Department of Experimental Medicine and Surgery, University of Rome Tor Vergata, Rome, Italy.
PLoS One. 2013 Jul 9;8(7):e68076. doi: 10.1371/journal.pone.0068076. Print 2013.
Dual/mixed-tropic HIV-1 strains are predominant in a significant proportion of patients, though little information is available regarding their replication-capacity and susceptibility against CCR5-antagonists in-vitro. The aim of the study was to analyze the replication-capacity and susceptibility to maraviroc of HIV-1 clinical isolates with different tropism characteristics in primary monocyte-derived-macrophages (MDM), peripheral-blood-mononuclear-cells (PBMC), and CD4(+) T-lymphocytes.
Twenty-three HIV-1 isolates were phenotipically and genotipically characterized as R5, X4 or dual (discriminated as R5(+)/X4, R5/X4, R5/X4(+)). Phenotypic-tropism was evaluated by multiple-cycles-assay on U87MG-CD4(+)-CCR5(+)-/CXCR4(+)-expressing cells. Genotypic-tropism prediction was obtained using Geno2Pheno-algorithm (false-positive-rate [FPR] = 10%). Replication-capacity and susceptibility to maraviroc were investigated in human-primary MDM, PBMC and CD4(+) T-cells. AMD3100 was used as CXCR4-inhibitor. Infectivity of R5/Dual/X4-viruses in presence/absence of maraviroc was assessed also by total HIV-DNA, quantified by real-time polymerase-chain-reaction.
Among 23 HIV-1 clinical isolates, phenotypic-tropism-assay distinguished 4, 17 and 2 viruses with R5-tropic, dual/mixed-, and X4-tropic characteristics, respectively. Overall, viruses defined as R5(+)/X4-tropic were found with the highest prevalence (10/23, 43.5%). The majority of isolates efficiently replicated in both PBMC and CD4(+) T-cells, regardless of their tropism, while MDM mainly sustained replication of R5- or R5(+)/X4-tropic isolates; strong correlation between viral-replication and genotypic-FPR-values was observed in MDM (rho = 0.710;p-value = 1.4e-4). In all primary cells, maraviroc inhibited viral-replication of isolates not only with pure R5- but also with dual/mixed tropism (mainly R5(+)/X4 and, to a lesser extent R5/X4 and R5/X4(+)). Finally, no main differences by comparing the total HIV-DNA with the p24-production in presence/absence of maraviroc were found.
Maraviroc is effective in-vitro against viruses with dual-characteristics in both MDM and lymphocytes, despite the potential X4-mediated escape. This suggests that the concept of HIV-entry through one of the two coreceptors "separately" may require revision, and that the use of CCR5-antagonists in patients with dual/mixed-tropic viruses may be a therapeutic-option that deserves further investigations in different clinical settings.
双重/混合嗜性 HIV-1 株在很大一部分患者中占优势，尽管有关其在体外对 CCR5 拮抗剂的复制能力和敏感性的信息很少。本研究的目的是分析不同嗜性特征的 HIV-1 临床分离株在原代单核细胞衍生的巨噬细胞（MDM）、外周血单核细胞（PBMC）和 CD4+T 淋巴细胞中的复制能力和对马拉维若的敏感性。
23 株 HIV-1 分离株通过 U87MG-CD4+CCR5+/CXCR4+表达细胞的多轮检测进行表型和基因型特征分析，确定其为 R5、X4 或双重（区分 R5(+)/X4、R5/X4、R5/X4(+)）。使用 Geno2Pheno 算法（假阳性率 [FPR] = 10%）预测基因型嗜性。在人原代 MDM、PBMC 和 CD4+T 细胞中研究复制能力和对马拉维若的敏感性。使用 AMD3100 作为 CXCR4 抑制剂。还通过实时聚合酶链反应定量检测 HIV-DNA 评估 R5/Dual/X4 病毒在存在/不存在马拉维若时的感染性。
在 23 株 HIV-1 临床分离株中，表型嗜性检测将 4、17 和 2 株病毒分别鉴定为 R5 嗜性、双重/混合嗜性和 X4 嗜性。总体而言，具有 R5(+)/X4 嗜性的病毒最常见（10/23，43.5%）。大多数分离株无论其嗜性如何，均能在 PBMC 和 CD4+T 细胞中有效复制，而 MDM 主要维持 R5 或 R5(+)/X4 嗜性分离株的复制；在 MDM 中观察到病毒复制与基因型 FPR 值之间存在很强的相关性（rho = 0.710;p 值 = 1.4e-4）。在所有原代细胞中，马拉维若不仅抑制了具有纯 R5-，而且还抑制了双重/混合嗜性（主要是 R5(+)/X4，其次是 R5/X4 和 R5/X4(+)）的分离株的病毒复制。最后，在存在/不存在马拉维若的情况下，通过比较总 HIV-DNA 与 p24 产生，未发现主要差异。
马拉维若在 MDM 和淋巴细胞中对具有双重特征的病毒具有体外疗效，尽管存在潜在的 X4 介导的逃逸。这表明，通过两种核心受体中的一种“分别”进入 HIV 的概念可能需要修订，并且在具有双重/混合嗜性病毒的患者中使用 CCR5 拮抗剂可能是一种值得在不同临床环境中进一步研究的治疗选择。
