Università Vita-Salute San Raffaele, Milano, Italy.
J Antimicrob Chemother. 2010 Mar;65(3):425-33. doi: 10.1093/jac/dkp477. Epub 2010 Jan 7.
To understand the dynamic viral evolution observed during failure on raltegravir-containing regimens, we studied the genotypic and phenotypic patterns of resistance to raltegravir and the residual replication capacity (rRC) of HIV-1 variants selected in vivo.
Clonal genotypic analyses were performed on sequential HIV-1 integrase sequences amplified from 11 failing patients and sampled every 4-24 weeks for up to 64 weeks. Fully replicating recombinant viruses were generated using modified vectors in which selected viral integrase genes amplified from patients' plasma were cloned. rRC was measured by a novel multiple cycle competition assay. Resistance to raltegravir and the rRC of resistant HIV-1 variants selected in vivo were evaluated in purified CD4+ T cells.
In all of the patients but one, failure was associated with selection of mutations in positions 143, 148 or 155. Unlike mutations at position 143 (Y143S/K/R), identified alone or in combination with others, mutations at position 148 and 155 were always found in combination. A wide range of resistance levels to raltegravir [from 10- to 770-fold change in 50% inhibitory concentration (IC(50)) compared with baseline] was observed using recombinant viral clones. Finally, rRC was not significantly altered in highly resistant variants.
Two patterns of viral evolution were observed in the resistant viral populations, driving the variants towards a fast (most of them with G140S + Q148H mutations) or progressive increase in resistance to raltegravir. These results may have implications either for the evaluation of genotypic results, or for the correct clinical use of the compound.
为了了解拉替拉韦治疗失败时观察到的病毒动态进化，我们研究了耐药基因型和表型模式以及体内选择的 HIV-1 变异体的残余复制能力(rRC)。
对 11 例治疗失败患者的 HIV-1 整合酶序列进行连续的克隆基因型分析，每 4-24 周采集一次，最长达 64 周。采用改良载体生成完全复制的重组病毒，其中从患者血浆中扩增的选定病毒整合酶基因克隆到载体中。采用新的多循环竞争测定法测量 rRC。在纯化的 CD4+T 细胞中评估了耐药 HIV-1 变异体对拉替拉韦的耐药性和体内选择的 rRC。
在所有患者中，除了 1 例患者，失败都与位置 143、148 或 155 处的突变选择有关。与位置 143(Y143S/K/R)处的突变不同，单独或与其他突变组合发现的位置 148 和 155 处的突变总是组合出现。使用重组病毒克隆，观察到拉替拉韦的耐药水平有很大差异[与基线相比，50%抑制浓度(IC(50))的 10-770 倍变化]。最后，高耐药变异体的 rRC 没有明显改变。
在耐药病毒群体中观察到两种病毒进化模式，使变异体快速(大多数为 G140S + Q148H 突变)或逐渐增加对拉替拉韦的耐药性。这些结果可能对基因型结果的评估或该化合物的正确临床应用有影响。
