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1.
Nat Commun ; 10(1): 2737, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31227699

RESUMO

Little is known about the genotypic make-up of HIV-1 DNA genomes during the earliest stages of HIV-1 infection. Here, we use near-full-length, single genome next-generation sequencing to longitudinally genotype and quantify subtype C HIV-1 DNA in four women identified during acute HIV-1 infection in Durban, South Africa, through twice-weekly screening of high-risk participants. In contrast to chronically HIV-1-infected patients, we found that at the earliest phases of infection in these four participants, the majority of viral DNA genomes are intact, lack APOBEC-3G/F-associated hypermutations, have limited genome truncations, and over one year show little indication of cytotoxic T cell-driven immune selections. Viral sequence divergence during acute infection is predominantly fueled by single-base substitutions and is limited by treatment initiation during the earliest stages of disease. Our observations provide rare longitudinal insights of HIV-1 DNA sequence profiles during the first year of infection to inform future HIV cure research.


Assuntos
DNA Viral/genética , Evolução Molecular , Genoma Viral/genética , Infecções por HIV/virologia , HIV-1/genética , Doença Aguda , Adulto , Fármacos Anti-HIV/uso terapêutico , Análise Mutacional de DNA , Feminino , Seguimentos , Infecções por HIV/sangue , Infecções por HIV/tratamento farmacológico , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Estudos Longitudinais , Mutação , Estudos Prospectivos , África do Sul , Carga Viral , Adulto Jovem
2.
J Clin Invest ; 129(3): 988-998, 2019 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30688658

RESUMO

Chromosomal integration of genome-intact HIV-1 sequences into the host genome creates a reservoir of virally infected cells that persists throughout life, necessitating indefinite antiretroviral suppression therapy. During effective antiviral treatment, the majority of these proviruses remain transcriptionally silent, but mechanisms responsible for viral latency are insufficiently clear. Here, we used matched integration site and proviral sequencing (MIP-Seq), an experimental approach involving multiple displacement amplification of individual proviral species, followed by near-full-length HIV-1 next-generation sequencing and corresponding chromosomal integration site analysis to selectively map the chromosomal positions of intact and defective proviruses in 3 HIV-1-infected individuals undergoing long-term antiretroviral therapy. Simultaneously, chromatin accessibility and gene expression in autologous CD4+ T cells were analyzed by assays for transposase-accessible chromatin using sequencing (ATAC-Seq) and RNA-Seq. We observed that in comparison to proviruses with defective sequences, intact HIV-1 proviruses were enriched for non-genic chromosomal positions and more frequently showed an opposite orientation relative to host genes. In addition, intact HIV-1 proviruses were preferentially integrated in either relative proximity to or increased distance from active transcriptional start sites and to accessible chromatin regions. These studies strongly suggest selection of intact proviruses with features of deeper viral latency during prolonged antiretroviral therapy, and may be informative for targeting the genome-intact viral reservoir.

3.
J Virol ; 2018 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-30305354

RESUMO

The extent to which viral genetic context influences HIV adaptation to Human Leukocyte Antigen (HLA) class I-restricted immune pressures remains incompletely understood. The Ugandan HIV epidemic, where major pandemic group M subtypes A1 and D co-circulate in a single host population, provides an opportunity to investigate this question. We characterized plasma HIV RNA gag, pol and nef sequences, along with host HLA genotypes, in 464 antiretroviral-naïve individuals chronically infected with HIV subtypes A1 or D. Using phylogenetically-informed statistical approaches, we identified HLA-associated polymorphisms and formally compared their strengths of selection between viral subtypes. A substantial number (32%) of HLA-associated polymorphisms identified in subtypes A1 and/or D had previously been reported in subtypes B, C and/or Circulating Recombinant Form (CRF) 01_AE, confirming the shared nature of many HLA-driven escape pathways regardless of viral genetic context. Nevertheless, 34% of identified HLA-associated polymorphisms were significantly differentially selected between subtypes A1 and D. Experimental investigation of select examples of subtype-specific escape revealed distinct underlying mechanisms with important implications for vaccine design: whereas some were attributable to subtype-specific sequence variation that influenced epitope-HLA binding, others were attributable to differential mutational barriers to immune escape. Overall, our results confirm HIV genetic context as a key modulator of viral adaptation to host cellular immunity and highlight the power of combined bioinformatic and mechanistic studies, paired with knowledge of epitope immunogenicity, to identify appropriate viral regions for inclusion in subtype-specific and universal HIV vaccine strategies.ImportanceThe identification of HIV polymorphisms reproducibly selected under pressure by specific HLA alleles, and the elucidation of their impact on viral function, can help identify immunogenic viral regions where immune escape incurs a fitness cost. However, our knowledge of HLA-driven escape pathways and their functional costs is largely limited to HIV subtype B, and to a lesser extent C. Our study represents the first characterization of HLA-driven adaptation pathways in HIV subtypes A1 and D, which dominate in East Africa, and the first statistically rigorous characterization of differential HLA-driven escape across viral subtypes. Results support a considerable impact of viral genetic context on HIV adaptation to host HLA, where HIV subtype-specific sequence variation influences both epitope-HLA binding and the fitness costs of escape. Integrated bioinformatic and mechanistic characterization of these and other instances of differential escape could aid rational CTL-based vaccine immunogen selection for both subtype-specific and universal HIV vaccines.

4.
PLoS Pathog ; 14(9): e1007257, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30180214

RESUMO

HIV-1 can downregulate HLA-C on infected cells, using the viral protein Vpu, and the magnitude of this downregulation varies widely between primary HIV-1 variants. The selection pressures that result in viral downregulation of HLA-C in some individuals, but preservation of surface HLA-C in others are not clear. To better understand viral immune evasion targeting HLA-C, we have characterized HLA-C downregulation by a range of primary HIV-1 viruses. 128 replication competent viral isolates from 19 individuals with effective anti-retroviral therapy, show that a substantial minority of individuals harbor latent reservoir virus which strongly downregulates HLA-C. Untreated infections display no change in HLA-C downregulation during the first 6 months of infection, but variation between viral quasispecies can be detected in chronic infection. Vpu molecules cloned from plasma of 195 treatment naïve individuals in chronic infection demonstrate that downregulation of HLA-C adapts to host HLA genotype. HLA-C alleles differ in the pressure they exert for downregulation, and individuals with higher levels of HLA-C expression favor greater viral downregulation of HLA-C. Studies of primary and mutant molecules identify 5 residues in the transmembrane region of Vpu, and 4 residues in the transmembrane domain of HLA-C, which determine interactions between Vpu and HLA. The observed adaptation of Vpu-mediated downregulation to host genotype indicates that HLA-C alleles differ in likelihood of mediating a CTL response that is subverted by viral downregulation, and that preservation of HLA-C expression is favored in the absence of these responses. Finding that latent reservoir viruses can downregulate HLA-C could have implications for HIV-1 cure therapy approaches in some individuals.

5.
J Clin Invest ; 128(9): 4074-4085, 2018 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-30024859

RESUMO

HIV posttreatment controllers (PTCs) represent a natural model of sustained HIV remission, but they are rare and little is known about their viral reservoir. We obtained 1,450 proviral sequences after near-full-length amplification for 10 PTCs and 16 posttreatment noncontrollers (NCs). Before treatment interruption, the median intact and total reservoir size in PTCs was 7-fold lower than in NCs, but the proportion of intact, defective, and total clonally expanded proviral genomes was not significantly different between the 2 groups. Quantification of total but not intact proviral genome copies predicted sustained HIV remission as 81% of NCs, but none of the PTCs had a total proviral genome greater than 4 copies per million peripheral blood mononuclear cells (PBMCs). The results highlight the restricted intact and defective HIV reservoir in PTCs and suggest that total proviral genome burden could act as the first biomarker for identifying PTCs. Total and defective but not intact proviral copy numbers correlated with levels of cell-associated HIV RNA, activated NK cell percentages, and both HIV-specific CD4+ and CD8+ responses. These results support the concept that defective HIV genomes can lead to viral antigen production and interact with both the innate and adaptive immune systems.

6.
AIDS Patient Care STDS ; 32(7): 257-264, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29985647

RESUMO

The prevalence of HIV pretreatment drug resistance (PDR) is increasing in sub-Saharan Africa. We sought to describe correlates of PDR and evaluate effects of PDR on clinical outcomes in rural Uganda. We analyzed data from the Uganda AIDS Rural Treatment Outcomes study, a cohort of antiretroviral therapy (ART)-naive adults with HIV (2005-2015). We performed resistance testing on pre-ART specimens. We defined PDR as any World Health Organization (WHO) 2009 surveillance drug resistance mutation and classified PDR level using the Stanford algorithm. We fit unadjusted and sex-stratified log binomial regression and Cox proportional hazard models to identify correlates of PDR and the impact of PDR on viral suppression, loss to follow-up (LTFU), and death. We analyzed data from 738 participants (median age 33 years, 69% female). Overall, prevalence of PDR was 3.5% (n = 26), owing mostly to resistance to non-nucleoside reverse transcriptase inhibitors. PDR increased over time in women (1.8% in those enrolling in clinic in 2001-2006, vs. 7.0% in 2007-2013; p = 0.006), but not in men (1.15% vs. 0.72%, p = 0.737). Lower pre-ART log10 HIV RNA was also associated with higher prevalence of PDR. We identified longer time to viral suppression among those with PDR compared with without PDR (0.5 and 0.3 years, respectively, p = 0.023), but there was no significant relationship with mortality or LTFU (p = 0.139). We observed increasing rates of PDR in women in southwestern Uganda. Implications of this trend, particularly to prevention of mother-to-child transmission programs in the region, require attention due to delayed viral suppression among those with PDR.

7.
Immunity ; 48(6): 1183-1194.e5, 2018 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-29802019

RESUMO

HIV-1 infection of CD4+ T cells leads to cytopathic effects and cell demise, which is counter to the observation that certain HIV-1-infected cells possess a remarkable long-term stability and can persist lifelong in infected individuals treated with suppressive antiretroviral therapy (ART). Using quantitative mass spectrometry-based proteomics, we showed that HIV-1 infection activated cellular survival programs that were governed by BIRC5, a molecular inhibitor of cell apoptosis that is frequently overexpressed in malignant cells. BIRC5 and its upstream regulator OX40 were upregulated in productively and latently infected CD4+ T cells and were functionally involved in maintaining their viability. Moreover, OX40-expressing CD4+ T cells from ART-treated patients were enriched for clonally expanded HIV-1 sequences, and pharmacological inhibition of BIRC5 resulted in a selective decrease of HIV-1-infected cells in vitro. Together, these findings suggest that BIRC5 supports long-term survival of HIV-1-infected cells and may lead to clinical strategies to reduce persisting viral reservoirs.

8.
AIDS ; 32(7): 921-926, 2018 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-29424775

RESUMO

OBJECTIVE: The aim of our study was to analyze the dynamics of HIV-DNA levels in CD4 T-cell subsets in individuals starting successful dolutegravir-based regimens. DESIGN: Twenty-seven individuals with acute infection (AI, n = 8) or chronic infection (CI, n = 5) and patients in virological success (VS, n = 10) or virological failure (VF, n = 4) on antiretroviral therapy (ART) who initiated a dolutegravir-based regimen were enrolled (NCT02557997). METHODS: CD4 T-cells from baseline and week 48 of successful treatment were sorted into effector memory (TEM), transitional memory (TTM), central memory (TCM) and naïve (TN) cell groups for total HIV-DNA measurements by qPCR. Bayesian methods were used to estimate the posterior probability of a HIV-DNA decrease more than 0.25 log copies/10 cells at week 48. RESULTS: All patients achieved HIV-RNA suppression at 48 weeks. At baseline and week 48, the highest contributions to the HIV-DNA-infected pool from CD4 T cells were observed in TTM cells in the AI group (62.4 and 60.2%, respectively), but in TCM cells for the CI, VS and VF groups (54.6 and 59.4%, 58.2 and 62.9%, 62.4 and 67.2%), respectively. HIV-DNA burden declined in all subsets after 48 weeks of treatment in the AI (probability (Pr) > 91%), CI (Pr > 52%) and VF (Pr > 52%) groups, but only in TEM cells in the VS group (Pr = 95%). CONCLUSION: Our study showed that dolutegravir-based treatment reduced the HIV-DNA cellular burden in individuals from the AI, CI and VF groups, though the reduction levels differed between the patient subgroups. Early treated patients had the highest probability of HIV-DNA reduction. Interestingly, in the aviremic VS group, HIV-DNA reduction was limited to TEM cells.

9.
PLoS Pathog ; 14(1): e1006792, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29324842

RESUMO

Therapeutic strategies aimed at achieving antiretroviral therapy (ART)-free HIV remission in infected individuals are under active investigation. Considering the vast majority of HIV-infected individuals experience plasma viral rebound upon cessation of therapy, clinical trials evaluating the efficacy of curative strategies would likely require inclusion of ART interruption. However, it is unclear what impact short-term analytical treatment interruption (ATI) and subsequent reinitiation of ART have on immunologic and virologic parameters of HIV-infected individuals. Here, we show a significant increase of HIV burden in the CD4+ T cells of infected individuals during ATI that was correlated with the level of plasma viral rebound. However, the size of the HIV reservoirs as well as immune parameters, including markers of exhaustion and activation, returned to pre-ATI levels 6-12 months after the study participants resumed ART. Of note, the proportions of near full-length, genome-intact and structurally defective HIV proviral DNA sequences were similar prior to ATI and following reinitiation of ART. In addition, there was no evidence of emergence of antiretroviral drug resistance mutations within intact HIV proviral DNA sequences following reinitiation of ART. These data demonstrate that short-term ATI does not necessarily lead to expansion of the persistent HIV reservoir nor irreparable damages to the immune system in the peripheral blood, warranting the inclusion of ATI in future clinical trials evaluating curative strategies.


Assuntos
Antirretrovirais/administração & dosagem , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1 , Carga Viral/efeitos dos fármacos , Adulto , Biomarcadores/análise , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/patologia , Linfócitos T CD8-Positivos/virologia , Estudos de Coortes , Esquema de Medicação , Feminino , Infecções por HIV/sangue , HIV-1/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Suspensão de Tratamento
10.
J Antimicrob Chemother ; 73(4): 1045-1053, 2018 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-29244129

RESUMO

Objectives: Better understanding of the dynamics of HIV reservoirs under ART is a critical step to achieve a functional HIV cure. Our objective was to assess the genetic diversity of archived HIV-1 DNA over 48 weeks in blood cells of individuals starting treatment with a dolutegravir-based regimen. Methods: Eighty blood samples were prospectively and longitudinally collected from 20 individuals (NCT02557997) including: acutely (n = 5) and chronically (n = 5) infected treatment-naive individuals, as well as treatment-experienced individuals who switched to a dolutegravir-based regimen and were either virologically suppressed (n = 5) or had experienced treatment failure (n = 5). The integrase and V3 loop regions of HIV-1 DNA isolated from PBMCs were analysed by pyrosequencing at baseline and weeks 4, 24 and 48. HIV-1 genetic diversity was calculated using Shannon entropy. Results: All individuals achieved or maintained viral suppression throughout the study. A low and stable genetic diversity of archived HIV quasispecies was observed in individuals starting treatment during acute infection. A dramatic reduction of the genetic diversity was observed at week 4 of treatment in the other individuals. In these patients and despite virological suppression, a recovery of the genetic diversity of the reservoirs was observed up to 48 weeks. Viral variants bearing dolutegravir resistance-associated substitutions at integrase position 50, 124, 230 or 263 were detected in five individuals (n = 5/20, 25%) from all groups except those who were ART-failing at baseline. None of these substitutions led to virological failure. Conclusions: These data demonstrate that the genetic diversity of the HIV-1 reservoir is reshaped following the initiation of a dolutegravir-based regimen and strongly suggest that HIV-1 can continue to replicate despite successful treatment.

11.
PLoS Med ; 14(11): e1002461, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29182633

RESUMO

BACKGROUND: Notwithstanding 1 documented case of HIV-1 cure following allogeneic stem cell transplantation (allo-SCT), several subsequent cases of allo-SCT in HIV-1 positive individuals have failed to cure HIV-1 infection. The aim of our study was to describe changes in the HIV reservoir in a single chronically HIV-infected patient on suppressive antiretroviral therapy who underwent allo-SCT for treatment of acute lymphoblastic leukemia. METHODS AND FINDINGS: We prospectively collected peripheral blood mononuclear cells (PBMCs) by leukapheresis from a 55-year-old man with chronic HIV infection before and after allo-SCT to measure the size of the HIV-1 reservoir and characterize viral phylogeny and phenotypic changes in immune cells. At day 784 post-transplant, when HIV-1 was undetectable by multiple measures-including PCR measurements of both total and integrated HIV-1 DNA, replication-competent virus measurement by large cell input quantitative viral outgrowth assay, and in situ hybridization of colon tissue-the patient consented to an analytic treatment interruption (ATI) with frequent clinical monitoring. He remained aviremic off antiretroviral therapy until ATI day 288, when a low-level virus rebound of 60 HIV-1 copies/ml occurred, which increased to 1,640 HIV-1 copies/ml 5 days later, prompting reinitiation of ART. Rebounding plasma HIV-1 sequences were phylogenetically distinct from proviral HIV-1 DNA detected in circulating PBMCs before transplantation. The main limitations of this study are the insensitivity of reservoir measurements, and the fact that it describes a single case. CONCLUSIONS: allo-SCT led to a significant reduction in the size of the HIV-1 reservoir and a >9-month-long ART-free remission from HIV-1 replication. Phylogenetic analyses suggest that the origin of rebound virus was distinct from the viruses identified pre-transplant in the PBMCs.


Assuntos
Infecções por HIV/terapia , Carga Viral/efeitos dos fármacos , Antirretrovirais/uso terapêutico , HIV/genética , Infecções por HIV/virologia , HIV-1/genética , Humanos , Leucócitos Mononucleares , Masculino , Pessoa de Meia-Idade , Filogenia , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Transplante de Células-Tronco/métodos , Carga Viral/fisiologia
12.
AIDS ; 31(17): 2345-2354, 2017 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-28832407

RESUMO

OBJECTIVES: HIV-1 subtypes A1 and D cocirculate in a rural community in Mbarara, Uganda. This study examines HIV-1 intersubtype recombination in this community under a full-genome sequencing context. We aim to estimate prevalence, examine time trends, and test for clinical correlates and outcomes associated with intersubtype recombinants. METHODS: Near-full-genome HIV-1 Sanger sequence data were collected from plasma samples of 504 treatment-naïve individuals, who then received protease inhibitor or nonnucleoside reverse transcriptase inhibitor-containing regimens and were monitored for up to 7.5 years. Subtypes were inferred by Los Alamos Recombinant Identification Program (RIP) 3.0 and compared with Sanger/REGA and MiSeq/RIP. 'Nonrecombinants' and 'recombinants' infections were compared in terms of pretherapy viral load, CD4 cell count, posttherapy time to virologic suppression, virologic rebound, first CD4 rise above baseline and sustained CD4 recovery. RESULTS: Prevalence of intersubtype recombinants varied depending on the genomic region examined: gag (15%), prrt (11%), int (8%), vif (10%), vpr (2%), vpu (9%), GP120 (8%), GP41 (18%), and nef (4%). Of the 200 patients with near-full-genome data, prevalence of intersubtype recombination was 46%; the most frequently observed recombinant was A1-D (25%). Sanger/REGA and MiSeq/RIP yielded generally consistent results. Phylogenetic tree revealed most recombinants did not share common ancestors. No temporal trend was observed (all P > 0.1). Subsequent subtype switches were detected in 27 of 143 (19%) study participants with follow-up sequences. Nonrecombinant versus recombinants infections were not significantly different in any pre nor posttherapy clinical correlates examined (all P > 0.2). CONCLUSION: Intersubtype recombination was highly prevalent (46%) in Uganda if the entire HIV genome was considered, but was neither associated with clinical correlates nor therapy outcomes.


Assuntos
Genótipo , Infecções por HIV/patologia , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/genética , Recombinação Genética , Adulto , Antirretrovirais/uso terapêutico , Contagem de Linfócito CD4 , Estudos Transversais , Feminino , Genoma Viral , Infecções por HIV/epidemiologia , HIV-1/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Estudos Longitudinais , Masculino , Prevalência , Análise de Sequência de DNA , Resposta Viral Sustentada , Resultado do Tratamento , Uganda/epidemiologia , Carga Viral
13.
J Clin Invest ; 127(7): 2689-2696, 2017 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-28628034

RESUMO

HIV-1 causes a chronic, incurable disease due to its persistence in CD4+ T cells that contain replication-competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating CD4+ T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, ex vivo-isolated CD4+ T cells, and subsets of functionally polarized memory CD4+ T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of CD4+ T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such clonally expanded CD4+ T cells constituted 62% of all analyzed genome-intact sequences in memory CD4 T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected over a duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized CD4+ T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected CD4+ T cells.


Assuntos
Genoma Viral/imunologia , HIV-1/fisiologia , Células Th1/imunologia , Células Th1/virologia , Latência Viral/imunologia , Adulto , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade
14.
AIDS ; 31(14): 1935-1943, 2017 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-28650381

RESUMO

OBJECTIVE: Long-acting rilpivirine is a candidate for preexposure prophylaxis (PrEP) for prevention of HIV-1 infection. However, rilpivirine resistance mutations at reverse transcriptase codon 138 (E138X) occur naturally in a minority of HIV-1-infected persons; in particular those expressing human leukocyte antigen (HLA)-B18 where reverse transcriptase-E138X arises as an immune escape mutation. We investigate the global prevalence, B18-linkage and replicative cost of reverse transcriptase-E138X and its regional implications for rilpivirine PrEP. METHODS: We analyzed linked reverse transcriptase-E138X/HLA data from 7772 antiretroviral-naive patients from 16 cohorts spanning five continents and five HIV-1 subtypes, alongside unlinked global reverse transcriptase-E138X and HLA frequencies from public databases. E138X-containing HIV-1 variants were assessed for in-vitro replication as a surrogate of mutation stability following transmission. RESULTS: Reverse transcriptase-E138X variants, where the most common were rilpivirine resistance-associated mutations E138A/G/K, were significantly enriched in HLA-B18-positive individuals globally (P = 3.5 × 10) and in all HIV-1 subtypes except A. Reverse transcriptase-E138X and B18 frequencies correlated positively in 16 cohorts with linked HIV/HLA genotypes (Spearman's R = 0.75; P = 7.6 × 10) and in unlinked HIV/HLA data from 43 countries (Spearman's R = 0.34, P = 0.02). Notably, reverse transcriptase-E138X frequencies approached (or exceeded) 10% in key epidemic regions (e.g. sub-Saharan Africa, Southeastern Europe) where B18 is more common. This, along with the observation that reverse transcriptase-E138X variants do not confer in-vitro replicative costs, supports their persistence, and ongoing accumulation in circulation over time. CONCLUSIONS: Results illustrate the potential for a natural immune-driven HIV-1 polymorphism to compromise antiretroviral-based prevention, particularly in key epidemic regions. Regional reverse transcriptase-E138X surveillance should be undertaken before use of rilpivirine PrEP.


Assuntos
Antirretrovirais/farmacologia , Farmacorresistência Viral , Infecções por HIV/prevenção & controle , HIV-1/imunologia , Evasão da Resposta Imune , Mutação de Sentido Incorreto , Profilaxia Pré-Exposição , Saúde Global , Infecções por HIV/virologia , Transcriptase Reversa do HIV/genética , HIV-1/enzimologia , HIV-1/genética , Antígeno HLA-B18/genética , Humanos , Polimorfismo Genético , Rilpivirina/farmacologia
15.
J Acquir Immune Defic Syndr ; 76(2): 188-192, 2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28639996

RESUMO

Despite a poor toxicity profile, zidovudine supersedes abacavir (ABC) as an alternative first-line agent in most international treatment guidelines because of concerns about HLA-B*57:01-related ABC-hypersensitivity. We detected one case of HLA-B*57:01 carriage among 513 HIV-infected individuals in Uganda, which, in combination with previous reports, supports the safety of ABC in the region.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Didesoxinucleosídeos/uso terapêutico , Infecções por HIV/tratamento farmacológico , Adulto , África ao Sul do Saara , Contagem de Linfócito CD4 , Estudos de Coortes , Feminino , Seguimentos , Técnicas de Genotipagem , HIV-1/efeitos dos fármacos , HIV-1/isolamento & purificação , Antígenos HLA-B/sangue , Antígenos HLA-B/genética , Humanos , Masculino , Projetos Piloto , Carga Viral
16.
J Virol ; 91(13)2017 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-28424286

RESUMO

There are marked differences in the spread and prevalence of HIV-1 subtypes worldwide, and differences in clinical progression have been reported. However, the biological reasons underlying these differences are unknown. Gag-protease is essential for HIV-1 replication, and Gag-protease-driven replication capacity has previously been correlated with disease progression. We show that Gag-protease replication capacity correlates significantly with that of whole isolates (r = 0.51; P = 0.04), indicating that Gag-protease is a significant contributor to viral replication capacity. Furthermore, we investigated subtype-specific differences in Gag-protease-driven replication capacity using large well-characterized cohorts in Africa and the Americas. Patient-derived Gag-protease sequences were inserted into an HIV-1 NL4-3 backbone, and the replication capacities of the resulting recombinant viruses were measured in an HIV-1-inducible reporter T cell line by flow cytometry. Recombinant viruses expressing subtype C Gag-proteases exhibited substantially lower replication capacities than those expressing subtype B Gag-proteases (P < 0.0001); this observation remained consistent when representative Gag-protease sequences were engineered into an HIV-1 subtype C backbone. We identified Gag residues 483 and 484, located within the Alix-binding motif involved in virus budding, as major contributors to subtype-specific replicative differences. In East African cohorts, we observed a hierarchy of Gag-protease-driven replication capacities, i.e., subtypes A/C < D < intersubtype recombinants (P < 0.0029), which is consistent with reported intersubtype differences in disease progression. We thus hypothesize that the lower Gag-protease-driven replication capacity of subtypes A and C slows disease progression in individuals infected with these subtypes, which in turn leads to greater opportunity for transmission and thus increased prevalence of these subtypes.IMPORTANCE HIV-1 subtypes are unevenly distributed globally, and there are reported differences in their rates of disease progression and epidemic spread. The biological determinants underlying these differences have not been fully elucidated. Here, we show that HIV-1 Gag-protease-driven replication capacity correlates with the replication capacity of whole virus isolates. We further show that subtype B displays a significantly higher Gag-protease-mediated replication capacity than does subtype C, and we identify a major genetic determinant of these differences. Moreover, in two independent East African cohorts we demonstrate a reproducible hierarchy of Gag-protease-driven replicative capacity, whereby recombinants exhibit the greatest replication, followed by subtype D, followed by subtypes A and C. Our data identify Gag-protease as a major determinant of subtype differences in disease progression among HIV-1 subtypes; furthermore, we propose that the poorer viral replicative capacity of subtypes A and C may paradoxically contribute to their more efficient spread in sub-Saharan Africa.


Assuntos
Progressão da Doença , Genótipo , Infecções por HIV/patologia , Infecções por HIV/virologia , HIV-1/fisiologia , Replicação Viral , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética , África Oriental , Variação Genética , HIV-1/classificação , HIV-1/genética , Humanos , Liberação de Vírus
17.
J Virol Methods ; 240: 32-41, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27865749

RESUMO

The HIV-1 accessory protein Vpu exhibits high inter- and intra- subtype genetic diversity that may influence Vpu function and possibly contribute to HIV-1 pathogenesis. However, scalable methods to evaluate genotype/phenotype relationships in natural Vpu sequences are limited, particularly those expressing the protein in CD4+ T-cells, the natural target of HIV-1 infection. A major impediment to assay scalability is the extensive genetic diversity within, and immediately upstream of, Vpu's initial 5' coding region, which has necessitated the design of oligonucleotide primers specific for each individual HIV-1 isolate (or subtype). To address this, we developed two universal forward primers, located in relatively conserved regions 38 and 90 bases upstream of Vpu, and a single universal reverse primer downstream of Vpu, which are predicted to cover the vast majority of global HIV-1 group M sequence diversity. We show that inclusion of up to 90 upstream bases of HIV-1 genomic sequence does not significantly influence in vitro Vpu expression or function when a Rev/Rev Response Element (RRE)-dependent expression system is used. We further assess the function of four diverse HIV-1 Vpu sequences, revealing reproducible and significant differences between them. Our approach represents a scalable option to measure the in vitro function of genetically diverse natural Vpu isolates in a CD4+ T-cell line.


Assuntos
Primers do DNA , Variação Genética , HIV-1/genética , Proteínas do Vírus da Imunodeficiência Humana/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Proteínas Virais Reguladoras e Acessórias/genética , Linfócitos T CD4-Positivos , Regulação para Baixo , Infecções por HIV/virologia , Proteínas do Vírus da Imunodeficiência Humana/metabolismo , Humanos , Elementos de Resposta , Proteínas Virais Reguladoras e Acessórias/metabolismo
18.
AIDS ; 30(11): 1781-8, 2016 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-27124899

RESUMO

BACKGROUND: Previous studies suggest that infection with non-R5-tropic subtype B HIV-1, compared with R5, is associated with a more rapid decline in CD4 cell count, but does not affect PI/(N)NRTI therapy outcome. Here, we explored clinical correlates associated with viral tropism in subtype A1 and D infections. METHODS: HIV-1 subtype A1 (n = 196) and D (n = 143) pretherapy plasma samples and up to 7.5 years of posttherapy virologic and CD4 data were collected from a cross-sectional cohort in Mbarara, Uganda. Tropism and subtype were inferred using env V3 (geno2pheno) and gp41 (RIP) Sanger sequences. For each subtype, R5 infection was compared with non-R5 in terms of: pretherapy viral load and CD4 cell count (Mann-Whitney tests), and therapy outcomes, including time to virologic suppression, postsuppression virologic rebound, CD4 decline and CD4 recovery (log-rank tests). RESULTS: A 94% of all patients in this study achieved virologic suppression within median 3 months posttherapy. In both subtypes, non-R5 infection was associated with lower pretherapy CD4 cell count (non-R5 vs. R5; A: median 57 vs. 147 cells/µl P = 0.005; D: 80 vs. 128 cells/µl P = 0.006). Multivariable linear regression confirmed that tropism, not subtype nor the interaction between subtype and tropism, was a significant predictor of pretherapy CD4 cell count (P < 0.0001). None of pretherapy viral load, time to virologic suppression, virologic rebound, CD4 decline nor CD4 recovery was significantly different (all P > 0.09). CONCLUSION: Regardless of HIV-1 subtype or tropism, the majority of patients in this Ugandan cohort responded to therapy, even though non-R5 infection was associated with lower pretherapy CD4 cell count.


Assuntos
Antirretrovirais/uso terapêutico , Genótipo , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/genética , Tropismo Viral , Adulto , Contagem de Linfócito CD4 , Estudos Transversais , Feminino , Infecções por HIV/tratamento farmacológico , HIV-1/isolamento & purificação , HIV-1/fisiologia , Humanos , Masculino , Análise de Sequência de DNA , Resultado do Tratamento , Uganda , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
19.
Curr Opin HIV AIDS ; 11(4): 383-7, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27023286

RESUMO

PURPOSE OF REVIEW: HIV-1 is able to create lasting reservoirs of virally infected cells that persist life-long and are extremely difficult to eradicate, thus necessitating indefinite antiretroviral therapy. Large numbers of studies suggest that CD4 T cells represent the major, and possibly the only cell type supporting HIV-1 long-term persistence. However, the ability to serve as long-term viral reservoirs may be confined to certain subpopulations of CD4 T cells with specific functional and developmental characteristics that HIV-1 can selectively exploit to propagate long-term viral survival within the host. Identification of CD4 T-cell subtypes that serve as hotspots for viral persistence may be critical for designing strategies to purge the immune system of persisting viral reservoirs. RECENT FINDINGS: Developmentally immature, long-lasting CD4 memory T-cell populations seem to contain the majority of latently HIV-1-infected cells that persist despite antiretroviral therapy in the peripheral blood. Emerging data suggest that functional polarization toward a T helper 17 (Th17), a T follicular helper cell or a regulatory T-cell lineage may also be associated with an increased ability to serve as a viral reservoir site. Atypical T cells such a γδ CD4 T cells or tissue-resident memory CD4 T cells may be predestined to serve as sites for HIV-1 persistence in specific tissues, but will require additional exploration in future studies. SUMMARY: Recent advances have increased awareness for the profound diversity and complexity of CD4 T-cell subpopulations serving as sites for HIV-1 persistence. Continuous technological and methodological improvements to interrogate viral reservoirs in distinct CD4 T-cell subpopulations may allow to define a more complete landscape of the HIV-1 reservoir composition in different T-cell subpopulations.


Assuntos
Linfócitos T CD4-Positivos/virologia , Infecções por HIV/virologia , HIV-1/fisiologia , Subpopulações de Linfócitos T/virologia , Latência Viral , Humanos
20.
J Antimicrob Chemother ; 70(7): 2090-6, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25755001

RESUMO

OBJECTIVES: To assess the prevalence of minority resistant variants (MRVs) at baseline and their impact on the virological response. The ANRS 139 TRIO trial evaluated the combination of raltegravir, etravirine and darunavir, plus an optimized background therapy, in 87% of cases. Patients were highly experienced and harboured multiresistant viruses, but were naive to the three drugs, and showed a high level of virological suppression. METHODS: Ultra-deep sequencing of reverse transcriptase, protease and integrase regions was performed at the trial baseline, and sequences were interpreted according to the ANRS algorithm. MRVs were assessed using MiSeq and 454 technologies (limit of detection 1%). RESULTS: At baseline, minority variants with at least one NRTI, one NNRTI, one PI, one major PI or an integrase inhibitor resistance-associated mutation were present in 46%, 45%, 68%, 24% and 13% of patients, respectively. When minority variants are taken into account, the prevalence of resistance to etravirine, darunavir and raltegravir at baseline was 29%, 40% and 9%, respectively. No difference was observed in the prevalence of MRVs between patients with virological failure and those with virological success, except a trend for patients exhibiting baseline etravirine MRVs (50% versus 26%, P = 0.09). CONCLUSIONS: We have shown a high level of MRVs at baseline in highly pre-treated patients harbouring multiresistant viruses. However, these MRVs were not associated with an increased risk of virological failure, except for a trend for etravirine MRVs.


Assuntos
Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/uso terapêutico , Farmacorresistência Viral , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , Darunavir/farmacologia , Darunavir/uso terapêutico , Integrase de HIV/genética , Protease de HIV/genética , Transcriptase Reversa do HIV/genética , HIV-1/genética , HIV-1/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Estudos Longitudinais , Mutação , Piridazinas/farmacologia , Piridazinas/uso terapêutico , Raltegravir Potássico/farmacologia , Raltegravir Potássico/uso terapêutico , Resultado do Tratamento , Carga Viral
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