Zhai W, Davies J, Shang D Z, Chan S W, Allain J P
Division of Transfusion Medicine, Department of Haematology, University of Cambridge, UK.
J Viral Hepat. 1999 Mar;6(2):115-24. doi: 10.1046/j.1365-2893.1999.00146.x.
The hypervariable region 1 (HVR1) of hepatitis C virus (HCV) may contain a potential neutralization site and the generation of human single-chain antibody fragments (scFv) to HVR1 may therefore provide a useful tool for the study of HCV. In this report, we have isolated and characterized three anti-HVR1 scFv clones from two patient-derived phage-displayed libraries and HCV HVR1 peptides. scFv S52/20 and S53/6 were selected with serologically cross-reactive HVR1 peptides. scFv p3f10 was obtained by screening the library from patient MH with an autologous HVR1 peptide. Nucleotide sequencing showed that the VH chains and Vkappa chains of all three scFv antibodies were derived from VH3 and Vkappa1 family germline V-genes, respectively. The specificity and affinity of the recombinant scFv antibodies were examined by enzyme-linked immunosorbent assay (ELISA) and an affinity biosensor, using HVR1 peptides. S52/20 scFv binding to S52 HVR1 peptide was blocked by preincubation with soluble peptide S52 and was partially competed by one of three HCV-infected patient sera. In addition, scFv S52/20 blocked the binding of HCV-susceptible Molt-4 cells to immobilized S52 peptide. This study demonstrates that recombinant human scFv antibodies to HCV HVR1 can be produced in vitro and directly confirms that HVR1 of HCV elicits highly specific antibodies. The very high specificity of these antibodies to HVR1 may limit their potential use in passive immunization therapy against HCV, and further engineering of the scFvs needs to be performed to generate broad-spectrum blocking scFvs.
丙型肝炎病毒（HCV）的高变区1（HVR1）可能包含一个潜在的中和位点，因此产生针对HVR1的人单链抗体片段（scFv）可能为HCV的研究提供一个有用的工具。在本报告中，我们从两个患者来源的噬菌体展示文库和HCV HVR1肽中分离并鉴定了三个抗HVR1 scFv克隆。scFv S52/20和S53/6是用血清学交叉反应性HVR1肽筛选出来的。scFv p3f10是通过用患者MH的自体HVR1肽筛选文库获得的。核苷酸测序表明，所有三个scFv抗体的VH链和Vκ链分别来源于VH3和Vκ1家族种系V基因。使用HVR1肽，通过酶联免疫吸附测定（ELISA）和亲和生物传感器检测重组scFv抗体的特异性和亲和力。S52/20 scFv与S52 HVR1肽的结合可被可溶性肽S52预孵育阻断，并被三份HCV感染患者血清中的一份部分竞争。此外，scFv S52/20阻断了HCV易感的Molt-4细胞与固定化S52肽的结合。本研究表明，针对HCV HVR1的重组人scFv抗体可以在体外产生，并直接证实HCV的HVR1能引发高度特异性抗体。这些抗体对HVR1的极高特异性可能会限制它们在针对HCV的被动免疫治疗中的潜在用途，需要对scFv进行进一步工程改造以产生广谱阻断性scFv。
