Ita Sergio, Agostinho Mayara R, Sullivan Katherine, Yub Han Seung, Akleh Rana, Johnson Welkin E, Fofana Ismael Ben F
1 Biology Department, Boston College , Chestnut Hill, Massachusetts.
2 Virology Program, Harvard Medical School , Boston, Massachusetts.
AIDS Res Hum Retroviruses. 2017 Aug;33(8):869-879. doi: 10.1089/AID.2016.0247. Epub 2017 Feb 16.
We have constructed a single chain fragment variable (scFv) phage display library from a simian immunodeficiency virus (SIV)-infected rhesus macaque that developed unusually high-titer neutralizing antibody responses against tier-3, neutralization-resistant SIVmac239. The library was screened using trimeric (gp140) and monomeric (gp120) forms of the SIVmac239 envelope (Env) glycoprotein. We also cloned variable-heavy and variable-light (VH-VL) antibody fragments from seven previously described rhesus macaque B-cell lines (BLCLs) that produce SIV gp120-specific monoclonal antibodies (mAbs). Thirty-two gp140-specific mAbs were selected along with 20 gp120-specific ones. gp120-specific mAbs were only from the VH4 family, while gp41-specific mAbs were primarily from VH1, followed by VH4 and VH3. Rhesus macaque BLCL-derived mAbs belonged primarily to the VH4 family of antibodies followed by VH3 and a smaller number of VH1s. A preferential VH combination with Vλ light chain was observed with phage display-selected SIV Env-specific mAbs (gp120 and gp140), but not with BLCL-derived antibodies or the unpanned library. None of the tested antibodies had detectable neutralizing activity against tier-3 SIVmac239. The majority of gp120-specifc mAbs potently neutralized tier-1 SIVmac316 with 50% inhibitory concentration (IC50) values below 1 μg/ml. For gp140-specific antibodies, which were all specific for the gp41-subunit, 2 out of 11 tested neutralized SIVmac316 (IC50 of 7 and 5 μg/ml, respectively). These data suggest an order of preferential VH segment usage for SIV-specific antibodies in rhesus macaques. These antibodies will be useful in assessing the contribution of non-neutralizing antibodies to inhibition of SIV infection in vitro and in vivo.
我们从一只感染了猴免疫缺陷病毒（SIV）的恒河猴构建了一个单链抗体可变区（scFv）噬菌体展示文库，该恒河猴针对3级中和抗性SIVmac239产生了异常高滴度的中和抗体反应。使用SIVmac239包膜（Env）糖蛋白的三聚体（gp140）和单体（gp120）形式筛选该文库。我们还从七个先前描述的产生SIV gp120特异性单克隆抗体（mAb）的恒河猴B细胞系（BLCL）中克隆了重链可变区和轻链可变区（VH-VL）抗体片段。选择了32个gp140特异性mAb以及20个gp120特异性mAb。gp120特异性mAb仅来自VH4家族，而gp41特异性mAb主要来自VH1，其次是VH4和VH3。恒河猴BLCL衍生的mAb主要属于VH4抗体家族，其次是VH3和少量的VH1。在噬菌体展示选择的SIV Env特异性mAb（gp120和gp140）中观察到VH与Vλ轻链的优先组合，但在BLCL衍生的抗体或未淘选的文库中未观察到。所测试的抗体均未对3级SIVmac239具有可检测的中和活性。大多数gp120特异性mAb能有效中和1级SIVmac316，50%抑制浓度（IC50）值低于1μg/ml。对于均对gp41亚基特异的gp140特异性抗体，11个测试中有2个中和了SIVmac316（IC50分别为7和5μg/ml）。这些数据表明恒河猴中SIV特异性抗体优先使用VH片段的顺序。这些抗体将有助于评估非中和抗体在体外和体内对抑制SIV感染的作用。
