Deep V3 Sequencing for HIV Type 1 Tropism in Treatment-Naive Patients: A Reanalysis of the MERIT Trial of Maraviroc

Swenson, Luke C.; Mo, Theresa; Dong, Winnie; Zhong, Xiaoyin; Woods, Conan K.; Thielen, Alexander; Jensen, Mark A.; Knapp, David J.H.F.; Chapman, Douglass; Portsmouth, Simon; Lewis, Marilyn; James, Ian; Heera, Jayvant; Valdez, Hernán; Harrigan, P. Richard

doi:10.1093/cid/cir493

Cited by 86 publications

(92 citation statements)

References 21 publications

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“…Of the various CCR5 antagonists, only three reached advanced stages of clinical trials and only maraviroc was approved first for salvage-based treatment and then for first-line treatment in combination with nucleoside analogs (51,52). For the most part, virologic failures with MVC-based treatment regimens have been poorly characterized and generally complicated by the preexistence of multidrug-resistant genotypes in these heavily treated patients (16,53,54). When MVC is used as a salvage-based treatment regimen, resistance commonly relates to the rapid selection of preexisting X4-using HIV-1 in the intrapatient HIV-1 population (16).…”

Section: Discussionmentioning

confidence: 99%

HIV-1 Resistance to Maraviroc Conferred by a CD4 Binding Site Mutation in the Envelope Glycoprotein gp120

Ratcliff

Shi

Arts

2013

J Virol

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Maraviroc (MVC) is a CCR5 antagonist that inhibits HIV-1 entry by binding to the coreceptor and inducing structural alterations in the extracellular loops. In this study, we isolated MVC-resistant variants from an HIV-1 primary isolate that arose after 21 weeks of tissue culture passage in the presence of inhibitor. gp120 sequences from passage control and MVC-resistant cultures were cloned into NL4-3 via yeast-based recombination followed by sequencing and drug susceptibility testing. Using 140 clones, three mutations were linked to MVC resistance, but none appeared in the V3 loop as was the case with previous HIV-1 strains resistant to CCR5 antagonists. Rather, resistance was dependent upon a single mutation in the C4 region of gp120. Chimeric clones bearing this N425K mutation replicated at high MVC concentrations and displayed significant shifts in 50% inhibitory concentrations (IC 50 s), characteristic of resistance to all other antiretroviral drugs but not typical of MVC resistance. Previous reports on MVC resistance describe an ability to use a drug-bound form of the receptor, leading to reduction in maximal drug inhibition. In contrast, our structural models on K425 gp120 suggest that this resistant mutation impacts CD4 interactions and highlights a novel pathway for MVC resistance.

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Section: Discussionmentioning

confidence: 99%

HIV-1 Resistance to Maraviroc Conferred by a CD4 Binding Site Mutation in the Envelope Glycoprotein gp120

Ratcliff

Shi

Arts

2013

J Virol

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“…The position-specific scoring matrix (PSSM) analyzes the entire V3 sequence and predicts X4 variants based on a scoring of the probability of the amino acid at each position being overrepresented among X4 sequences versus R5 sequences (10). The Geno2Pheno (G2P) algorithm (11) also analyzes the entire V3 sequence and provides a quantitative score (false-positive rate [FPR]) representing the probability of falsely predicting an R5 variant as an X4 variant; the validity of this approach has been assessed in several clinical trials (12)(13)(14)(15)(16). G2P has been widely used in research and clinical settings, but it requires a preset FPR cutoff to call X4 variants.…”

mentioning

confidence: 99%

Deep Sequencing of the HIV-1 env Gene Reveals Discrete X4 Lineages and Linkage Disequilibrium between X4 and R5 Viruses in the V1/V2 and V3 Variable Regions

Zhou

Bednar

Sturdevant

et al. 2016

J Virol

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HIV-1 requires the CD4 receptor and a coreceptor (CCR5 [R5 phenotype] or CXCR4 [X4 phenotype]) to enter cells. Coreceptor tropism can be assessed by either phenotypic or genotypic analysis, the latter using bioinformatics algorithms to predict tropism based on the env V3 sequence. We used the Primer ID sequencing strategy with the MiSeq sequencing platform to reveal the structure of viral populations in the V1/V2 and C2/V3 regions of the HIV-1 env gene in 30 late-stage and 6 early-stage subjects. We also used endpoint dilution PCR followed by cloning of env genes to create pseudotyped virus to explore the link between genotypic predictions and phenotypic assessment of coreceptor usage. We found out that the most stringently sequence-based calls of X4 variants (Geno2Pheno false-positive rate [FPR] of <2%) formed distinct lineages within the viral population, and these were detected in 24 of 30 late-stage samples (80%), which was significantly higher than what has been seen previously by using other approaches. Non-X4 lineages were not skewed toward lower FPR scores in X4-containing populations. Phenotypic assays showed that variants with an intermediate FPR (2 to 20%) could be either X4/dual-tropic or R5 variants, although the X4 variants made up only about 25% of the lineages with an FPR of <10%, and these variants carried a distinctive sequence change. Phylogenetic analysis of both the V1/V2 and C2/V3 regions showed evidence of recombination within but very little recombination between the X4 and R5 lineages, suggesting that these populations are genetically isolated. IMPORTANCEPrimer ID sequencing provides a novel approach to study genetic structures of viral populations. X4 variants may be more prevalent than previously reported when assessed by using next-generation sequencing (NGS) and with a greater depth of sampling than single-genome amplification (SGA). Phylogenetic analysis to identify lineages of sequences with intermediate FPR values may provide additional information for accurately predicting X4 variants by using V3 sequences. Limited recombination occurs between X4 and R5 lineages, suggesting that X4 and R5 variants are genetically isolated and may be replicating in different cell types or that X4/R5 recombinants have reduced fitness. H uman immunodeficiency virus type 1 (HIV-1) requires the CD4 receptor and a coreceptor to infect host cells. Entry is mediated by the viral envelope protein (Env), which is processed to give two associated subunits, gp120 and gp41, that assemble into a trimeric structure embedded in the viral envelope/membrane. The binding of gp120 to CD4 triggers a structural change that exposes its variable region 3 (V3) loop, which is part of the coreceptor domain (1, 2). The majority of transmitted/founder (T/F) viruses and viruses isolated from the clinically latent stage in HIV-infected subjects use CCR5 as the coreceptor, making the virus CCR5 tropic (or an R5 virus). The virus can evolve to use CXCR4 (CXCR4 tropic [or an X4 virus]), and viruses that have switched core...

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“…Processing of the clinical specimens and generation of the HIV envelope PCR amplicons were previously described. 8 Prior to CS-PCR, the first-round PCR products were quantified using a double-stranded DNA quantification reagent (Quant-It PicoGreen; Invitrogen) and normalized to *3 · 10 8 amplicons per ll. One microliter of the normalized template was amplified by real-time CS-PCR, in duplicate, as described above.…”

Section: Figmentioning

confidence: 99%

“…Next-generation sequencing may prove to be the best method of detection, as it is not only sensitive, but can also give a fairly accurate estimation of the percent X4 in a population. 7,8 However, next-generation sequencing is still fairly expensive and handling the data can be onerous. Phenotyping is another option for X4 detection, but many laboratories do not have the facilities or expertise to develop a sensitive culture-based assay, and commercial assays, such as Trofile, are costly.…”

Section: Introductionmentioning

confidence: 99%

Development of a Novel Codon-Specific Polymerase Chain Reaction for the Detection of CXCR4-Utilizing HIV Type 1 Subtype B

McLaughlin¹,

Swenson

et al. 2013

AIDS Research and Human Retroviruses

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Insight concerning the switch in HIV-1 coreceptor use will lead to a better understanding of HIV-1 pathogenesis and host-virus dynamics. Predicting CXCR4 utilization by analyzing HIV-1 envelope consensus sequences is highly specific, but minority variants in the viral population are often missed resulting in low sensitivity. Commercial phenotypic assays are costly, and the development of sensitive in-house phenotypic assays to detect CXCR4-using HIV may not be feasible for some laboratories. A sensitive, inexpensive genotyping assay was developed to detect viral sequences associated with CXCR4-utilizing virus (X4). Codon-specific primer pairs were used to detect X4-associated codons at five positions in the HIV-1 envelope V3 loop (11, 13, 24, 25, and 32). Sixty plasma samples from HIV-1-infected individuals were analyzed by consensus sequencing and codonspecific PCR (CS-PCR). Forty-six of these were also phenotyped by Trofile or Enhanced Sensitivity Trofile (ESTA). CS-PCR detected X4 variants 17% more often than 11/24/25 consensus sequencing alone (n = 60), 30% more often than Trofile (n = 27), and in a limited data set, 16% more often than ESTA (n = 19). CS-PCR combined with consensus sequencing had approximately 80% concordance with ESTA.

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Deep V3 Sequencing for HIV Type 1 Tropism in Treatment-Naive Patients: A Reanalysis of the MERIT Trial of Maraviroc

Abstract: Reanalysis of the MERIT trial using deep V3 loop sequencing indicates that, had patients originally been screened using this method, the maraviroc arm would have likely been found to be noninferior to the efavirenz arm.

Cited by 86 publications

References 21 publications

HIV-1 Resistance to Maraviroc Conferred by a CD4 Binding Site Mutation in the Envelope Glycoprotein gp120

HIV-1 Resistance to Maraviroc Conferred by a CD4 Binding Site Mutation in the Envelope Glycoprotein gp120

Deep Sequencing of the HIV-1 env Gene Reveals Discrete X4 Lineages and Linkage Disequilibrium between X4 and R5 Viruses in the V1/V2 and V3 Variable Regions

Development of a Novel Codon-Specific Polymerase Chain Reaction for the Detection of CXCR4-Utilizing HIV Type 1 Subtype B

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