Impact of maraviroc-resistant and low-CCR5- adapted mutations induced by in vitro passage on sensitivity to anti-envelope neutralizing antibodies.
Impact of maraviroc-resistant and low-CCR5- adapted mutations induced by in vitro passage on sensitivity to anti-envelope neutralizing antibodies.
复制标题
体外传代诱导的马拉韦罗抗性和低 CCR5 适应突变对抗包膜中和抗体敏感性的影响。
DOI：
10.1099/vir.0.062885-0
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发表时间：
2014
影响因子：
3.8
通讯作者：
Shuzo Matsushita.
中科院分区：
文献类型：
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作者：
Kazuhisa Yoshimura;Shigeyoshi Harada;Samatchaya Boonchawalit;Yoko Kawanami;Shuzo Matsushita.
本研究的目的是使用人类免疫缺陷病毒1型B亚型临床分离株（HIV-1 KP-5）体外产生马拉韦罗（MVC）耐药病毒，以了解对MVC耐药的机制。为了在体外筛选对MVC具有抗性的HIV-1变异体，我们将高表达趋化因子（C-C基序）受体5（CCR 5）的PM 1/CCR 5细胞暴露于HIV-1 KP-5，然后在MVC存在下连续传代。我们还在低CCR 5表达的PM 1细胞中传代HIV-1 KP-5，以确定低CCR 5适应的取代，并比较MVC选择的变体的Env序列。用MVC（10 µM）传代48次后，HIV-1 KP-5获得耐药表型[最大抑制百分比（MPI）24%]，而低CCR 5适应变体对MVC的敏感性较低（IC 50 ~200 nM），但MPI未降低。 在MVC选择的和低CCR 5适应的变体中观察到的常见取代选自V1、V3和V5中的准种。在14次传代后，MVC选择的变体在CCR 5 N-末端结合位点和V3（V200 I、T297 I、K305 R和M434 I）周围具有取代。低CCR 5适应的感染性克隆变得对抗CD 4 bs和CD 4 i mAb敏感，但对抗V3 mAb和自体血浆IgG不敏感。相反，MVC选择的克隆变得对测试的抗包膜（Env）mAb和自体血浆IgG高度敏感。这些发现表明，使用MVC结合的CCR 5进入所需的四个MVC抗性突变导致Env的构象变化，其与对抗Env中和抗体敏感的表型相关。
The aim of this study was to generate maraviroc (MVC)-resistant virusesin vitrousing a human immunodeficiency virus type 1 subtype B clinical isolate (HIV-1KP-5) to understand the mechanism(s) of resistance to MVC. To select HIV-1 variants resistant to MVCin vitro, we exposed high-chemokine (C-C motif) receptor 5 (CCR5)-expressing PM1/CCR5 cells to HIV-1KP-5followed by serial passage in the presence of MVC. We also passaged HIV-1KP-5in PM1 cells, which were low CCR5 expressing to determine low-CCR5-adapted substitutions and compared the Env sequences of the MVC-selected variants. Following 48 passages with MVC (10 µM), HIV-1KP-5acquired a resistant phenotype [maximal per cent inhibition (MPI) 24 %], whilst the low-CCR5-adapted variant had low sensitivity to MVC (IC50~200 nM), but not reduction of the MPI. The common substitutions observed in both the MVC-selected and low-CCR5-adapted variants were selected from the quasi-species, in V1, V3 and V5. After 14 passages, the MVC-selected variants harboured substitutions around the CCR5 N-terminal-binding site and V3 (V200I, T297I, K305R and M434I). The low-CCR5-adapted infectious clone became sensitive to anti-CD4bs and CD4i mAbs, but not to anti-V3 mAb and autologous plasma IgGs. Conversely, the MVC-selected clone became highly sensitive to the anti-envelope (Env) mAbs tested and the autologous plasma IgGs. These findings suggest that the four MVC-resistant mutations required for entry using MVC-bound CCR5 result in a conformational change of Env that is associated with a phenotype sensitive to anti-Env neutralizing antibodies.