HIV-1广谱中和活性感染者假病毒的中和表型分析

目的分析HIV-1广谱中和活性感染者基于膜蛋白基因(env)的假病毒对自体血浆和代表性广谱单克隆中和抗体(bm NAbs)的中和表型。方法单基因组扩增(SGA)广谱中和活性感染者第0月血浆的env基因并克隆至pc D-NA~(TM)3.1 Directional TOPO表达载体,将env表达质粒与HIV-1骨架质粒p SG3△env共转染293T/17细胞制备假病毒,用假病毒分别与自体连续时间点血浆和代表性bm NAbs进行中和实验分析中和表型。结果共获得11株功能性假病毒,假病毒对8个连续时间点自体血浆的中和敏感性呈现先升高(第0~15月),后下降(第16~32月),之后再上升(第33~45月)的波动。所有假病毒对10E8、PGT121和VRC01中和敏感(IC_(50)6μg/m L);各有18.2%(2株)的假病毒对12A21和2G12高度敏感(IC_(50)1μg/m L);所有病毒均对PGT135呈中和抗性。结论假病毒对当前时间点血浆的中和敏感性低,对之后时间点血浆中和敏感性增强,提示感染者体内病毒与中和抗体的动态进化过程。同一时间点假病毒对特定bm-NAbs的敏感性差异明显,提示感染者准种毒株间中和表型的复杂性。

Neutralization phenotype of the Env-pseudoviruses from an HIV-1 infected individual with broadly neutralizing activity

Objective To analyze the neutralization phenotype of Env-pseudoviruses from an HIV-1 infected individual with broadly neutralization activity against autologous plasma samples and representative broadly monoclonal neutralizing antibodies (bmNAbs).Methods Single genome amplification (SGA) of full length env genes and sequencing were performed on plasma sample at zero month time point from an HIV-1 infected individual with broadly neutralization activity.The env genes were then cloned into pcDNATM3.1 Directional TOPO expression vector.The derived env expression plasmids were cotransfected into 293T/17 cells with HIV-1 backbone plasmid pSG3 △ env to prepare pseudoviruses.And the neutralization phenotype of the pseudoviruses were identified through neutralization experiments with autologous plasma samples and representative bmNAbs.Results Totally,11 functional Env-pseudoviruses were constructed.The neutralization sensitivity of the pseudoviruses to autologous plasma samples at 8 consecutive time points showed a fluctuation of first increase (0-15 months),subsequent decline (16-32 months),and then rise (33-45 months).All Env-pseudotyped viruses were sensitive to 10E8,PGT121 and VRC01 (IC50＜6 μg/mL) but were highly resistant to PGT135.18.2％(2) of the viruses were highly sensitive to 12A21 and 2G12 (IC50＜1 μg/mL),respectively.Conclusion The pseudoviruses were less sensitive against the concurrent plasma samples,and the sensitivity increased overtime against later plasma samples,suggesting dynamic evolution of the viruses and the neutralizing antibodies in this individual with broadly neutralization activity.Pseudoviruses,even from the same time point,differed significantly in its sensitivity to specific bmNAbs,suggesting complexity of the neutralization phenotype between the virus quasispecies.