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1.
Top Antivir Med ; 27(3): 111-121, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31634862

RESUMO

The 2019 edition of the IAS-USA drug resistance mutations list updates the Figure last published in January 2017. The mutations listed are those that have been identified by specific criteria for evidence and drugs described. The Figure is designed to assist practitioners in identifying key mutations associated with resistance to antiretroviral drugs, and therefore, in making clinical decisions regarding antiretroviral therapy.

2.
PLoS Comput Biol ; 15(4): e1006849, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30978183

RESUMO

Quantitative viral outgrowth assays (QVOA) use limiting dilutions of CD4+ T cells to measure the size of the latent HIV-1 reservoir, a major obstacle to curing HIV-1. Efforts to reduce the reservoir require assays that can reliably quantify its size in blood and tissues. Although QVOA is regarded as a "gold standard" for reservoir measurement, little is known about its accuracy and precision or about how cell storage conditions or laboratory-specific practices affect results. Owing to this lack of knowledge, confidence intervals around reservoir size estimates-as well as judgments of the ability of therapeutic interventions to alter the size of the replication-competent but transcriptionally inactive latent reservoir-rely on theoretical statistical assumptions about dilution assays. To address this gap, we have carried out a Bayesian statistical analysis of QVOA reliability on 75 split samples of peripheral blood mononuclear cells (PBMC) from 5 antiretroviral therapy (ART)-suppressed participants, measured using four different QVOAs at separate labs, estimating assay precision and the effect of frozen cell storage on estimated reservoir size. We found that typical assay results are expected to differ from the true value by a factor of 1.6 to 1.9 up or down. Systematic assay differences comprised a 24-fold range between the assays with highest and lowest scales, likely reflecting differences in viral outgrowth readout and input cell stimulation protocols. We also found that controlled-rate freezing and storage of samples did not cause substantial differences in QVOA compared to use of fresh cells (95% probability of < 2-fold change), supporting continued use of frozen storage to allow transport and batched analysis of samples. Finally, we simulated an early-phase clinical trial to demonstrate that batched analysis of pre- and post-therapy samples may increase power to detect a three-fold reservoir reduction by 15 to 24 percentage points.


Assuntos
Infecções por HIV/virologia , HIV-1 , Carga Viral/métodos , Latência Viral , Fármacos Anti-HIV/uso terapêutico , Teorema de Bayes , Linfócitos T CD4-Positivos/virologia , Biologia Computacional , Simulação por Computador , Infecções por HIV/tratamento farmacológico , HIV-1/fisiologia , Humanos , Leucócitos Mononucleares/virologia , Funções Verossimilhança , Cadeias de Markov , Método de Monte Carlo , Reprodutibilidade dos Testes , Carga Viral/estatística & dados numéricos , Replicação Viral
3.
J Biol Chem ; 294(14): 5576-5589, 2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30745362

RESUMO

Histone deacetylase (HDAC) inhibitors (HDACis) have been widely tested in clinical trials for their ability to reverse HIV latency but have yielded only limited success. One HDACi, suberoylanilide hydroxamic acid (SAHA), exhibits off-target effects on host gene expression predicted to interfere with induction of HIV transcription. Romidepsin (RMD) has higher potency and specificity for class I HDACs implicated in maintaining HIV provirus in the latent state. More robust HIV reactivation has indeed been achieved with RMD use ex vivo than with SAHA; however, reduction of viral reservoir size has not been observed in clinical trials. Therefore, using RNA-Seq, we sought to compare the effects of SAHA and RMD on gene expression in primary CD4+ T cells. Among the genes whose expression was modulated by both HDACi agents, we identified genes previously implicated in HIV latency. Two genes, SMARCB1 and PARP1, whose modulation by SAHA and RMD is predicted to inhibit HIV reactivation, were evaluated in the major maturation subsets of CD4+ T cells and were consistently either up- or down-regulated by both HDACi compounds. Our results indicate that despite having different potencies and HDAC specificities, SAHA and RMD modulate an overlapping set of genes, implicated in HIV latency regulation. Some of these genes merit exploration as additional targets to improve the therapeutic outcomes of "shock and kill" strategies. The overall complexity of HDACi-induced responses among host genes with predicted stimulatory or inhibitory effects on HIV expression likely contributes to differential HDACi potencies and dictates the outcome of HIV reactivation.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Depsipeptídeos/farmacologia , HIV-1/fisiologia , Inibidores de Histona Desacetilases/farmacologia , Ativação Viral/efeitos dos fármacos , Vorinostat/farmacologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Poli(ADP-Ribose) Polimerase-1/biossíntese , Proteína SMARCB1/biossíntese , Transcrição Genética/efeitos dos fármacos , Latência Viral/efeitos dos fármacos
4.
Retrovirology ; 16(1): 4, 2019 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-30770748

RESUMO

Latently infected CD4 lymphocytes preclude cure of HIV infection, even with the most effective antiretroviral therapy. The replication competent latent HIV reservoir has been quantified with the terminal dilution quantitative viral outgrowth assay, which induces virus propagation in CD4+ T cell culture supernatants following cellular activation. Efforts to improve the sensitivity of this inefficient assay have introduced more sensitive p24 ELISA and RNA PCR based endpoints, but these more sensitive endpoints have raised the question whether they are measuring induced replication competent or defective virions. Here we performed parallel terminal dilution assays with CD4 lymphocytes from subjects effectively treated with antiretroviral therapy. An HIV integrase inhibitor was incorporated into one set of parallel cultures to compare the frequency of cells that can be induced to produce virions to those that produce virus that can propagate and amplify with co-culture in permissive cells. The majority of cells that can be induced to generate virus particles are producing replication competent virus, thus justifying more sensitive and faster assays of this reservoir.


Assuntos
Linfócitos T CD4-Positivos/virologia , Infecções por HIV/virologia , HIV/fisiologia , Carga Viral/métodos , Ativação Viral , Latência Viral , Replicação Viral , Antirretrovirais/uso terapêutico , Infecções por HIV/tratamento farmacológico , Humanos
6.
EBioMedicine ; 36: 113-121, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30316868

RESUMO

BACKGROUND: Improved assays are critical to better characterize the HIV reservoir and to reliably evaluate candidate intervention strategies. Here we describe different methods to quantify the HIV reservoir. METHODS: We developed an optimized quantitative viral outgrowth assay (QVOA) to quantify the frequency of cells harboring replication-competent HIV, which is simpler and more sensitive than classical QVOAs. We also developed new inducible RNA assays that concomitantly measure the frequency of cell-associated [ca-] (gag and tat-rev) and cell-free [cf-] HIV RNA after three days of anti-CD3/CD28 stimulation. FINDINGS: The median frequency of the infected cells measured after induction was 94 IQR[60-132], 16 IQR [9-29] and 2.9 IQR[1.9-6.8] cells/106 CD4+ T-cells for ca-RNA gag and tat-rev, and cf-RNA, respectively. There are a large proportion of transcription-competent proviruses (ca-RNA) that seemed unable to form complete virions (cf-RNA), suggesting post-transcriptional blocks or defective proviruses. Importantly, the median frequency of infected CD4+ T-cells as estimated by 3-day inducible cf-RNA assay was not statistically different from the frequency measured by the QVOA (median of 3.3 [1.9-6.2] IUPM). The latently infected cells detected by the inducible cf-RNA assay correlated highly with the QVOA ( r= 0.67, p < .001), and both assays were equivalent in 60% of the samples tested, suggesting that most cells induced to produce virions are generating replication-competent virus. INTERPRETATION: These inducible RNA assays provide more sensitivity and a greater dynamic range for the monitoring of reduction of the reservoir by eradication strategies. Such assays may serve as robust and useful tools for clinical investigations of the HIV reservoir.

7.
J Clin Microbiol ; 56(12)2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30232127

RESUMO

We utilized pulsed-field gel electrophoresis (PFGE) to purify high-molecular-weight DNA from HIV-infected cells. This purification, in combination with our previously described droplet digital PCR (ddPCR) assay, was used to develop a method to quantify proviral integrated HIV DNA free of lower-molecular-weight species of HIV DNA. Episomal 2-long-terminal-repeat (2-LTR) circles were completely cleared from HIV DNA samples. Technical replicates of the complete assay, starting with the same specimens, resulted in no statistical differences in quantification of integrated HIV gag sequences in cellular DNA from cells from HIV-infected subjects after prolonged treatment with antiretroviral therapy (ART). The PFGE ddPCR assay was compared to the Alu-gag quantitative PCR (qPCR) assay, the most widely used assay to measure proviral integrated HIV DNA. Spearman's rho nonparametric correlation determined PFGE ddPCR results to be positively correlated with Alu-gag qPCR results (r = 0.7052; P = 0.0273). In summary, PFGE ddPCR is a sensitive, reproducible, and robust method to measure proviral integrated HIV DNA and is theoretically more accurate than previously described assays, because it is a direct measure of integrated HIV DNA.

8.
Clin Infect Dis ; 2018 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-30052811

RESUMO

Background: Contemporary antiretroviral therapies (ART) and management strategies have diminished both human immunodeficiency virus (HIV) treatment failure and the acquired resistance to drugs in resource-rich regions, but transmission of drug-resistant viruses has not similarly decreased. In low- and middle-income regions, ART roll-out has improved outcomes, but has resulted in increasing acquired and transmitted resistances. Our objective was to review resistance to ART drugs and methods to detect it, and to provide updated recommendations for testing and monitoring for drug resistance in HIV-infected individuals. Methods: A volunteer panel of experts appointed by the International Antiviral (formerly AIDS) Society-USA reviewed relevant peer-reviewed data that were published or presented at scientific conferences. Recommendations were rated according to the strength of the recommendation and quality of the evidence, and reached by full panel consensus. Results: Resistance testing remains a cornerstone of ART. It is recommended in newly-diagnosed individuals and in patients in whom ART has failed. Testing for transmitted integrase strand-transfer inhibitor resistance is currently not recommended, but this may change as more resistance emerges with widespread use. Sanger-based and next-generation sequencing approaches are each suited for genotypic testing. Testing for minority variants harboring drug resistance may only be considered if treatments depend on a first-generation nonnucleoside analogue reverse transcriptase inhibitor. Different HIV-1 subtypes do not need special considerations regarding resistance testing. Conclusions: Testing for HIV drug resistance in drug-naive individuals and in patients in whom antiretroviral drugs are failing, and the appreciation of the role of testing, are crucial to the prevention and management of failure of ART.

9.
Front Immunol ; 9: 603, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29706951

RESUMO

The greatest obstacle to a cure for HIV is the provirus that integrates into the genome of the infected cell and persists despite antiretroviral therapy. A "shock and kill" approach has been proposed as a strategy for an HIV cure whereby drugs and compounds referred to as latency-reversing agents (LRAs) are used to "shock" the silent provirus into active replication to permit "killing" by virus-induced pathology or immune recognition. The LRA most utilized to date in clinical trials has been the histone deacetylase (HDAC) inhibitor-vorinostat. Potentially, pathological off-target effects of vorinostat may result from the activation of human endogenous retroviruses (HERVs), which share common ancestry with exogenous retroviruses including HIV. To explore the effects of HDAC inhibition on HERV transcription, an unbiased pharmacogenomics approach (total RNA-Seq) was used to evaluate HERV expression following the exposure of primary CD4+ T cells to a high dose of vorinostat. Over 2,000 individual HERV elements were found to be significantly modulated by vorinostat, whereby elements belonging to the ERVL family (e.g., LTR16C and LTR33) were predominantly downregulated, in contrast to LTR12 elements of the HERV-9 family, which exhibited the greatest signal, with the upregulation of 140 distinct elements. The modulation of three different LTR12 elements by vorinostat was confirmed by droplet digital PCR along a dose-response curve. The monitoring of LTR12 expression during clinical trials with vorinostat may be indicated to assess the impact of this HERV on the human genome and host immunity.

10.
J Virol ; 92(3)2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29142136

RESUMO

Residual viremia is common during antiretroviral therapy (ART) and could be caused by ongoing low-level virus replication or by release of viral particles from infected cells. ART intensification should impact ongoing viral propagation but not virion release. Eighteen acutely infected men were enrolled in a randomized controlled trial and monitored for a median of 107 weeks. Participants started ART with (n = 9) or without (n = 9) intensification with maraviroc (MVC) within 90 days of infection. Levels of HIV DNA and cell-free RNA were quantified by droplet digital PCR. Deep sequencing of C2-V3 env, gag, and pol (454 Roche) was performed on longitudinally collected plasma and peripheral blood mononuclear cell (PBMC) samples while on ART. Sequence data were analyzed for evidence of evolution by (i) molecular diversity analysis, (ii) nonparametric test for panmixia, and (iii) tip date randomization within a Bayesian framework. There was a longitudinal decay of HIV DNA after initiation of ART with no difference between MVC intensification groups (-0.08 ± 0.01 versus -0.09 ± 0.01 log10 copies/week in MVC+ versus MVC- groups; P = 0.62). All participants had low-level residual viremia (median, 2.8 RNA copies/ml). Across participants, medians of 56 (interquartile range [IQR], 36 to 74), 29 (IQR, 25 to 35), and 40 (IQR, 31 to 54) haplotypes were generated for env, gag, and pol regions, respectively. There was no clear evidence of viral evolution during ART and no difference in viral diversity or population structure from individuals with or without MVC intensification. Further efforts focusing on elucidating the mechanism(s) of viral persistence in various compartments using recent sequencing technologies are still needed, and potential low-level viral replication should always be considered in cure strategies.IMPORTANCE Residual viremia is common among HIV-infected people on ART. It remains controversial if this viremia is a consequence of propagating infection. We hypothesized that molecular evolution would be detectable during viral propagation and that therapy intensified with the entry inhibitor maraviroc would demonstrate less evolution. We performed a randomized double-blinded treatment trial with 18 acutely infected men (standard ART versus standard ART plus maraviroc). From longitudinally collected blood plasma and cells, levels of HIV DNA and cell-free HIV RNA were quantified by droplet digital PCR, and HIV DNA (env, gag, and pol coding regions) was deep sequenced (454 Roche). Investigating people who started ART during the earliest stages of their HIV infection, when viral diversity is low, provides an opportunity to detect evidence of viral evolution. Despite using a battery of analytical techniques, no clear and consistent evidence of viral propagation for over 90 weeks of observation could be discerned.


Assuntos
Antagonistas dos Receptores CCR5/uso terapêutico , Cicloexanos/uso terapêutico , Infecções por HIV/tratamento farmacológico , Triazóis/uso terapêutico , Viremia/tratamento farmacológico , Replicação Viral/efeitos dos fármacos , Adulto , Terapia Antirretroviral de Alta Atividade , Teorema de Bayes , California , DNA Viral/sangue , Método Duplo-Cego , Feminino , Infecções por HIV/virologia , HIV-1/genética , HIV-1/fisiologia , Humanos , Masculino , Maraviroc , RNA Viral/sangue , Carga Viral , Adulto Jovem
11.
AIDS ; 32(4): 443-450, 2018 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-29239894

RESUMO

OBJECTIVE: Given the emerging appreciation for the role of antibody-dependent effector functions and IgG subclass distribution among spontaneous controllers of HIV, we sought to determine whether antibody-associated features diverged in early HIV infection between patients who ultimately became controllers versus those who became progressors. METHODS: IgG was purified from plasma from nine acutely infected patients who subsequently controlled HIV spontaneously (controllers) and 10 acutely infected individuals who did not control viremia (progressors). Antibody profiles were compared at weeks 4, 12, 24 and 48 postinfection. Levels of clade B gp120-specific, gp140-specific and gp41-specific IgG antibody subclasses were measured. In addition, gp120-specific antibody-dependent cellular phagocytosis, rapid fluorescent antibody-dependent cellular cytotoxicity and antibody-dependent cellular viral inhibition were all assessed. RESULTS: Although no single antibody-related measurement was significantly associated with long-term HIV control, combinations of antibody-associated variables were able to accurately differentiate controllers and progressors. In contrast to controllers, progressors showed greater dynamic changes in gp120-specific subclass selection profiles, with increasing levels of Env-specific IgG2 antibodies and losses in Env-specific IgG3 antibodies. Moreover, progressors, but not controllers, lost antibody-dependent cellular viral inhibition function over time. Together, these results highlight changes in IgG subclass selection profiles in progressive, but not controlled, HIV infection. CONCLUSION: This study suggests that the temporal variation and maintenance of Env-specific IgG subclasses during acute HIV infection are predictive of eventual disease control. The maintenance of gp120-specific and gp140-specific IgG3 may contribute to control of disease in spontaneous controllers. Thus, strategies to induce stable IgG3 responses may preserve control of the viral reservoir.

12.
Lancet Infect Dis ; 18(2): 188-197, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29158101

RESUMO

BACKGROUND: Antiretroviral therapy (ART) that enables suppression of HIV replication has been successfully rolled out at large scale to HIV-positive patients in low-income and middle-income countries. WHO guidelines for these regions define failure of ART with a lenient threshold of viraemia (HIV RNA viral load ≥1000 copies per mL). We investigated the occurrence of detectable viraemia during ART below this threshold and its effect on treatment outcomes in a large South African cohort. METHODS: In this observational cohort study, we included HIV-positive adults registered between Jan 1, 2007, and May 1, 2016, at 57 clinical sites in South Africa, who were receiving WHO-recommended ART regimens and viral load monitoring. Low-level viraemia was defined as the occurrence of at least one viral load measurement of 51-999 copies per mL during ART. Outcomes were WHO-defined virological failure (one or more viral load measurement of ≥1000 copies per mL) and switch to second-line ART. Risks were estimated with Cox proportional hazard models. FINDINGS: 70 930 patients were included in the analysis, of whom 67 644 received first-line ART, 1476 received second-line ART, and 1810 received both. Median duration of follow-up was 124 weeks (IQR 56-221) for patients on first-line ART and 101 weeks (IQR 51-178) for patients on second-line ART. Low-level viraemia occurred in 16 013 (23%) of 69 454 patients, with an incidence of 11·5 per 100 person-years of follow-up (95% CI 11·4-11·7), during first-line ART. Virological failure during follow-up occurred in 14 380 (22%) of 69 454 patients on first-line ART. Low-level viraemia was associated with increased hazards of virological failure (hazard ratio [HR] 2·6, 95% CI 2·5-2·8; p<0·0001) and switch to second-line ART (HR 5·2, 4·4-6·1; p<0·0001]) compared with virological suppression of less than 50 copies per mL. Risk of virological failure increased further with higher ranges and persistence of low-level viraemia. INTERPRETATION: In this large cohort, low-level viraemia occurred frequently and increased the risk of virological failure and switch to second-line ART. Strategies for management of low-level viraemia need to be incorporated into WHO guidelines to meet UNAIDS-defined targets aimed at halting the global HIV epidemic. FUNDING: None.

13.
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
14.
PLoS Med ; 14(11): e1002417, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29112956

RESUMO

BACKGROUND: It is unknown if extremely early initiation of antiretroviral therapy (ART) may lead to long-term ART-free HIV remission or cure. As a result, we studied 2 individuals recruited from a pre-exposure prophylaxis (PrEP) program who started prophylactic ART an estimated 10 days (Participant A; 54-year-old male) and 12 days (Participant B; 31-year-old male) after infection with peak plasma HIV RNA of 220 copies/mL and 3,343 copies/mL, respectively. Extensive testing of blood and tissue for HIV persistence was performed, and PrEP Participant A underwent analytical treatment interruption (ATI) following 32 weeks of continuous ART. METHODS AND FINDINGS: Colorectal and lymph node tissues, bone marrow, cerebral spinal fluid (CSF), plasma, and very large numbers of peripheral blood mononuclear cells (PBMCs) were obtained longitudinally from both participants and were studied for HIV persistence in several laboratories using molecular and culture-based detection methods, including a murine viral outgrowth assay (mVOA). Both participants initiated PrEP with tenofovir/emtricitabine during very early Fiebig stage I (detectable plasma HIV-1 RNA, antibody negative) followed by 4-drug ART intensification. Following peak viral loads, both participants experienced full suppression of HIV-1 plasma viremia. Over the following 2 years, no further HIV could be detected in blood or tissue from PrEP Participant A despite extensive sampling from ileum, rectum, lymph nodes, bone marrow, CSF, circulating CD4+ T cell subsets, and plasma. No HIV was detected from tissues obtained from PrEP Participant B, but low-level HIV RNA or DNA was intermittently detected from various CD4+ T cell subsets. Over 500 million CD4+ T cells were assayed from both participants in a humanized mouse outgrowth assay. Three of 8 mice infused with CD4+ T cells from PrEP Participant B developed viremia (50 million input cells/surviving mouse), but only 1 of 10 mice infused with CD4+ T cells from PrEP Participant A (53 million input cells/mouse) experienced very low level viremia (201 copies/mL); sequence confirmation was unsuccessful. PrEP Participant A stopped ART and remained aviremic for 7.4 months, rebounding with HIV RNA of 36 copies/mL that rose to 59,805 copies/mL 6 days later. ART was restarted promptly. Rebound plasma HIV sequences were identical to those obtained during acute infection by single-genome sequencing. Mathematical modeling predicted that the latent reservoir size was approximately 200 cells prior to ATI and that only around 1% of individuals with a similar HIV burden may achieve lifelong ART-free remission. Furthermore, we observed that lymphocytes expressing the tumor marker CD30 increased in frequency weeks to months prior to detectable HIV-1 RNA in plasma. This study was limited by the small sample size, which was a result of the rarity of individuals presenting during hyperacute infection. CONCLUSIONS: We report HIV relapse despite initiation of ART at one of the earliest stages of acute HIV infection possible. Near complete or complete loss of detectable HIV in blood and tissues did not lead to indefinite ART-free HIV remission. However, the small numbers of latently infected cells in individuals treated during hyperacute infection may be associated with prolonged ART-free remission.


Assuntos
Antirretrovirais/uso terapêutico , Biomarcadores/análise , Infecções por HIV/tratamento farmacológico , HIV-1 , Adulto , Citometria de Fluxo , Infecções por HIV/imunologia , Infecções por HIV/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Estudos Prospectivos , Recidiva , Prevenção Secundária , Resultado do Tratamento
15.
Curr Opin HIV AIDS ; 12(5): 423-424, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28771448
16.
J Clin Microbiol ; 55(8): 2544-2553, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28592548

RESUMO

The search for a cure for HIV infection has highlighted the need for increasingly sensitive and precise assays to measure viral burden in various tissues and body fluids. We describe the application of a standardized assay for HIV-1 RNA in multiple specimen types. The fully automated Aptima HIV-1 Quant Dx assay (Aptima assay) is FDA cleared for blood plasma HIV-1 RNA quantitation. In this study, the Aptima assay was applied for the quantitation of HIV RNA in peripheral blood mononuclear cells (PBMCs; n = 72), seminal plasma (n = 20), cerebrospinal fluid (CSF; n = 36), dried blood spots (DBS; n = 104), and dried plasma spots (DPS; n = 104). The Aptima assay was equivalent to or better than commercial assays or validated in-house assays for the quantitation of HIV RNA in CSF and seminal plasma. For PBMC specimens, the sensitivity of the Aptima assay in the detection of HIV RNA decayed as background uninfected PBMC counts increased; proteinase K treatment demonstrated some benefit in restoring signal at higher levels of background PBMCs. Finally, the Aptima assay yielded 100% detection rates of DBS in participants with plasma HIV RNA levels of ≥35 copies/ml and 100% detection rates of DPS in participants with plasma HIV RNA levels of ≥394 copies/ml. The Aptima assay can be applied to a variety of specimens from HIV-infected subjects to measure HIV RNA for studies of viral persistence and cure strategies. It can also detect HIV in dried blood and plasma specimens, which may be of benefit in resource-limited settings.


Assuntos
Automação Laboratorial/métodos , HIV-1/isolamento & purificação , RNA Viral/análise , Carga Viral/métodos , HIV-1/genética , Humanos
17.
J Virol ; 91(17)2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28615205

RESUMO

Understanding whether the neutralizing antibody (NAb) response impacts HIV-1 superinfection and how superinfection subsequently modulates the NAb response can help clarify correlates of protection from HIV exposures and better delineate pathways of NAb development. We examined associations between the development of NAb and the occurrence of superinfection in a well-characterized, antiretroviral therapy (ART)-naive, primary infection cohort of men who have sex with men. Deep sequencing was applied to blood plasma samples from the cohort to detect cases of superinfection. We compared the NAb activity against autologous and heterologous viruses between 10 participants with intrasubtype B superinfection and 19 monoinfected controls, matched to duration of infection and risk behavior. Three to 6 months after primary infection, individuals who would later become superinfected had significantly weaker NAb activity against tier 1 subtype B viruses (P = 0.003 for SF-162 and P = 0.017 for NL4-3) and marginally against autologous virus (P = 0.054). Lower presuperinfection NAb responses correlated with weaker gp120 binding and lower plasma total IgG titers. Soon after superinfection, the NAb response remained lower, but between 2 and 3 years after primary infection, NAb levels strengthened and reached those of controls. Superinfecting viruses were typically not susceptible to neutralization by presuperinfection plasma. These observations suggest that recently infected individuals with a delayed NAb response against primary infecting and tier 1 subtype B viruses are more susceptible to superinfection.IMPORTANCE Our findings suggest that within the first year after HIV infection, a relatively weak neutralizing antibody response against primary and subtype-specific neutralization-sensitive viruses increases susceptibility to superinfection in the face of repeated exposures. As natural infection progresses, the immune response strengthens significantly in some superinfected individuals. These findings will inform HIV vaccine design by providing testable correlates of protection from initial HIV infection.


Assuntos
Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/classificação , Superinfecção/imunologia , Adulto , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , California , Estudos de Casos e Controles , Anticorpos Anti-HIV/sangue , Proteína gp120 do Envelope de HIV/imunologia , Infecções por HIV/virologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Masculino , Testes de Neutralização , Superinfecção/virologia , Adulto Jovem
18.
Virology ; 508: 127-133, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28527342

RESUMO

Quantification of cell-associated replication-competent HIV, in blood samples from patients with undetectable plasma viremia, requires specialized culture conditions that include exogenous pan T cell stimulation. Different research groups have used several stimuli for this purpose; however, the relative efficacies of these T cell stimuli to induce productive HIV replication from latently infected cells ex vivo have not been systematically evaluated. To this end, we compared four commonly used T cell stimuli: 1) irradiated allogeneic cells plus phytohaemagglutinin (PHA); 2) PHA alone; 3) phorbol myristate acetate plus Ionomycin; and 4) immobilized αCD3 plus αCD28 antibodies. End-point dilutions of patient CD4 T cells were performed, using virion RNA production to quantify HIV induction. Our results demonstrated that these activation approaches were not equivalent and that antibody cross-linking of CD3 and CD28 membrane receptors was the most effective means to activate HIV replication from a resting cell state, closely followed by stimulation with irradiated allogeneic cells plus PHA.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/virologia , HIV-1/fisiologia , Ativação Linfocitária , Latência Viral , Replicação Viral , Adulto , Fármacos Anti-HIV/uso terapêutico , Linfócitos T CD4-Positivos/virologia , Células Cultivadas , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , HIV-1/genética , Humanos , Masculino , Ativação Viral
19.
Nat Med ; 23(5): 638-643, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28414330

RESUMO

Despite years of fully suppressive antiretroviral therapy (ART), HIV persists in its hosts and is never eradicated. One major barrier to eradication is that the virus infects multiple cell types that may individually contribute to HIV persistence. Tissue macrophages are critical contributors to HIV pathogenesis; however, their specific role in HIV persistence during long-term suppressive ART has not been established. Using humanized myeloid-only mice (MoM), we demonstrate that HIV infection of tissue macrophages is rapidly suppressed by ART, as reflected by a rapid drop in plasma viral load and a dramatic decrease in the levels of cell-associated viral RNA and DNA. No viral rebound was observed in the plasma of 67% of the ART-treated animals at 7 weeks after ART interruption, and no replication-competent virus was rescued from the tissue macrophages obtained from these animals. In contrast, in a subset of animals (∼33%), a delayed viral rebound was observed that is consistent with the establishment of persistent infection in tissue macrophages. These observations represent the first direct evidence, to our knowledge, of HIV persistence in tissue macrophages in vivo.


Assuntos
Infecções por HIV/virologia , HIV-1/fisiologia , Macrófagos/virologia , Animais , Fármacos Anti-HIV/uso terapêutico , Terapia Antirretroviral de Alta Atividade , Medula Óssea , DNA Viral , Eletroforese em Gel de Campo Pulsado , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Transplante de Células-Tronco Hematopoéticas , Humanos , Imuno-Histoquímica , Lactonas , Leucócitos Mononucleares , Fígado , Macrófagos Alveolares/virologia , Camundongos , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares , Fenóis , RNA Viral , Baço , Linfócitos T , Carga Viral , Latência Viral , Replicação Viral
20.
J Infect Dis ; 215(10): 1506-1513, 2017 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-28419276

RESUMO

Background.: Investigations into which human immunodeficiency virus type 1 (HIV-1) sequence features may be selected for transmission during sexual exposure have been hampered by the small number of characterized transmission pairs in individual studies. Methods.: To boost statistical power to detect differences in glycosylation, length, and electrical charge in the HIV-1 V1-V4 coding region, we reanalyzed all available 2485 env sequences derived from 114 subjects representing 58 transmission pairs from previous studies using mixed-effects linear regression and an approach to approximate the unobserved transmitted virus. Results.: The recipient partner had a shorter V1-V4 region and fewer potential N-linked glycosylation sites (PNGS) than sequences from the source partner. We also detected a trend toward more PNGS and lower isoelectric points in transmitted sequences with source partner and the evolutionary tendency to shorten V1-V4 sequences, reduce the number of PNGS, and lower isoelectric points in the recipient following transmission. Conclusions.: By using all available well-characterized env sequences from transmission pairs via sexual exposure, we were able to identify several important virologic factors that may be important in the development of biomedical preventive interventions.


Assuntos
Proteína gp120 do Envelope de HIV/genética , Infecções por HIV/transmissão , Infecções por HIV/virologia , HIV-1/genética , Fragmentos de Peptídeos/genética , Análise de Variância , Evolução Molecular , Glicosilação , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/metabolismo , HIV-1/patogenicidade , Humanos
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