Barnas Douglas, Koontz Dianna, Bazmi Holly, Bixby Christian, Jemsek Joseph, Mellors John W
Department of Medicine, Division Infectious Diseases, University of Pittsburgh, Pittsburgh, PA, USA.
Antivir Ther. 2010;15(3):437-41. doi: 10.3851/IMP1539.
The rapid failure of initial therapy with combinations of nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) that exclude zidovudine has not been fully explained by standard virus population analyses of HIV type-1 (HIV-1) drug resistance. We therefore investigated HIV-1 genotype and phenotype at the single genome level in samples from patients on a failing regimen of tenofovir (TNV), didanosine (ddI) and lamivudine (3TC).
Single genome sequencing was performed on 9 failure samples containing both K65R and M184V mutations by standard genotype, either as wild-type/mutant mixtures (6/9) or as mutant only (3/9). Recombinant clones with different combinations of observed mutations were generated and tested for NRTI susceptibility.
Of the 204 single genome sequences analysed, 50% were K65R/M184V double mutants, 38% were M184V single mutants, 10% were M184I single mutants and only 1% (2 sequences) were K65R single mutants. Phenotypic testing of recombinant clones showed a significant increase in resistance for double mutants: mean fold resistance to abacavir, ddI and TNV was 6.5, 4.3 and 1.6 for K65R/M184V double mutants versus 2.5, 1.9 and 0.6 for M184V single mutants, respectively (P<0.001).
Mutants with K65R and M184V linked on the same genome were the most common HIV-1 variants in samples analysed from patients failing TNV, ddI and 3TC with both mutations detected by standard genotype. The double mutant exhibited reduced susceptibility to all three NRTIs in the regimen. This resistant phenotype, resulting from just two linked point mutations, likely contributes to rapid failure of NRTI combinations that exclude zidovudine.
不包含齐多夫定的核苷/核苷酸逆转录酶抑制剂（NRTIs）联合初始治疗的快速失败，尚未通过1型人类免疫缺陷病毒（HIV-1）耐药性的标准病毒群体分析得到充分解释。因此，我们在接受替诺福韦（TNV）、去羟肌苷（ddI）和拉米夫定（3TC）联合治疗方案但治疗失败的患者样本中，在单基因组水平上研究了HIV-1基因型和表型。
对9个同时含有K65R和M184V突变的治疗失败样本进行单基因组测序，采用标准基因型，其中6个样本为野生型/突变体混合物，3个样本仅为突变体。构建含有观察到的不同突变组合的重组克隆，并检测其对NRTIs的敏感性。
在分析的204个单基因组序列中，50%为K65R/M184V双突变体，38%为M184V单突变体，10%为M184I单突变体，只有1%（2个序列）为K65R单突变体。重组克隆的表型检测显示双突变体的耐药性显著增加：K65R/M184V双突变体对阿巴卡韦、ddI和TNV的平均耐药倍数分别为6.5、4.3和1.6，而M184V单突变体分别为2.5、1.9和0.6（P<0.001）。
在接受TNV、ddI和3TC治疗失败且通过标准基因型检测到两种突变的患者样本中，同一基因组上同时存在K65R和M184V的突变体是最常见的HIV-1变体。双突变体对该治疗方案中的所有三种NRTIs的敏感性均降低。仅由两个连锁点突变导致的这种耐药表型，可能是不包含齐多夫定的NRTIs联合治疗快速失败的原因。
