[8] D’huyvetter M, Vincke C, Xavier C, et al. Targeted radionuclide therapy with a 177lu-labeled anti-her2 nanobody. Theranostics, 2014,4(7):708-720.
doi: 10.7150/thno.8156 pmid: 24883121
[9] Oliveira S, Van Dongen G a M S, Walsum M S-V, et al. Rapid visualization of human tumor xenografts through optical imaging with a near-infrared fluorescent anti-epidermal growth factor receptor nanobody. Molecular Imaging, 2012, 11(1): 7290.2011.00025.
pmid: 22418021
[10] Abulrob A, Sprong H, Van Bergen En Henegouwen P, et al. The blood-brain barrier transmigrating single domain antibody: mechanisms of transport and antigenic epitopes in human brain endothelial cells. Journal of Neurochemistry, 2005,95(4):1201-1214.
doi: 10.1111/j.1471-4159.2005.03463.x pmid: 16271053
[11] Bannas P, Hambach J, Koch-Nolte F. Nanobodies and nanobody-based human heavy chain antibodies as antitumor therapeutics. Frontiers in Immunology, 2017,8:1603-1603.
doi: 10.3389/fimmu.2017.01603 pmid: 29213270
[12] Muyldermans S, Baral T N, Retarnozzo V C, et al. Camelid immunoglobulins and nanobody technology. Veterinary Immunology and Immunopathology, 2009,128(1-3):178-183.
doi: 10.1016/j.vetimm.2008.10.299 pmid: 19026455
[13] Harmsen M M, De Haard H J. Properties, production, and applications of camelid single-domain antibody fragments. Applied Microbiology and Biotechnology, 2007,77(1):13-22.
doi: 10.1007/s00253-007-1142-2 pmid: 17704915
[14] Stijlemans B, Conrath K, Cortez-Retamozo V, et al. Efficient targeting of conserved cryptic epitopes of infectious agents by single domain antibodies - african trypanosomes as paradigm. Journal of Biological Chemistry, 2004,279(2):1256-1261.
doi: 10.1074/jbc.M307341200 pmid: 14527957
[15] Saerens D, Conrath K, Govaert J, et al. Disulfide bond introduction for general stabilization of immunoglobulin heavy-chain variable domains. Journal of Molecular Biology, 2008,377(2):478-488.
doi: 10.1016/j.jmb.2008.01.022 pmid: 18262543
[16] Li M, Fan X D, Liu J, et al. Selection by phage display of nanobodies directed against hypoxia inducible factor-1 alpha (hif-1 alpha). Biotechnology and Applied Biochemistry, 2015,62(6):738-745.
doi: 10.1002/bab.1340 pmid: 25556956
[17] Dumoulin M, Conrath K, Van Meirhaeghe A, et al. Single-domain antibody fragments with high conformational stability. Protein Science, 2002,11(3):500-515.
doi: 10.1110/ps.34602 pmid: 11847273
[18] Omidfar K, Daneshpour M. Advances in phage display technology for drug discovery. Expert Opinion on Drug Discovery, 2015,10(6):651-669.
doi: 10.1517/17460441.2015.1037738 pmid: 25910798
[19] Van Audenhove I, Boucherie C, Pieters L, et al. Stratifying fascin and cortactin function in invadopodium formation using inhibitory nanobodies and targeted subcellular delocalization. FASEB Journal, 2014,28(4):1805-1818.
doi: 10.1096/fj.13-242537 pmid: 24414419
[20] Van Den Abbeele A, De Clercq S, De Ganck A, et al. A llama-derived gelsolin single-domain antibody blocks gelsolin-g-actin interaction. Cellular and Molecular Life Sciences, 2010,67(9):1519-1535.
doi: 10.1007/s00018-010-0266-1 pmid: 20140750
[21] Van Audenhove I, Van Impe K, Ruano-Gallego D, et al. Mapping cytoskeletal protein function in cells by means of nanobodies. Cytoskeleton, 2013,70(10):604-622.
doi: 10.1002/cm.21122 pmid: 23818458
[22] Su C, Nguyen V K, Nei M. Adaptive evolution of variable region genes encoding an unusual type of immunoglobulin in camelids. Molecular Biology and Evolution, 2002,19(3):205-215.
doi: 10.1093/oxfordjournals.molbev.a004073 pmid: 11861879
[23] Harmsen M M, Ruuls R C, Nijman I J, et al. Llama heavy-chain v regions consist of at least four distinct subfamilies revealing novel sequence features. Molecular Immunology, 2000,37(10):579-590.
doi: 10.1016/s0161-5890(00)00081-x pmid: 11163394
[24] Vincke C, Loris R, Saerens D, et al. General strategy to humanize a camelid single-domain antibody and identification of a universal humanized nanobody scaffold. Journal of Biological Chemistry, 2009,284(5):3273-3284.
doi: 10.1074/jbc.M806889200 pmid: 19010777
[25] Vaneycken I, Govaert J, Vincke C, et al. In vitro analysis and in vivo tumor targeting of a humanized, grafted nanobody in mice using pinhole spect/micro-ct. Journal of Nuclear Medicine, 2010,51(7):1099-1106.
doi: 10.2967/jnumed.109.069823 pmid: 20554727
[26] Liu W, Song H, Chen Q, et al. Recent advances in the selection and identification of antigen-specific nanobodies. Molecular Immunology, 2018,96:37-47.
doi: 10.1016/j.molimm.2018.02.012 pmid: 29477934
[27] Salvador J P, Vilaplana L, Marco M P. Nanobody: outstanding features for diagnostic and therapeutic applications. Analytical and Bioanalytical Chemistry, 2019,411(9):1703-1713.
doi: 10.1007/s00216-019-01633-4 pmid: 30734854
[28] Sheehan J, Marasco W A. Phage and yeast display, 2015,3(1).
[29] Hussack G, Baral T N, Baardsnes J, et al. A novel affinity tag, abtag, and its application to the affinity screening of single-domain antibodies selected by phage display. Frontiers in Immunology, 2017,8:1406-1406.
doi: 10.3389/fimmu.2017.01406 pmid: 29163485
[30] Koide A, Tereshko V, Uysal S, et al. Exploring the capacity of minimalist protein interfaces: interface energetics and affinity maturation to picomolar kd of a single-domain antibody with a flat paratope. Journal of Molecular Biology, 2007,373(4):941-953.
doi: 10.1016/j.jmb.2007.08.027 pmid: 17888451
[31] Li M, Fan X, Liu J, et al. Selection by phage display of nanobodies directed against hypoxia inducible factor-1α (hif-1α). Biotechnology and Applied Biochemistry, 2015,62(6):738-45.
doi: 10.1002/bab.1340 pmid: 25556956
[32] Vanmarsenille C, Del Olmo I D, Elseviers J, et al. Nanobodies targeting conserved epitopes on the major outer membrane protein of campylobacter as potential tools for control of campylobacter colonization. Veterinary Research, 2017,48(1):86.
doi: 10.1186/s13567-017-0491-9 pmid: 29216932
[33] Keller M A, Stiehm E R. Passive immunity in prevention and treatment of infectious diseases. Clinical Microbiology Reviews, 2000,13(4):602-614.
doi: 10.1128/cmr.13.4.602-614.2000 pmid: 11023960
[34] Ruano-Gallego D, Yara D A, Di Ianni L, et al. A nanobody targeting the translocated intimin receptor inhibits the attachment of enterohemorrhagic E. Coli to human colonic mucosa. PLoS Pathogens, 2019,15(8):e1008031.
doi: 10.1371/journal.ppat.1008031 pmid: 31465434
[35] Bakshi S, Garcia R S, Van Der Weken H, et al. Evaluating single-domain antibodies as carriers for targeted vaccine delivery to the small intestinal epithelium. Journal of Controlled Release, 2020,321:416-429.
doi: 10.1016/j.jconrel.2020.01.033 pmid: 31981657
[36] Van Der Vaart J M, Pant N, Wolvers D, et al. Reduction in morbidity of rotavirus induced diarrhoea in mice by yeast produced monovalent llama-derived antibody fragments. Vaccine, 2006,24(19):4130-4137.
doi: 10.1016/j.vaccine.2006.02.045 pmid: 16616802
[37] Martin M C, Pant N, Ladero V, et al. Integrative expression system for delivery of antibody fragments by lactobacilli. Applied and Environmental Microbiology, 2011,77(6):2174-2179.
doi: 10.1128/AEM.02690-10 pmid: 21257814
[38] Tokuhara D, Alvarez B, Mejima M, et al. Rice-based oral antibody fragment prophylaxis and therapy against rotavirus infection. Journal of Clinical Investigation, 2013,123(9):3829-3838.
doi: 10.1172/JCI70266 pmid: 23925294
[39] Gray E R, Brookes J C, Caillat C, et al. Unravelling the molecular basis of high affinity nanobodies against hiv p24: in vitro functional, structural, and in silico insights. ACS Infectious Diseases, 2017,3(7):479-491.
doi: 10.1021/acsinfecdis.6b00189 pmid: 28591513
[40] Zhu M, Gong X, Hu Y, et al. Streptavidin-biotin-based directional double nanobody sandwich elisa for clinical rapid and sensitive detection of influenza h5n1. Journal of Translational Medicine, 2014,12:352.
doi: 10.1186/s12967-014-0352-5 pmid: 25526777
[41] Rasoulinejad S, Gargari S L M. Aptamer-nanobody based elasa for specific detection of acinetobacter baumannii isolates. Journal of Biotechnology, 2016,231:46-54.
doi: 10.1016/j.jbiotec.2016.05.024 pmid: 27234880
[42] Deckers N, Saerens D, Kanobana K, et al. Nanobodies, a promising tool for species-specific diagnosis of taenia solium cysticercosis. International Journal for Parasitology, 2009,39(5):625-633.
doi: 10.1016/j.ijpara.2008.10.012 pmid: 19041315
[43] He Y, Ren Y, Guo B, et al. Development of a specific nanobody and its application in rapid and selective determination of salmonella enteritidis in milk. Food Chemistry, 2020,310:125942.
doi: 10.1016/j.foodchem.2019.125942 pmid: 31830714
[44] Melli L J, Zylberman V, Hiriart Y, et al. Development and evaluation of a novel vhh-based immunocapture assay for high-sensitivity detection of shiga toxin type 2 (stx2) in stool samples. Journal of Clinical Microbiology, 2020,58(3):e01566-19.
doi: 10.1128/JCM.01566-19 pmid: 31826960
[45] Cao J, Zhong N, Wang G, et al. Nanobody-based sandwich reporter system for living cell sensing influenza a virus infection. Scientific Reports, 2019,9(1):15899.
doi: 10.1038/s41598-019-52258-7 pmid: 31685871
[46] Caljon G, Caveliers V, Lahoutte T, et al. Using microdialysis to analyse the passage of monovalent nanobodies through the blood-brain barrier. British Journal of Pharmacology, 2012,165(7):2341-2353.
doi: 10.1111/j.1476-5381.2011.01723.x pmid: 22013955
[47] Nye S, Whitley R J, Kong M. Viral infection in the development and progression of pediatric acute respiratory distress syndrome. Frontiers in Pediatrics, 2016,4:128.
doi: 10.3389/fped.2016.00128 pmid: 27933286
[48] Detalle L, Stohr T, Palomo C, et al. Generation and characterization of alx-0171, a potent novel therapeutic nanobody for the treatment of respiratory syncytial virus infection. Antimicrobial Agents and Chemotherapy, 2016,60(1):6-13.
doi: 10.1128/AAC.01802-15 pmid: 26438495
[49] Mora A L, Detalle L, Gallup J M, et al. Delivery of alx-0171 by inhalation greatly reduces respiratory syncytial virus disease in newborn lambs. mAbs, 2018,10(5):778-795.
doi: 10.1080/19420862.2018.1470727 pmid: 29733750
[50] Palomo C, Mas V, Detalle L, et al. Trivalency of a nanobody specific for the human respiratory syncytial virus fusion glycoprotein drastically enhances virus neutralization and impacts escape mutant selection. Antimicrobial Agents and Chemotherapy, 2016,60(11):6498-6509.
doi: 10.1128/AAC.00842-16 pmid: 27550346
[51] Broadbent L, Parke H G, Ferguson L J, et al. Comparative therapeutic potential of alx-0171 and palivizumab against rsv clinical isolate infection of well-differentiated primary pediatric bronchial epithelial cell cultures. Bio Rxiv, 2019: 800326.
[52] Tillib S V, Ivanova T I, Vasilev L A, et al. Formatted single-domain antibodies can protect mice against infection with influenza virus (h5n2). Antiviral Research, 2013,97(3):245-254.
doi: 10.1016/j.antiviral.2012.12.014 pmid: 23274623
[53] Tome-Amat J, Ramos I, Amanor F, et al. Influenza a virus utilizes low-affinity, high-avidity interactions with the nuclear import machinery to ensure infection and immune evasion. Journal of Virology, 2019,93(1):e01046.
doi: 10.1128/JVI.01046-18 pmid: 30305352
[54] Ibanez L I, De Filette M, Hultberg A, et al. Nanobodies with in vitro neutralizing activity protect mice against h5n1 influenza virus infection. The Journal of Infectious Diseases, 2011,203(8):1063-1072.
doi: 10.1093/infdis/jiq168 pmid: 21450996
[55] Wei G, Meng W, Guo H, et al. Potent neutralization of influenza a virus by a single-domain antibody blocking m2 ion channel protein. PLoS One, 2011,6(12):e28309.
doi: 10.1371/journal.pone.0028309 pmid: 22164266
[56] Laursen N S, Friesen R H E, Zhu X, et al. Universal protection against influenza infection by a multidomain antibody to influenza hemagglutinin. Science, 2018,362(6414):598-602.
doi: 10.1126/science.aaq0620 pmid: 30385580
[57] Stalin Raj V, Okba N M A, Gutierrez-Alvarez J, et al. Chimeric camel/human heavy-chain antibodies protect against mers-cov infection. Science Advances, 2018, 4(8): eaas9667.
doi: 10.1126/sciadv.aas9667 pmid: 30101189
[58] Wrapp D, De Vlieger D, Corbett K S, et al. Structural basis for potent neutralization of betacoronaviruses by single-domain camelid antibodies. Cell, 2020,181(5):1004-1015.
doi: 10.1016/j.cell.2020.04.031 pmid: 32375025
[59] Zhao G Y, He L, Sun S H, et al. A novel nanobody targeting Middle East respiratory syndrome coronavirus (mers-cov) receptor-binding domain has potent cross-neutralizing activity and protective efficacy against mers-cov. Journal of Virology, 2018,92(18):e00837.
doi: 10.1128/JVI.00837-18 pmid: 29950421
[60] Dong J, Huang B, Jia Z, et al. Development of multi-specific humanized llama antibodies blocking sars-cov-2/ace2 interaction with high affinity and avidity. Emerg Microbes Infect, 2020,9(1):1034-1036.
doi: 10.1080/22221751.2020.1768806 pmid: 32403995
[61] Limited B G. Beroni group r&d team identifies 24 types of nanobodies for rapid detection and treatment of coronavirus (covid-19). [2020-05-08]. https://www.globenewswire.com/news-release/2020/05/08/2030400/0/en/Beroni-Group-R-D-Team-Identifies-24-Types-of-Nanobodies-for-Rapid-Detection-and-Treatment-of-Coronavirus-COVID-19.html.
[62] Wrapp D, De Vlieger D, Corbett K S, et al. Structural basis for potent neutralization of betacoronaviruses by single-domain camelid antibodies. Cell, 2020, 181(5): 1004-1015.e15.
doi: 10.1016/j.cell.2020.04.031 pmid: 32375025
[63] Huo J, Le Bas A, Ruza R R, et al. Neutralizing nanobodies bind sars-cov-2 spike rbd and block interaction with ace2. Nature Structural & Molecular Biology, 2020.
doi: 10.1038/s41594-020-0504-7 pmid: 32958947
[64] Walter J D, Hutter C a J, Zimmermann I, et al. Synthetic nanobodies targeting the sars-cov-2 receptor-binding domain. Bio Rxiv, 2020: 2020.04.16.045419.
[65] Nieto G V, Jara R, Himelreichs J, et al. Fast isolation of sub-nanomolar affinity alpaca nanobody against the spike rbd of sars-cov-2 by combining bacterial display and a simple single-step density gradient selection. Bio Rxiv, 2020: 2020.06.09.137935.
[66] Li T, Cai H, Yao H, et al. Potent synthetic nanobodies against sars-cov-2 and molecular basis for neutralization. Bio Rxiv, 2020: 2020.06.09.143438.
[67] Custódio T F, Das H, Sheward D J, et al. Selection, biophysical and structural analysis of synthetic nanobodies that effectively neutralize sars-cov-2. Bio Rxiv, 2020: 2020.06.23.165415.
[68] Hanke L, Vidakovics M, Sheward D, et al. An alpaca nanobody neutralizes sars-cov-2 by blocking receptor interaction. 2020.
[69] Matz J, Kessler P, Bouchet J, et al. Straightforward selection of broadly neutralizing single-domain antibodies targeting the conserved cd4 and coreceptor binding sites of hiv-1 gp120. Journal of Virology, 2013,87(2):1137-1149.
doi: 10.1128/JVI.00461-12 pmid: 23152508
[70] Koch K, Kalusche S, Torres J L, et al. Selection of nanobodies with broad neutralizing potential against primary hiv-1 strains using soluble subtype c gp140 envelope trimers. Scientific Reports, 2017,7(1):8390.
doi: 10.1038/s41598-017-08273-7 pmid: 28827559
[71] Weiss R A, Verrips C T. Nanobodies that neutralize hiv. Vaccines, 2019,7(3):77.
doi: 10.3390/vaccines7030077
[72] Van Hout A, Klarenbeek A, Bobkov V, et al. Cxcr4-targeting nanobodies differentially inhibit cxcr4 function and hiv entry. Biochemical Pharmacology, 2018,158:402-412.
doi: 10.1016/j.bcp.2018.10.015 pmid: 30342024
[73] Boons E, Li G, Vanstreels E, et al. A stably expressed llama single-domain intrabody targeting rev displays broad-spectrum anti-hiv activity. Antiviral Research, 2014,112:91-102.
doi: 10.1016/j.antiviral.2014.10.007 pmid: 25453342
[74] Lutje Hulsik D, Liu Y Y, Strokappe N M, et al. A gp41 mper-specific llama vhh requires a hydrophobic cdr3 for neutralization but not for antigen recognition. PLoS Pathogens, 2013,9(3):e1003202.
doi: 10.1371/journal.ppat.1003202 pmid: 23505368
[75] Daka A, Peer D. Rnai-based nanomedicines for targeted personalized therapy. Advanced Drug Delivery Reviews, 2012,64(13):1508-1521.
doi: 10.1016/j.addr.2012.08.014 pmid: 22975009
[76] Cunha-Santos C, Perdigao P R L, Martin F, et al. Inhibition of hiv replication through sirna carried by cxcr4-targeted chimeric nanobody. Cellular and Molecular Life Sciences, 2019.
doi: 10.1007/s00018-020-03678-6 pmid: 33078208
[77] Li S F, Zhang W, Jiang K P, et al. Nanobody against the e7 oncoprotein of human papillomavirus 16. Molecular Immunology, 2019,109:12-19.
doi: 10.1016/j.molimm.2019.02.022 pmid: 30849663
[78] Woodham A W, Cheloha R W, Ling J J, et al. Nanobody-antigen conjugates elicit hpv-specific antitumor immune responses. Cancer Immunology Research, 2018,6(7):870-880.
doi: 10.1158/2326-6066.CIR-17-0661 pmid: 29792298
[79] De Martel C, Ferlay J, Franceschi S, et al. Global burden of cancers attributable to infections in 2008: a review and synthetic analysis. The Lancet Oncology, 2012,13(6):607-615.
doi: 10.1016/S1470-2045(12)70137-7 pmid: 22575588
[80] Walsh R, Nuttall S, Revill P, et al. Targeting the hepatitis b virus precore antigen with a novel ignar single variable domain intrabody. Virology, 2011,411(1):132-141.
doi: 10.1016/j.virol.2010.12.034 pmid: 21239030
[81] Tarr A W, Lafaye P, Meredith L, et al. An alpaca nanobody inhibits hepatitis c virus entry and cell-to-cell transmission. Hepatology, 2013,58(3):932-939.
doi: 10.1002/hep.26430 pmid: 23553604
[82] Koromyslova A D, Hausman G S. Nanobodies targeting norovirus capsid reveal functional epitopes and potential mechanisms of neutralization. Biophysical Journal, 2018,114(3):219a-219a.
[83] Geoghegan E M, Zhang H, Desai P J, et al. Antiviral activity of a single-domain antibody immunotoxin binding to glycoprotein d of herpes simplex virus 2. Antimicrobial Agents and Chemotherapy, 2015,59(1):527-535.
doi: 10.1128/AAC.03818-14 pmid: 25385102
[84] Conrath K E, Lauwereys M, Galleni M, et al. Beta-lactamase inhibitors derived from single-domain antibody fragments elicited in the camelidae. Antimicrobial Agents and Chemotherapy, 2001,45(10):2807-2812.
doi: 10.1128/AAC.45.10.2807-2812.2001 pmid: 11557473
[85] Barta M L, Shearer J P, Arizmendi O, et al. Single-domain antibodies pinpoint potential targets within shigella invasion plasmid antigen d of the needle tip complex for inhibition of type iii secretion. Journal of Biological Chemistry, 2017,292(40):16677-16687.
doi: 10.1074/jbc.M117.802231 pmid: 28842484
[86] King M T, Huh I, Shenai A, et al. Structural basis of vhh-mediated neutralization of the food-borne pathogen listeria monocytogenes. Journal of Biological Chemistry, 2018,293(35):13626-13635.
doi: 10.1074/jbc.RA118.003888 pmid: 29976754
[87] Ebrahimizadeh W, Mousavi Gargari S, Rajabibazl M, et al. Isolation and characterization of protective anti-lps nanobody against v. Cholerae o1 recognizing inaba and ogawa serotypes. Applied Microbiology and Biotechnology, 2013,97(10):4457-4466.
doi: 10.1007/s00253-012-4518-x pmid: 23135228
[88] Baral T N, Magez S, Stijlemans B, et al. Experimental therapy of african trypanosomiasis with a nanobody-conjugated human trypanolytic factor. Nature Medicine, 2006,12(5):580-584.
doi: 10.1038/nm1395 pmid: 16604085
[89] Arias J L Unciti-Broceta J D Maceira J, et al. Nanobody conjugated plga nanoparticles for active targeting of african trypanosomiasis. Journal of Controlled Release, 2015,197:190-198.
doi: 10.1016/j.jconrel.2014.11.002 pmid: 25445702
[90] Unciti-Broceta J D, Arias J L, Maceira J, et al. Specific cell targeting therapy bypasses drug resistance mechanisms in african trypanosomiasis. PLoS Pathogens, 2015,11(6):e1004942.
doi: 10.1371/journal.ppat.1004942 pmid: 26110623
[91] Wu Y, Jiang S, Ying T. Single-domain antibodies as therapeutics against human viral diseases. Frontiers in Immunology, 2017,8:1802.
doi: 10.3389/fimmu.2017.01802 pmid: 29326699
[92] Keyaerts M, Xavier C, Heemskerk J, et al. Phase i study of 68ga-her2-nanobody for pet/ct assessment of her2 expression in breast carcinoma. Journal of Nuclear Medicine, 2016,57(1):27-33.
doi: 10.2967/jnumed.115.162024 pmid: 26449837
[93] Lemke J, Von Karstedt S, Zinngrebe J, et al. Getting trail back on track for cancer therapy. Cell Death and Differentiation, 2014,21(9):1350-1364.
doi: 10.1038/cdd.2014.81 pmid: 24948009
[94] Wu Y, Li C, Xia S, et al. Identification of human single-domain antibodies against sars-cov-2. Cell Host & Microbe, 2020, 27(6): 891-898.e5.
doi: 10.1016/j.chom.2020.04.023 pmid: 32413276
[95] Dolk E, Van Der Vaart M, Lutje Hulsik D, et al. Isolation of llama antibody fragments for prevention of dandruff by phage display in shampoo. Applied and Environmental Microbiology, 2005,71(1):442-450.
doi: 10.1128/AEM.71.1.442-450.2005 pmid: 15640220
[96] 2018新型抗体药物论坛. 传遍了朋友圈的羊驼-纳米抗体平台究竟是什么.[ 2018-10-26]. http://meeting.bioon.com/2018NA/news-detail/c8098462ec0323e6.
[1] 赵晓煜,徐祺玲,赵晓东,安云飞. 基因治疗慢病毒载体的转导增强策略*[J]. 中国生物工程杂志, 2021, 41(8): 52-58.
[2] 陈晨,胡劲超,曹姗姗,门冬. 新型冠状病毒抗原快速检测研发现状及展望*[J]. 中国生物工程杂志, 2021, 41(6): 119-128.
[3] 马巧妮,王萌,朱兴全. 重组酶介导扩增技术及其在病原微生物快速检测中的应用进展*[J]. 中国生物工程杂志, 2021, 41(6): 45-49.
[4] 吕慧中,赵晨辰,朱链,许娜. 外泌体靶向递药在肿瘤治疗中的进展[J]. 中国生物工程杂志, 2021, 41(5): 79-86.
[5] 时忠林,崔俊生,杨柯,胡安中,李亚楠,刘勇,邓国庆,朱灿灿,朱灵. 基于微流控芯片的核酸等温扩增技术研究进展 *[J]. 中国生物工程杂志, 2021, 41(2/3): 116-128.
[6] 范月蕾,王跃,王恒哲,李丹丹,毛开云. 新型冠状病毒体外诊断技术研发现状与展望 *[J]. 中国生物工程杂志, 2021, 41(2/3): 150-161.
[7] 原博,王杰文,康广博,黄鹤. 双特异性纳米抗体的研究进展及其应用 *[J]. 中国生物工程杂志, 2021, 41(2/3): 78-88.
[8] 唐德平,邢梦洁,宋文涛,姚慧慧,毛爱红. microRNA治疗在癌症及其他疾病中的研究进展*[J]. 中国生物工程杂志, 2021, 41(11): 64-73.
[9] 吴忧,辛林. 新的药物传递系统:外泌体作为药物载体递送*[J]. 中国生物工程杂志, 2020, 40(9): 28-35.
[10] 蔺士新,刘东晨,雷云,熊盛,谢秋玲. TNF-α纳米抗体的筛选、表达及特异性检测 *[J]. 中国生物工程杂志, 2020, 40(7): 15-21.
[11] 毛开云,赵若春,王跃,范月蕾,江洪波. 全球细胞治疗CMO/CDMO行业发展态势分析 *[J]. 中国生物工程杂志, 2020, 40(6): 106-112.
[12] 李玉,张晓. 日本细胞治疗监管双轨制的经验及启示 *[J]. 中国生物工程杂志, 2020, 40(1-2): 174-179.
[13] 潘彤彤,陈永平. 重型/危重型新型冠状病毒肺炎关键治疗技术研究进展[J]. 中国生物工程杂志, 2020, 40(1-2): 78-83.
[14] 赵萍,杨艳萍. 人冠状病毒感染诊断技术专利态势分析[J]. 中国生物工程杂志, 2020, 40(1-2): 51-56.
[15] 林福玉,刘金毅,程永庆. 重组人干扰素α1b抗新型冠状病毒的基础和临床研究进展[J]. 中国生物工程杂志, 2020, 40(12): 1-7.
主管：中国科学院 　主办：中国科学院文献情报中心 　中国生物技术发展中心 　中国生物工程学会