Novel single-cell-level phenotypic assay for residual drug susceptibility and reduced replication capacity of drug-resistant human immunodeficiency virus type 1.
Human immunodeficiency virus type 1 (HIV-1)-infected individuals who develop drug-resistant virus during antiretroviral therapy may derive benefit from continued treatment for two reasons. First, drug-resistant viruses can retain partial susceptibility to the drug combination. Second, therapy selects for drug-resistant viruses that may have reduced replication capacities relative to archived, drug-sensitive viruses. We developed a novel single-cell-level phenotypic assay that allows these two effects to be distinguished and compared quantitatively. Patient-derived gag-pol sequences were cloned into an HIV-1 reporter virus that expresses an endoplasmic reticulum-retained Env-green fluorescent protein fusion. Flow cytometric analysis of single-round infections allowed a quantitative analysis of viral replication over a 4-log dynamic range. The assay faithfully reproduced known in vivo drug interactions occurring at the level of target cells. Simultaneous analysis of single-round infections by wild-type and resistant viruses in the presence and absence of the relevant drug combination divided the benefit of continued nonsuppressive treatment into two additive components, residual virus susceptibility to the drug combination and selection for drug-resistant variants with diminished replication capacities. In some patients with drug resistance, the dominant circulating viruses retained significant susceptibility to the combination. However, in other cases, the dominant drug-resistant viruses showed no residual susceptibility to the combination but had a reduced replication capacity relative to the wild-type virus. In this case, simplification of the regimen might still allow adequate suppression of the wild-type virus. In a third pattern, the resistant viruses had no residual susceptibility to the relevant drug regimen but nevertheless had a replication capacity equivalent to that of wild-type virus. In such cases, there is no benefit to continued treatment. Thus, the ability to simultaneously analyze residual susceptibility and reduced replication capacity of drug-resistant viruses may provide a basis for rational therapeutic decisions in the setting of treatment failure.
Zhang
Haili
H
Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Zhou
Yan
Y
Alcock
Cecily
C
Kiefer
Tara
T
Monie
Daphne
D
Siliciano
Janet
J
Li
Quan
Q
Pham
Paul
P
Cofrancesco
Joseph
J
Persaud
Deborah
D
Siliciano
Robert F
RF
D000328:Adult; D023241:Antiretroviral Therapy, Highly Active; D002648:Child; D002675:Child, Preschool; D024882:Drug Resistance, Viral; D005434:Flow Cytometry; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D015658:HIV Infections / Q000821:virology; D016333:HIV Protease / Q000235:genetics; D054303:HIV Reverse Transcriptase / Q000235:genetics; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D006801:Humans; D019169:Jurkat Cells; D008164:Luminescent Proteins / Q000235:genetics; D008826:Microbial Sensitivity Tests / Q000379:methods; D010641:Phenotype; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014779:Virus Replication
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D054303:HIV Reverse Transcriptase; D016333:HIV Protease