Institut de Recerca de la SIDA irsiCaixa - HIVACAT, Hospital Universitari Germans Trias i Pujol, Universitat Autònoma de Barcelona, Catalonia, Spain.
PLoS One. 2013 Aug 1;8(8):e67085. doi: 10.1371/journal.pone.0067085. Print 2013.
Technically, HIV-1 tropism can be evaluated in plasma or peripheral blood mononuclear cells (PBMCs). However, only tropism testing of plasma HIV-1 has been validated as a tool to predict virological response to CCR5 antagonists in clinical trials. The preferable tropism testing strategy in subjects with undetectable HIV-1 viremia, in whom plasma tropism testing is not feasible, remains uncertain.
METHODS & RESULTS: We designed a proof-of-concept study including 30 chronically HIV-1-infected individuals who achieved HIV-1 RNA <50 copies/mL during at least 2 years after first-line ART initiation. First, we determined the diagnostic accuracy of 454 and population sequencing of gp120 V3-loops in plasma and PBMCs, as well as of MT-2 assays before ART initiation. The Enhanced Sensitivity Trofile Assay (ESTA) was used as the technical reference standard. 454 sequencing of plasma viruses provided the highest agreement with ESTA. The accuracy of 454 sequencing decreased in PBMCs due to reduced specificity. Population sequencing in plasma and PBMCs was slightly less accurate than plasma 454 sequencing, being less sensitive but more specific. MT-2 assays had low sensitivity but 100% specificity. Then, we used optimized 454 sequence data to investigate viral evolution in PBMCs during viremia suppression and only found evolution of R5 viruses in one subject. No de novo CXCR4-using HIV-1 production was observed over time. Finally, Slatkin-Maddison tests suggested that plasma and cell-associated V3 forms were sometimes compartmentalized.
The absence of tropism shifts during viremia suppression suggests that, when available, testing of stored plasma samples is generally safe and informative, provided that HIV-1 suppression is maintained. Tropism testing in PBMCs may not necessarily produce equivalent biological results to plasma, because the structure of viral populations and the diagnostic performance of tropism assays may sometimes vary between compartments. Thereby, proviral DNA tropism testing should be specifically validated in clinical trials before it can be applied to routine clinical decision-making.
从技术上讲，HIV-1 嗜性可在血浆或外周血单核细胞（PBMC）中进行评估。然而，只有血浆 HIV-1 嗜性检测已被验证可作为预测临床试验中 CCR5 拮抗剂病毒学应答的工具。在血浆 HIV-1 无法检测到的 HIV-1 血症患者中，其优选的嗜性检测策略仍不确定，这些患者的病毒血症无法检测。
我们设计了一项概念验证研究，包括 30 名慢性 HIV-1 感染者，他们在首次接受一线 ART 治疗后至少 2 年内 HIV-1 RNA<50 拷贝/mL。首先，我们确定了 454 测序和群体测序在血浆和 PBMC 中以及在 ART 前的 MT-2 检测的准确性，该方法作为技术参考标准。增强敏感性 Trofile 检测（ESTA）用于作为参考标准。与 ESTA 相比，血浆病毒的 454 测序具有最高的一致性。由于特异性降低，PBMC 中的 454 测序准确性降低。血浆和 PBMC 中的群体测序准确性略低于血浆 454 测序，其敏感性较低，但特异性较高。MT-2 检测的敏感性较低，但特异性为 100%。然后，我们使用优化的 454 序列数据来研究病毒血症抑制期间 PBMC 中的病毒进化，仅在一名患者中发现 R5 病毒的进化。随着时间的推移，未观察到新的 CXCR4 使用 HIV-1 的产生。最后，Slatkin-Maddison 检验表明，血浆和细胞相关的 V3 形式有时是分隔的。
在病毒血症抑制期间没有发生嗜性转变，这表明在病毒血症得到抑制的情况下，检测储存的血浆样本通常是安全且有信息价值的，前提是 HIV-1 得到抑制。PBMC 中的嗜性检测不一定能产生与血浆等同的生物学结果，因为病毒群体的结构和嗜性检测的诊断性能有时会在不同的部位之间发生变化。因此，在将前病毒 DNA 嗜性检测应用于常规临床决策之前，应在临床试验中对其进行专门验证。
