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1.
Open Forum Infect Dis ; 11(5): ofae212, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38756763

RESUMO

Background: Persistence of HIV-1 in reservoirs necessitates life-long antiretroviral therapy (ART). There are conflicting data using genetic analysis on whether persistence includes an actively replicating reservoir with strong evidence arguing against replication. Methods: We investigated the possibility of ongoing viral evolution during suppressive therapy by comparing near full-length viral genomic sequences using phylogenetic analysis of viral RNA in plasma before therapy initiation early after infection and from virus induced to grow from the latent reservoir after a period of suppressive ART. We also focused our analysis on evidence of selective pressure by drugs in the treatment regimen and at sites of selective pressure by the adaptive immune response. Results: Viral genomes induced to grow from the latent reservoir from 10 participants with up to 9 years on suppressive ART were highly similar to the nearly homogeneous sequences in plasma taken early after infection at ART initiation. This finding was consistent across the entire genome and when the analysis focused on sites targeted by the drug regimen and by host selective pressure of antibody and cytotoxic T cells. The lack of viral evolution away from pretherapy sequences in spite of demonstrated selective pressure is most consistent with a lack of viral replication during reservoir persistence. Conclusions: These results do not support ongoing viral replication as a mechanism of HIV-1 persistence during suppressive ART.

2.
J Infect Dis ; 229(3): 743-752, 2024 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-38349333

RESUMO

BACKGROUND: The histone deacetylase inhibitor vorinostat (VOR) can reverse human immunodeficiency virus type 1 (HIV-1) latency in vivo and allow T cells to clear infected cells in vitro. HIV-specific T cells (HXTCs) can be expanded ex vivo and have been safely administered to people with HIV (PWH) on antiretroviral therapy. METHODS: Six PWH received infusions of 2 × 107 HXTCs/m² with VOR 400 mg, and 3 PWH received infusions of 10 × 107 HXTCs/m² with VOR. The frequency of persistent HIV by multiple assays including quantitative viral outgrowth assay (QVOA) of resting CD4+ T cells was measured before and after study therapy. RESULTS: VOR and HXTCs were safe, and biomarkers of serial VOR effect were detected, but enhanced antiviral activity in circulating cells was not evident. After 2 × 107 HXTCs/m² with VOR, 1 of 6 PWH exhibited a decrease in QVOA, and all 3 PWH exhibited such declines after 10 × 107 HXTCs/m² and VOR. However, most declines did not exceed the 6-fold threshold needed to definitively attribute decline to the study intervention. CONCLUSIONS: These modest effects provide support for the strategy of HIV latency reversal and reservoir clearance, but more effective interventions are needed to yield the profound depletion of persistent HIV likely to yield clinical benefit. Clinical Trials Registration. NCT03212989.


Assuntos
Infecções por HIV , HIV-1 , Humanos , Vorinostat/uso terapêutico , Vorinostat/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Inibidores de Histona Desacetilases/farmacologia , Linfócitos T CD4-Positivos , Terapia Baseada em Transplante de Células e Tecidos , Latência Viral
3.
Clin Exp Immunol ; 215(3): 279-290, 2024 02 19.
Artigo em Inglês | MEDLINE | ID: mdl-37950348

RESUMO

CD8 T cells recognize infected and cancerous cells via their T-cell receptor (TCR), which binds peptide-MHC complexes on the target cell. The affinity of the interaction between the TCR and peptide-MHC contributes to the antigen sensitivity, or functional avidity, of the CD8 T cell. In response to peptide-MHC stimulation, the TCR-CD3 complex and CD8 co-receptor are downmodulated. We quantified CD3 and CD8 downmodulation following stimulation of human CD8 T cells with CMV, EBV, and HIV peptides spanning eight MHC restrictions, observing a strong correlation between the levels of CD3 and CD8 downmodulation and functional avidity, regardless of peptide viral origin. In TCR-transduced T cells targeting a tumor-associated antigen, changes in TCR-peptide affinity were sufficient to modify CD3 and CD8 downmodulation. Correlation analysis and generalized linear modeling indicated that CD3 downmodulation was the stronger correlate of avidity. CD3 downmodulation, simply measured using flow cytometry, can be used to identify high-avidity CD8 T cells in a clinical context.


Assuntos
Linfócitos T CD8-Positivos , Receptores de Antígenos de Linfócitos T , Humanos , Regulação para Baixo , Receptores de Antígenos de Linfócitos T/genética , Antígenos CD8/metabolismo , Peptídeos/metabolismo , Complexo CD3/metabolismo
4.
J Clin Invest ; 132(8)2022 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-35426377

RESUMO

Latency reversal strategies for HIV cure using inhibitor of apoptosis protein (IAP) antagonists (IAPi) induce unprecedented levels of latent reservoir expression without immunotoxicity during suppressive antiretroviral therapy (ART). However, full targeting of the reservoir may require combinatorial approaches. A Jurkat latency model screen for IAPi combination partners demonstrated synergistic latency reversal with bromodomain (BD) and extraterminal domain protein inhibitors (BETi). Mechanistic investigations using CRISPR-CAS9 and single-cell RNA-Seq informed comprehensive ex vivo evaluations of IAPi plus pan-BET, bD-selective BET, or selective BET isoform targeting in CD4+ T cells from ART-suppressed donors. IAPi+BETi treatment resulted in striking induction of cell-associated HIV gag RNA, but lesser induction of fully elongated and tat-rev RNA compared with T cell activation-positive controls. IAPi+BETi resulted in HIV protein induction in bulk cultures of CD4+ T cells using an ultrasensitive p24 assay, but did not result in enhanced viral outgrowth frequency using a standard quantitative viral outgrowth assay. This study defines HIV transcriptional elongation and splicing as important barriers to latent HIV protein expression following latency reversal, delineates the roles of BET proteins and their BDs in HIV latency, and provides a rationale for exploration of IAPi+BETi in animal models of HIV latency.


Assuntos
Infecções por HIV , HIV-1 , Animais , Linfócitos T CD4-Positivos , Infecções por HIV/tratamento farmacológico , Infecções por HIV/genética , HIV-1/fisiologia , Proteínas do Vírus da Imunodeficiência Humana , NF-kappa B/genética , NF-kappa B/metabolismo , Proteínas Nucleares/metabolismo , RNA , Fatores de Transcrição/metabolismo , Ativação Viral , Latência Viral
5.
J Infect Dis ; 225(5): 856-861, 2022 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-34562096

RESUMO

We tested the combination of a broadly neutralizing HIV antibody with the latency reversal agent vorinostat (VOR). Eight participants received 2 month-long cycles of VRC07-523LS with VOR. Low-level viremia, resting CD4+ T-cell-associated HIV RNA (rca-RNA) was measured, and intact proviral DNA assay (IPDA) and quantitative viral outgrowth assay (QVOA) were performed at baseline and posttreatment. In 3 participants, IPDA and QVOA declines were accompanied by significant declines of rca-RNA. However, no IPDA or QVOA declines clearly exceeded assay variance or natural decay. Increased resistance to VRC07-523LS was not observed. This combination therapy did not reduce viremia or the HIV reservoir. Clinical Trials Registration. NCT03803605.


Assuntos
Infecções por HIV , HIV-1 , Anticorpos Amplamente Neutralizantes , Linfócitos T CD4-Positivos , HIV-1/genética , Humanos , Viremia/tratamento farmacológico , Latência Viral , Vorinostat/uso terapêutico
6.
J Virol ; 95(6)2021 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-33361426

RESUMO

The HIV proviral reservoir is the major barrier to cure. The predominantly replication-defective proviral landscape makes the measurement of virus that is likely to cause rebound upon antiretroviral therapy (ART)-cessation challenging. To address this issue, novel assays to measure intact HIV proviruses have been developed. The intact proviral DNA assay (IPDA) is a high-throughput assay that uses two probes to exclude the majority of defective proviruses and determine the frequency of intact proviruses, albeit without sequence confirmation. Quadruplex PCR with four probes (Q4PCR) is a lower-throughput assay that uses limiting dilution long-distance PCR amplification followed by quantitative PCR (qPCR) and near-full-length genome sequencing (nFGS) to estimate the frequency of sequence-confirmed intact proviruses and provide insight into their clonal composition. To explore the advantages and limitations of these assays, we compared IPDA and Q4PCR measurements from 39 ART-suppressed people living with HIV. We found that IPDA and Q4PCR measurements correlated with one another, but frequencies of intact proviral DNA differed by approximately 19-fold. This difference may be in part due to inefficiencies in long-distance PCR amplification of proviruses in Q4PCR, leading to underestimates of intact proviral frequencies. In addition, nFGS analysis within Q4PCR explained that some of this difference is explained by proviruses that are classified as intact by IPDA but carry defects elsewhere in the genome. Taken together, this head-to-head comparison of novel intact proviral DNA assays provides important context for their interpretation in studies to deplete the HIV reservoir and shows that together the assays bracket true reservoir size.IMPORTANCE The intact proviral DNA assay (IPDA) and quadruplex PCR (Q4PCR) represent major advances in accurately quantifying and characterizing the replication-competent HIV reservoir. This study compares the two novel approaches for measuring intact HIV proviral DNA in samples from 39 antiretroviral therapy (ART)-suppressed people living with HIV, thereby informing ongoing efforts to deplete the HIV reservoir in cure-related trials.


Assuntos
Infecções por HIV/virologia , HIV-1/genética , Técnicas de Diagnóstico Molecular/métodos , Provírus/genética , Antirretrovirais/uso terapêutico , Sequência de Bases , Linfócitos T CD4-Positivos/virologia , DNA Viral/genética , Genes env/genética , Genoma Viral/genética , Infecções por HIV/tratamento farmacológico , HIV-1/isolamento & purificação , HIV-1/fisiologia , Reação em Cadeia da Polimerase , Polimorfismo Genético , Provírus/isolamento & purificação , Provírus/fisiologia , Carga Viral , Sequência de Empacotamento Viral/genética , Latência Viral
7.
J Infect Dis ; 224(1): 92-100, 2021 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-33216132

RESUMO

BACKGROUND: The replication-competent human immunodeficiency virus (HIV) reservoir is the major barrier to cure. The quantitative viral outgrowth assay (QVOA), the gold-standard method to quantify replication-competent HIV, is resource intensive, which limits its application in large clinical trials. The intact proviral DNA assay (IPDA) requires minimal cell input relative to QVOA and quantifies both defective and intact proviral HIV DNA, the latter potentially serving as a surrogate marker for replication-competent provirus. However, there are limited cross-sectional and longitudinal data on the relationship between IPDA and QVOA measurements. METHODS: QVOA and IPDA measurements were performed on 156 resting CD4 T-cell (rCD4) samples from 83 antiretroviral therapy-suppressed HIV-positive participants. Longitudinal QVOA and IPDA measurements were performed on rCD4 from 29 of these participants. RESULTS: Frequencies of intact, defective, and total proviruses were positively associated with frequencies of replication-competent HIV. Longitudinally, decreases in intact proviral frequencies were strikingly similar to that of replication-competent virus in most participants. In contrast, defective proviral DNA frequencies appeared relatively stable over time in most individuals. CONCLUSIONS: Changes in frequencies of IPDA-derived intact proviral DNA and replication-competent HIV measured by QVOA are similar. IPDA is a promising high-throughput approach to estimate changes in the frequency of the replication-competent reservoir.


Assuntos
Antirretrovirais/uso terapêutico , DNA Viral/análise , HIV/isolamento & purificação , Provírus/isolamento & purificação , Adulto , Estudos Transversais , Feminino , HIV/efeitos dos fármacos , HIV/crescimento & desenvolvimento , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Provírus/crescimento & desenvolvimento , Estudos Retrospectivos
8.
Elife ; 92020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-33021198

RESUMO

HIV-1-specific CD8+ T cells are an important component of HIV-1 curative strategies. Viral variants in the HIV-1 reservoir may limit the capacity of T cells to detect and clear virus-infected cells. We investigated the patterns of T cell escape variants in the replication-competent reservoir of 25 persons living with HIV-1 (PLWH) durably suppressed on antiretroviral therapy (ART). We identified all reactive T cell epitopes in the HIV-1 proteome for each participant and sequenced HIV-1 outgrowth viruses from resting CD4+ T cells. All non-synonymous mutations in reactive T cell epitopes were tested for their effect on the size of the T cell response, with a≥50% loss defined as an escape mutation. The majority (68%) of T cell epitopes harbored no detectable escape mutations. These findings suggest that circulating T cells in PLWH on ART could contribute to control of rebound and could be targeted for boosting in curative strategies.


Assuntos
Fármacos Anti-HIV/farmacologia , Linfócitos T CD4-Positivos/virologia , Infecções por HIV/imunologia , Latência Viral/efeitos dos fármacos , Adulto , Idoso , Estudos de Coortes , Epitopos/imunologia , Feminino , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Filogenia , Carga Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
9.
Front Immunol ; 11: 1971, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32849659

RESUMO

Quantifying the inducible HIV reservoir provides an estimate of the frequency of quiescent HIV-infected cells in humans as well as in animal models, and can help ascertain the efficacy of latency reversing agents (LRAs). The quantitative viral outgrowth assay (QVOA) is used to measure inducible, replication competent HIV and generate estimations of reservoir size. However, traditional QVOA is time and labor intensive and requires large amounts of lymphocytes. Given the importance of reproducible and accurate assessment of both reservoir size and LRA activity in cure strategies, efforts to streamline the QVOA are of high priority. We developed a modified QVOA, the Digital ELISA Viral Outgrowth or DEVO assay, with ultra-sensitive p24 readout, capable of femtogram detection of HIV p24 protein in contrast to the picogram limitations of traditional ELISA. For each DEVO assay, 8-12 × 106 resting CD4 + T cells from aviremic, ART-treated HIV + participants are plated in limiting dilution and maximally stimulated with PHA, IL-2 and uninfected allogeneic irradiated PBMC. CD8-depleted PHA blasts from an uninfected donor or HIV-permissive cells (e.g., Molt4/CCR5) are added to the cultures and virus allowed to amplify for 8-12 days. HIV p24 from culture supernatant is measured at day 8 by Simoa (single molecule array, ultra-sensitive p24 assay) confirmed at day 12, and infectious units per million CD4 + T cells (IUPM) are calculated using the maximum likelihood method. In all DEVO assays performed, HIV p24 was detected in the supernatant of cultures as early as 8 days post stimulation. Importantly, DEVO IUPM values at day 8 were comparable or higher than traditional QVOA IUPM values obtained at day 15. Interestingly, DEVO IUPM values were similar with or without the addition of allogeneic CD8-depleted target PHA blasts or HIV permissive cells traditionally used to expand virus. The DEVO assay uses fewer resting CD4 + T cells and provides an assessment of reservoir size in less time than standard QVOA. This assay offers a new platform to quantify replication competent HIV during limited cell availability. Other potential applications include evaluating LRA activity, and measuring clearance of infected cells during latency clearance assays.


Assuntos
Proteína do Núcleo p24 do HIV/metabolismo , Infecções por HIV/diagnóstico , Infecções por HIV/virologia , HIV-1/fisiologia , Carga Viral , Replicação Viral , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Ensaio de Imunoadsorção Enzimática , Infecções por HIV/imunologia , Humanos , Sensibilidade e Especificidade
10.
AIDS ; 34(13): 1923-1931, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32773474

RESUMO

OBJECTIVES: The aim of this study was to evaluate penetration of antiretrovirals into compartments and efficacy of a dual, NRTI-sparing regimen in acute HIV infection (AHI). DESIGN: Single-arm, open-label pilot study of participants with AHI initiating ritonavir-boosted darunavir 800 mg once daily and etravirine 400 mg once daily or 200 mg twice daily within 30 days of AHI diagnosis. METHODS: Efficacy was defined as HIV RNA less than 200 copies/ml by week 24. Optional sub-studies included pharmacokinetics analysis from genital fluids (weeks 0-4, 12, 48), cerebrospinal fluid (CSF) (weeks 2-4, 24 and 48) and endoscopic biopsies (weeks 4-12 and 36-48). Neuropsychological performance was assessed at weeks 0, 24 and 48. RESULTS: Fifteen AHI participants were enrolled. Twelve (80%) participants achieved HIV RNA less than 200 copies/ml by week 24. Among 12 participants retained through week 48, nine (75%) remained suppressed to less than 50 copies/ml. The median time from ART initiation to suppression less than 200 and less than 50 copies/ml was 59 and 86 days, respectively. The penetration ratios for etravirine and darunavir in gut associated lymphoid tissue were 19.2 and 3.05, respectively. Most AHI participants achieving viral suppression experienced neurocognitive improvement. Of the three participants without overall improvement in neurocognitive functioning as measured by impairment ratings (more than two tests below 1 SD), two had virologic failure. CONCLUSION: NRTI-sparing ART started during AHI resulted in rapid viral suppression similar to NRTI-based regimens. More novel and compact two-drug treatments for AHI should be considered. Early institution of ART during AHI appears to improve overall neurocognitive function and may reduce the risk of subsequent neurocognitive impairment. CLINICALTRIALS.GOV:: NCT00855413.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Terapia Antirretroviral de Alta Atividade , Darunavir/farmacocinética , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Ritonavir/farmacocinética , Adulto , Contagem de Linfócito CD4 , Darunavir/uso terapêutico , Farmacorresistência Viral , Quimioterapia Combinada , Feminino , HIV/genética , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Ritonavir/uso terapêutico , Resultado do Tratamento , Carga Viral/efeitos dos fármacos
11.
J Infect Dis ; 222(11): 1843-1852, 2020 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-32496542

RESUMO

BACKGROUND: Persistent HIV infection of long-lived resting CD4 T cells, despite antiretroviral therapy (ART), remains a barrier to HIV cure. Women have a more robust type 1 interferon response during HIV infection relative to men, contributing to lower initial plasma viremia. As lower viremia during acute infection is associated with reduced frequency of latent HIV infection, we hypothesized that women on ART would have a lower frequency of latent HIV compared to men. METHODS: ART-suppressed, HIV seropositive women (n = 22) were matched 1:1 to 22 of 39 ART-suppressed men. We also compared the 22 women to all 39 men, adjusting for age and race as covariates. We measured the frequency of latent HIV using the quantitative viral outgrowth assay, the intact proviral DNA assay, and total HIV gag DNA. We also performed activation/exhaustion immunophenotyping on peripheral blood mononuclear cells and quantified interferon-stimulated gene (ISG) expression in CD4 T cells. RESULTS: We did not observe evident sex differences in the frequency of persistent HIV in resting CD4 T cells. Immunophenotyping and CD4 T-cell ISG expression analysis revealed marginal differences across the sexes. CONCLUSIONS: Differences in HIV reservoir frequency and immune activation appear to be small across sexes during long-term suppressive therapy.


Assuntos
Antirretrovirais/uso terapêutico , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , Latência Viral , Adulto , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Estudos Transversais , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Expressão Gênica , HIV-1/genética , Humanos , Leucócitos Mononucleares , Masculino , Pessoa de Meia-Idade , Fatores Sexuais
12.
Sci Rep ; 10(1): 5134, 2020 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-32198428

RESUMO

Approaches to deplete persistent HIV infection are needed. We investigated the combined impact of the latency reversing agent vorinostat (VOR) and AGS-004, an autologous dendritic cell immunotherapeutic, on the HIV reservoir. HIV+, stably treated participants in whom resting CD4+ T cell-associated HIV RNA (rca-RNA) increased after VOR exposure ex vivo and in vivo received 4 doses of AGS-004 every 3 weeks, followed by VOR every 72 hours for 30 days, and then the cycle repeated. Change in VOR-responsive host gene expression, HIV-specific T cell responses, low-level HIV viremia, rca-RNA, and the frequency of resting CD4+ T-cell infection (RCI) was measured at baseline and after each cycle. No serious treatment-related adverse events were observed among five participants. As predicted, VOR-responsive host genes responded uniformly to VOR dosing. Following cycles of AGS-004 and VOR, rca-RNA decreased significantly in only two participants, with a significant decrease in SCA observed in one of these participants. However, unlike other cohorts dosed with AGS-004, no uniform increase in HIV-specific immune responses following vaccination was observed. Finally, no reproducible decline of RCI, defined as a decrease of >50%, was observed. AGS-004 and VOR were safe and well-tolerated, but no substantial impact on RCI was measured. In contrast to previous clinical data, AGS-004 did not induce HIV-specific immune responses greater than those measured at baseline. More efficacious antiviral immune interventions, perhaps paired with more effective latency reversal, must be developed to clear persistent HIV infection.


Assuntos
Células Dendríticas/transplante , Infecções por HIV/terapia , HIV-1/efeitos dos fármacos , Inibidores de Histona Desacetilases/uso terapêutico , Imunoterapia Adotiva/métodos , Vorinostat/uso terapêutico , Adulto , Linfócitos T CD4-Positivos/imunologia , Humanos , Memória Imunológica/imunologia , Masculino , Pessoa de Meia-Idade , Linfócitos T Citotóxicos/imunologia , Pesquisa Translacional Biomédica , Resultado do Tratamento , Vacinação
13.
Mol Ther Methods Clin Dev ; 15: 9-17, 2019 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-31534983

RESUMO

HIV infection induces a robust T cell response that is sustained by high viremia, but falls following the onset of antiretroviral therapy (ART). Relatively little has been reported on the subsequent stability of the HIV-specific T cell response in individuals on durable therapy. Such data are critical for powering clinical trials testing T cell-based immunotherapies. In a cross-sectional study, HIV-specific T cell responses were detectable by ex vivo interferon (IFN)-γ ELISpot (average ∼1,100 spot-forming units [SFUs]/106 peripheral blood mononuclear cells) in persons living with HIV (PLWH; n = 34), despite median durable ART suppression of 5.0 years. No substantial association was detected between the summed HIV-specific T cell response and the size of the replication-competent HIV reservoir. T cell responses were next measured in participants sampled weekly, monthly, or yearly. HIV-specific T cell responses were highly stable over the time periods examined; within-individual variation ranged from 16% coefficient of variation (CV) for weekly to 27% CV for yearly sampling. These data were used to generate power calculations for future immunotherapy studies. The stability of the HIV-specific T cell response in suppressed PLWH will enable powered studies of small sizes (e.g., n = 6-12), facilitating rapid and iterative testing for T cell-based immunotherapies against HIV.

14.
JCI Insight ; 4(12)2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31217357

RESUMO

In HIV-infected individuals on long-term antiretroviral therapy (ART), more than 40% of the infected cells are in clones. Although most HIV proviruses present in individuals on long-term ART are defective, including those in clonally expanded cells, there is increasing evidence that clones carrying replication-competent proviruses are common in patients on long-term ART and form part of the HIV reservoir that makes it impossible to cure HIV infection with current ART alone. Given the importance of clonal expansion in HIV persistence, we determined how soon after HIV acquisition infected clones can grow large enough to be detected (clones larger than ca. 1 × 105 cells). We studied 12 individuals sampled in early HIV infection (Fiebig stage III-V/VI) and 5 who were chronically infected. The recently infected individuals were started on ART at or near the time of diagnosis. We isolated more than 6,500 independent integration sites from peripheral blood mononuclear cells before ART was initiated and after 0.5-18 years of suppressive ART. Some infected clones could be detected approximately 4 weeks after HIV infection and some of these clones persisted for years. The results help to explain how the reservoir is established early and persists for years.


Assuntos
Infecções por HIV/patologia , Infecções por HIV/virologia , HIV-1/fisiologia , Leucócitos Mononucleares/virologia , Adulto , Fármacos Anti-HIV/uso terapêutico , Células Clonais/virologia , Progressão da Doença , Infecções por HIV/tratamento farmacológico , Humanos , Provírus/fisiologia , Fatores de Tempo , Carga Viral , Integração Viral , Replicação Viral
15.
Mol Ther ; 26(10): 2496-2506, 2018 10 03.
Artigo em Inglês | MEDLINE | ID: mdl-30249388

RESUMO

Adoptive T cell therapy has had dramatic successes in the treatment of virus-related malignancies and infections following hematopoietic stem cell transplantation. We adapted this method to produce ex vivo expanded HIV-specific T cells (HXTCs), with the long-term goal of using HXTCs as part of strategies to clear persistent HIV infection. In this phase 1 proof-of-concept study (NCT02208167), we administered HXTCs to antiretroviral therapy (ART)-suppressed, HIV-infected participants. Participants received two infusions of 2 × 107 cells/m2 HXTCs at a 2-week interval. Leukapheresis was performed at baseline and 12 weeks post-infusion to measure the frequency of resting cell infection by the quantitative viral outgrowth assay (QVOA). Overall, participants tolerated HXTCs, with only grade 1 adverse events (AEs) related to HXTCs. Two of six participants exhibited a detectable increase in CD8 T cell-mediated antiviral activity following the two infusions in some, but not all, assays. As expected, however, in the absence of a latency reversing agent, no meaningful decline in the frequency of resting CD4 T cell infection was detected. HXTC therapy in ART-suppressed, HIV-infected individuals appears safe and well tolerated, without any clinical signs of immune activation, likely due to the low residual HIV antigen burden present during ART.


Assuntos
Terapia Antirretroviral de Alta Atividade/métodos , Terapia Baseada em Transplante de Células e Tecidos , Infecções por HIV/terapia , Linfócitos T/transplante , Adulto , Idoso , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Terapia Genética , Infecções por HIV/genética , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/imunologia , HIV-1/patogenicidade , Humanos , Masculino , Pessoa de Meia-Idade , Linfócitos T/imunologia , Ativação Viral/genética , Ativação Viral/imunologia , Replicação Viral/genética , Replicação Viral/imunologia
16.
AIDS Res Hum Retroviruses ; 34(1): 111-122, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28636433

RESUMO

AGS-004 consists of matured autologous dendritic cells co-electroporated with in vitro transcribed RNA encoding autologous HIV antigens. In an open-label, single arm sub-study of AGS-004-003, AGS-004 was administered monthly to suppressed participants who started antiretroviral therapy (ART) during acute HIV infection. HIV-1 specific T cell responses were measured by multicolor flow cytometry after 3-4 doses. The frequency of resting CD4+ T-cell infection (RCI) was measured by quantitative viral outgrowth assay. Participants demonstrating increased immune response postvaccination were eligible for analytic treatment interruption (ATI). AGS-004 induced a positive immune response defined as ≥2-fold increase from baseline in the number of multifunctional HIV-1 specific CD28+/CD45RA- CD8+ effector/memory cytoxic T-lymphocytes (CTLs) in all six participants. All participants underwent ATI with rebound viremia at a median of 29 days. Immune correlates between time to viral rebound and the induction of effector CTLs were determined. Baseline RCI was low in most participants (0.043-0.767 IUPM). One participant had a >2-fold decrease (0.179-0.067 infectious units per million [IUPM]) in RCI at week 10. One participant with the lowest RCI had the longest ATI. AGS-004 dendritic cell administration increased multifunctional HIV-specific CD28+/CD45RA- CD8+ memory T cell responses in all participants, but did not permit sustained ART interruption. However, greater expansion of CD28-/CCR7-/CD45RA- CD8+ effector T cell responses correlated with a longer time to viral rebound. AGS-004 may be a useful tool to augment immune responses in the setting of latency reversal and eradication strategies.


Assuntos
Células Dendríticas/imunologia , Infecções por HIV/imunologia , Infecções por HIV/terapia , Imunogenicidade da Vacina , Imunoterapia/métodos , Doença Aguda , Adolescente , Adulto , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Feminino , HIV-1 , Humanos , Masculino , Pessoa de Meia-Idade , RNA Viral , Carga Viral , Viremia , Adulto Jovem
17.
EBioMedicine ; 23: 52-58, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28803740

RESUMO

Latently human immunodeficiency virus (HIV)-infected cells are transcriptionally quiescent and invisible to clearance by the immune system. To demonstrate that the latency reversing agent vorinostat (VOR) induces a window of vulnerability in the latent HIV reservoir, defined as the triggering of viral antigen production sufficient in quantity and duration to allow for recognition and clearance of persisting infection, we developed a latency clearance assay (LCA). The LCA is a quantitative viral outgrowth assay (QVOA) that includes the addition of immune effectors capable of clearing cells expressing viral antigen. Here we show a reduction in the recovery of replication-competent virus from VOR exposed resting CD4 T cells following addition of immune effectors for a discrete period. TAKE HOME MESSAGE: VOR exposure leads to sufficient production of viral protein on the cell surface, creating a window of vulnerability within this latent reservoir in antiretroviral therapy (ART)-suppressed HIV-infected individuals that allows the clearance of latently infected cells by an array of effector mechanisms.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Ácidos Hidroxâmicos/farmacologia , Latência Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Antígenos Virais/imunologia , Terapia Antirretroviral de Alta Atividade , Contagem de Linfócito CD4 , Relação CD4-CD8 , Linfócitos T CD8-Positivos/metabolismo , Infecções por HIV/tratamento farmacológico , Antígenos HLA/imunologia , Humanos , Peptídeos/imunologia , Carga Viral , Latência Viral/imunologia , Replicação Viral/imunologia , Vorinostat
18.
J Clin Invest ; 127(8): 3126-3135, 2017 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-28714868

RESUMO

BACKGROUND: The histone deacetylase (HDAC) inhibitor vorinostat (VOR) can increase HIV RNA expression in vivo within resting CD4+ T cells of aviremic HIV+ individuals. However, while studies of VOR or other HDAC inhibitors have reported reversal of latency, none has demonstrated clearance of latent infection. We sought to identify the optimal dosing of VOR for effective serial reversal of HIV latency. METHODS: In a study of 16 HIV-infected, aviremic individuals, we measured resting CD4+ T cell-associated HIV RNA ex vivo and in vivo following a single exposure to VOR, and then in vivo after a pair of doses separated by 48 or 72 hours, and finally following a series of 10 doses given at 72-hour intervals. RESULTS: Serial VOR exposures separated by 72 hours most often resulted in an increase in cell-associated HIV RNA within circulating resting CD4+ T cells. VOR was well tolerated by all participants. However, despite serial reversal of latency over 1 month of VOR dosing, we did not observe a measurable decrease (>0.3 log10) in the frequency of latent infection within resting CD4+ T cells. CONCLUSIONS: These findings outline parameters for the experimental use of VOR to clear latent infection. Latency reversal can be achieved by VOR safely and repeatedly, but effective depletion of persistent HIV infection will require additional advances. In addition to improvements in latency reversal, these advances may include the sustained induction of potent antiviral immune responses capable of recognizing and clearing the rare cells in which HIV latency has been reversed. TRIAL REGISTRATION: Clinicaltrials.gov NCT01319383. FUNDING: NIH grants U01 AI095052, AI50410, and P30 CA016086 and National Center for Advancing Translational Sciences grant KL2 TR001109.


Assuntos
Linfócitos T CD4-Positivos/efeitos dos fármacos , Esquema de Medicação , Infecções por HIV/tratamento farmacológico , Inibidores de Histona Desacetilases/administração & dosagem , Ácidos Hidroxâmicos/administração & dosagem , Latência Viral/efeitos dos fármacos , Adulto , Idoso , Fármacos Anti-HIV/farmacologia , Feminino , HIV-1/fisiologia , Humanos , Masculino , Pessoa de Meia-Idade , RNA Viral/análise , Fatores de Tempo , Resultado do Tratamento , Ativação Viral , Vorinostat
19.
AIDS ; 30(18): 2815-2822, 2016 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-27662549

RESUMO

BACKGROUND: Updated guidelines recommend immediate antiretroviral treatment (ART) during acute HIV infection (AHI), but efficacy data on regimens during AHI are limited. METHODS: We provide final data on a prospective, single-arm 96-week open-label study of once-daily emtricitabine/tenofovir/efavirenz initiated during AHI. The primary endpoint was the proportion of responders with HIV RNA less than 200 copies/ml by week 24. We examined time to viral suppression, retention, and CD8 cell activation through week 96 in relation to baseline characteristics. RESULTS: Between January 2005 and December 2011, 92 AHI participants enrolled. Most participants (78%) were men who have sex with men (MSM), and 42% were young MSM (18-25 years of age). Two participants withdrew leaving 90 patients for analysis. Eighty-one (90%) remained on therapy and achieved viral suppression to less than 200 copies/ml by week 24, and 71 (79%) to less than 50 copies/ml at week 48. The median time from ART initiation to suppression less than 200 copies/ml was 65 days (range 7-523) and to less than 50 copies/ml was 105 days (range 14-523). The frequency of immune activation declined from a median of 67% to 16% through week 96. Retention on study was maintained in 92% of participants at week 48 and in 83% through week 96. Among 75 participants retained through week 96, 92% were suppressed to less than 50 copies/ml. Among 39 young MSM, 79% completed a week 96 visit and 67% were suppressed at week 96. CONCLUSION: ART during AHI resulted in rapid and sustained viral suppression with high rates of retention in care and on ART in this cohort including a large proportion of young MSM.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Benzoxazinas/administração & dosagem , Emtricitabina/administração & dosagem , Infecções por HIV/tratamento farmacológico , Tenofovir/administração & dosagem , Adolescente , Adulto , Idoso , Alcinos , Linfócitos T CD8-Positivos/imunologia , Ciclopropanos , Combinação de Medicamentos , Feminino , Humanos , Masculino , Adesão à Medicação , Pessoa de Meia-Idade , Estudos Prospectivos , RNA Viral/sangue , Resposta Viral Sustentada , Resultado do Tratamento , Carga Viral , Adulto Jovem
20.
J Acquir Immune Defic Syndr ; 71(1): 111-9, 2016 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-26761274

RESUMO

OBJECTIVE: To describe demographic and behavioral characteristics of persons with acute HIV infection (AHI) over time. METHODS: We conducted a retrospective assessment of AHI identified through the Screening and Tracing Active Transmission (STAT) program from 2003 to 2012 in North Carolina (NC). AHI was identified using pooled nucleic acid amplification for antibody negative samples and individual HIV-1 RNA for antibody indeterminate samples. The STAT program provides rapid notification and evaluation. We compared STAT-collected demographic and risk characteristics with all persons requesting tests and all non-AHI diagnoses from the NC State Laboratory of Public Health. RESULTS: The STAT Program identified 236 AHI cases representing 3.4% (95% confidence interval: 3.0% to 3.9%) of all HIV diagnoses. AHI cases were similar to those diagnosed during established HIV. On pretest risk-assessments, AHI cases were predominately black (69.1%), male (80.1%), young (46.8% < 25 years), and men who have sex with men (MSM) (51.7%). Per postdiagnosis interviews, the median age decreased from 35 (interquartile range 25-42) to 27 (interquartile range 22-37) years, and the proportion <25 years increased from 23.8% to 45.2% (trend P = 0.04) between 2003 and 2012. AHI men were more likely to report MSM risk post-diagnosis than on pretest risk-assessments (64%-82.9%; P < 0.0001). Post-diagnosis report of MSM risk in men with AHI increased from 71.4% to 96.2%. CONCLUSIONS: In NC, 3.4% of individuals diagnosed with HIV infection have AHI. AHI screening provides a real-time source of incidence trends, improves the diagnostic yield of HIV testing, and offers an opportunity to limit onward transmission.


Assuntos
Infecções por HIV/epidemiologia , Programas de Rastreamento/estatística & dados numéricos , Doença Aguda , Adolescente , Adulto , Infecções por HIV/diagnóstico , HIV-1 , Homossexualidade Masculina/estatística & dados numéricos , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , North Carolina/epidemiologia , Estudos Retrospectivos , Fatores de Risco , Adulto Jovem
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