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1.
Alcohol Clin Exp Res ; 2021 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-34773280

RESUMO

BACKGROUND: Alcohol, insulin resistance (IR), and hepatitis C (HCV) are all significant contributors to adverse outcomes of chronic liver disease. Latinos are disproportionately affected by these risk factors. We investigated the relationship between alcohol use and insulin action in a prospective cohort of Latino individuals with and without HCV. METHODS: One hundred fifty-three nondiabetic Latino individuals (60 HCV+, 93 HCV-) underwent clinical evaluation and metabolic testing; 56 had repeat testing over a median follow-up of 1.5 years. Peripheral IR and hepatic IR were measured via steady-state plasma glucose (SSPG) and endogenous glucose production during a two-step, 240-min insulin suppression test. Insulin secretion (IS) was measured using the graded glucose infusion test. Alcohol use was categorized as none, moderate (≤1 drink/day for women and ≤2 drinks/day for men), and heavy (>moderate). Multivariable models including HCV status assessed associations of alcohol use with baseline SSPG, hepatic IR and IS, and changes in these parameters over time. RESULTS: Overall, the median age was 44 years, 63.4% were male, 66.7% overweight/ obese, and 31.9% had heavy lifetime alcohol use while 60.4% had moderate lifetime alcohol use. SSPG and IS were similar by levels of alcohol use at baseline and alcohol use was not statistically significantly associated with change in these measures over time. However, lifetime daily heavy alcohol use (vs. not heavy, coef 2.4 µU-mg/kg-min-ml, p = 0.04) and HCV status (coef 4.4 µU-mg/kg-min-ml, p = 0.0003) were independently associated with higher baseline hepatic IR, and current heavy alcohol use was associated with greater change in hepatic IR in follow-up (coef 5.8 µU-mg/kg-min-ml, p = 0.03). CONCLUSIONS: In this cohort of Latino individuals, lifetime and current heavy alcohol use influenced hepatic IR and its change over time. Strategies to decrease rates of heavy alcohol use or increase abstinence along with lifestyle modification and anti-HCV therapy to reduce metabolic risk are critical to prevent adverse liver and metabolic outcomes in Latino individuals.

2.
Artigo em Inglês | MEDLINE | ID: mdl-33713954

RESUMO

New all-oral regimens for rifampin-resistant tuberculosis (RR-TB) are being scaled up globally. Measurement of drug concentrations in hair assesses long-term drug exposure. Delamanid (DLM) is likely to be a key component of future RR-TB treatment regimens, but a method to describe its quantification in hair via liquid chromatography-tandem mass spectrometry (LC-MS/MS) has not previously been described. We developed and validated a simple, fast, sensitive, and accurate LC-MS/MS method for quantifying DLM and its metabolite DM-6705 in small hair samples. We pulverized and extracted two milligrams of hair in methanol at 37 °C for two hours, and diluted 1:1 with water. A gradient elution method eluted DLM, DM-6705, and the internal standard OPC 14714 within 3 min, bringing overall analysis time to 5.5 min. The method has limits of detection (LOD) of 0.0003 ng/mg for DLM and 0.003 ng/mg for DM-6705. The established linear dynamic ranges are 0.003-2.1 ng/mg and 0.03-21 ng/mg for DLM and DM-6705, respectively. Eleven of 12 participant hair samples had concentrations within DLM's linear dynamic range, while all 12 samples had concentrations within the quantifiable range for DM-6705. The ranges of concentrations observed in these clinical samples for DLM and DM-6705 were 0.004-0.264 ng/mg hair and 0.412-12.041 ng/mg hair respectively. We demonstrate that while DLM was detected in hair at very low levels, its primary metabolite DM-6705 had levels approximately 100 times higher. Measuring DM-6705 in hair may accurately reflect long-term adherence to DLM-containing regimens for drug-resistant TB.


Assuntos
Cromatografia Líquida/métodos , Cabelo/química , Nitroimidazóis/análise , Oxazóis/análise , Espectrometria de Massas em Tandem/métodos , Humanos , Limite de Detecção , Modelos Lineares , Nitroimidazóis/uso terapêutico , Oxazóis/uso terapêutico , Reprodutibilidade dos Testes , Tuberculose , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico
3.
EBioMedicine ; 65: 103241, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33647768

RESUMO

BACKGROUND: One strategy being pursued to clear latently infected cells that persist in people living with HIV (PLWH) on antiretroviral therapy (ART) is to activate latent HIV infection with a latency reversing agent (LRA). Surrogate markers that accurately measure virus production following an LRA are needed. METHODS: We quantified cell-associated unspliced (US), multiply spliced (MS) and supernatant (SN) HIV RNA by qPCR from total and resting CD4+ T cells isolated from seven PLWH on ART before and after treatment ex vivo with different LRAs, including histone deacetylase inhibitors (HDACi). MS and plasma HIV RNA were also quantified from PLWH on ART (n-11) who received the HDACi panobinostat. FINDINGS: In total and resting CD4+ T cells from PLWH on ART, detection of US RNA was common while detection of MS RNA was infrequent. Primers used to detect MS RNA, in contrast to US RNA, bound sites of the viral genome that are commonly mutated or deleted in PLWH on ART. Following ex vivo stimulation with LRAs, we identified a strong correlation between the fold change increase in SN and MS RNA, but not the fold change increase in SN and US RNA. In PLWH on ART who received panobinostat, MS RNA was significantly higher in samples with detectable compared to non0detectable plasma HIV RNA. INTERPRETATION: Following administration of an LRA, quantification of MS RNA is more likely to reflect an increase in virion production and is therefore a better indicator of meaningful latency reversal. FUNDING: NHMRC, NIH DARE collaboratory.


Assuntos
HIV-1/genética , Splicing de RNA , RNA Viral/sangue , Latência Viral/fisiologia , Antirretrovirais/farmacologia , Antirretrovirais/uso terapêutico , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , Proliferação de Células/efeitos dos fármacos , Infecções por HIV/tratamento farmacológico , Infecções por HIV/patologia , Infecções por HIV/virologia , HIV-1/fisiologia , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Humanos , Poli-Hidroxialcanoatos/farmacologia , RNA Viral/metabolismo , Acetato de Tetradecanoilforbol/farmacologia , Vorinostat/farmacologia , Vorinostat/uso terapêutico
4.
BMC Infect Dis ; 21(1): 99, 2021 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-33482745

RESUMO

BACKGROUND: Treatment monitoring of drug-resistant tuberculosis (DR-TB) in resource-limited settings is challenging. We developed a multi-analyte assay for eleven anti-TB drugs in small hair samples as an objective metric of drug exposure. METHODS: Small hair samples were collected from participants at various timepoints during directly observed RR-TB treatment at an inpatient tertiary referral facility in South Africa (DR-TB cohort). We assessed qualitative determination (i.e., detection above limit of detection) of bedaquiline, linezolid, clofazimine, pretomanid, levofloxacin, moxifloxacin, pyrazinamide, isoniazid, ethambutol, ethionamide, and prothionamide in an LC-MS/MS index panel assay against a reference standard of inpatient treatment records. Because treatment regimens prior to hospitalization were not available, we also analyzed specificity (for all drugs except isoniazid) using an external cohort of HIV-positive patients treated for latent TB infection with daily isoniazid (HIV/LTBI cohort) in Uganda. RESULTS: Among the 57 DR-TB patients (58% with pre-XDR/XDR-TB; 70% HIV-positive) contributing analyzable hair samples, the sensitivity of the investigational assay was 94% or higher for all drugs except ethionamide (58.5, 95% confidence interval [CI], 40.7-99.9). Assay specificity was low across all tested analytes within the DR-TB cohort; conversely, assay specificity was 100% for all drugs in the HIV/LTBI cohort. CONCLUSIONS: Hair drug concentrations reflect long-term exposure, and multiple successive regimens commonly employed in DR-TB treatment may result in apparent false-positive qualitative and falsely elevated quantitative hair drug levels when prior treatment histories within the hair growth window are not known.


Assuntos
Antituberculosos/análise , Monitoramento de Medicamentos/métodos , Cabelo/química , Rifampina/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Adulto , Antituberculosos/uso terapêutico , Cromatografia Líquida , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mycobacterium tuberculosis/efeitos dos fármacos , Sensibilidade e Especificidade , Espectrometria de Massas em Tandem , Tuberculose/tratamento farmacológico
5.
J Infect Dis ; 224(7): 1209-1218, 2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-32147687

RESUMO

BACKGROUND: Evaluations of human immunodeficiency virus (HIV) curative interventions require reliable and efficient quantification of replication-competent latent reservoirs. The "classic" quantitative viral outgrowth assay (QVOA) has been regarded as the reference standard, although prohibitively resource and labor intensive. We compared 6 "next-generation" viral outgrowth assays, using polymerase chain reaction or ultrasensitive p24 to assess their suitability as scalable proxies for QVOA. METHODS: Next-generation QVOAs were compared with classic QVOA using single leukapheresis-derived samples from 5 antiretroviral therapy-suppressed HIV-infected participants and 1 HIV-uninfected control; each laboratory tested blinded batches of 3 frozen and 1 fresh sample. Markov chain Monte Carlo methods estimated extra-Poisson variation at aliquot, batch, and laboratory levels. Models also estimated the effect of testing frozen versus fresh samples. RESULTS: Next-generation QVOAs had similar estimates of variation to QVOA. Assays with ultrasensitive readout reported higher infectious units per million values than classic QVOA. Within-batch testing had 2.5-fold extra-Poisson variation (95% credible interval [CI], 2.1-3.5-fold) for next-generation assays. Between-laboratory variation increased extra-Poisson variation to 3.4-fold (95% CI, 2.6-5.4-fold). Frozen storage did not substantially alter infectious units per million values (-18%; 95% CI, -52% to 39%). CONCLUSIONS: The data offer cautious support for use of next-generation QVOAs as proxies for more laborious QVOA, while providing greater sensitivities and dynamic ranges. Measurement of latent reservoirs in eradication strategies would benefit from high throughput and scalable assays.

6.
J Clin Microbiol ; 58(12)2020 11 18.
Artigo em Inglês | MEDLINE | ID: mdl-32967900

RESUMO

Detection of residual plasma viremia in antiretroviral therapy (ART)-suppressed HIV-infected individuals is critical for characterizing the latent reservoir and evaluating the impact of cure interventions. Ultracentrifugation-based single-copy assays are sensitive but labor intensive. Fully automated replicate testing using a standard clinical viral load assay was evaluated as a high-throughput alternative for the quantification of low-level viremia. Four plasma samples from blood donors with acute HIV-1 infection and one viral culture supernatant were serially diluted into 25-ml samples to nominal viral loads ranging from 39 to <0.5 copies (cp)/ml. Each dilution was tested with 45 replicates (reps) using 0.5 ml/rep with the Aptima HIV-1 Quant assay. The nominal and estimated viral loads based on the single-hit Poisson model were compared, and a hybrid Poisson digital model for calibrated viral load estimation was derived. Testing performed using 45 reps on longitudinal plasma samples from 50 ART-suppressed individuals in the Reservoir Assay Validation and Evaluation Network (RAVEN) study cohort (range of 1 to 19 years of continuous ART suppression) showed a median viral load of 0.54 cp/ml (interquartile range [IQR], 0.22 to 1.46 cp/ml) and a 14% (95% confidence interval [CI], 9% to 19%) decline in viral load for each additional year in duration suppressed. Within the RAVEN cohort, the expected false-negative rate for detection at lower rep numbers using 9 and 18 reps was 26% and 14%, respectively. Residual plasma viremia levels positively correlated with cell-associated HIV RNA and DNA. The performance characteristics of the replicate Aptima assay support its use for quantifying residual plasma viremia to study the latent HIV reservoir and cure interventions.


Assuntos
Infecções por HIV , HIV-1 , Terapia Antirretroviral de Alta Atividade , Infecções por HIV/diagnóstico , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Humanos , RNA Viral , Carga Viral , Viremia/diagnóstico , Viremia/tratamento farmacológico , Latência Viral
7.
JAMA ; 323(21): 2151-2159, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32484533

RESUMO

Importance: Reducing cesarean delivery rates in the US is an important public health goal; despite evidence of the safety of vaginal birth after cesarean delivery, most women have scheduled repeat cesarean deliveries. A decision support tool could help increase trial-of-labor rates. Objective: To analyze the effect of a patient-centered decision support tool on rates of trial of labor and vaginal birth after cesarean delivery and decision quality. Design, Setting, and Participants: Multicenter, randomized, parallel-group clinical trial conducted in Boston, Chicago, and the San Francisco Bay area. A total of 1485 English- or Spanish-speaking women with 1 prior cesarean delivery and no contraindication to trial of labor were enrolled between January 2016 and January 2019; follow-up was completed in June 2019. Interventions: Participants were randomized to use a tablet-based decision support tool prior to 25 weeks' gestation (n=742) or to receive usual care (without the tool) (n=743). Main Outcomes and Measures: The primary outcome was trial of labor; vaginal birth was the main secondary outcome. Other secondary outcomes focused on maternal and neonatal outcomes and decision quality. Results: Among 1485 patients (mean age, 34.0 [SD, 4.5] years), 1470 (99.0%) completed the trial (n = 735 in both randomization groups) and were included in the analysis. Trial-of-labor rates did not differ significantly between intervention and control groups (43.3% vs 46.2%, respectively; adjusted absolute risk difference, -2.78% [95% CI, -7.80% to 2.25%]; adjusted relative risk, 0.94 [95% CI, 0.84-1.05]). There were no statistically significant differences in vaginal birth rates (31.8% in both groups; adjusted absolute risk difference, -0.04% [95% CI, -4.80% to 4.71%]; adjusted relative risk, 1.00 [95% CI, 0.86-1.16]) or in any of the other 6 clinical maternal and neonatal secondary outcomes. There also were no significant differences between the intervention and control groups in the 5 decision quality measures (eg, mean decisional conflict scores were 17.2 and 17.5, respectively; adjusted mean difference, -0.38 [95% CI, -1.81 to 1.05]; scores >25 are considered clinically important). Conclusions and Relevance: Among women with 1 previous cesarean delivery, use of a decision support tool compared with usual care did not significantly change the rate of trial of labor. Further research may be needed to assess the efficacy of this tool in other clinical settings or when implemented at other times in pregnancy.


Assuntos
Técnicas de Apoio para a Decisão , Participação do Paciente , Assistência Centrada no Paciente , Prova de Trabalho de Parto , Nascimento Vaginal Após Cesárea/estatística & dados numéricos , Adulto , Cesárea/tendências , Computadores , Tomada de Decisões , Feminino , Humanos , Gravidez , Inquéritos e Questionários
8.
J Vis Exp ; (159)2020 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-32510502

RESUMO

Drug resistant-tuberculosis (DR-TB) is a growing public health threat, and assessment of therapeutic drug levels may have important clinical benefits. Plasma drug levels are the current gold standard assessment, but require phlebotomy and a cold chain, and capture only very recent adherence. Our method uses hair, a matrix that is easily collected and reflective of long-term adherence, to test for 11 anti-TB medications. Previous work by our group shows that antiretroviral drug levels in hair are associated with HIV outcomes. Our method for DR-TB drugs uses 2 mg of hair (3 cm proximal to the root), which is pulverized and extracted in methanol. Samples are analyzed with a single LC-MS/MS method, quantifying 11 drugs in a 16 min run. Lower limits of quantification (LLOQs) for the 11 drugs range from 0.01 ng/mg to 1 ng/mg. Drug presence is confirmed by comparing ratios of two mass spectrometry transitions. Samples are quantified using the area ratio of the drug to the deuterated, 15N-, or 13C-labeled drug isotopologue. We used a calibration curve ranging from 0.001-100 ng/mg. Application of the method to a convenience sample of hair samples collected from DR-TB patients on directly observed therapy (DOT) indicated drug levels in hair within the linear dynamic range of nine of the eleven drugs (isoniazid, pyrazinamide, ethambutol, linezolid, levofloxacin, moxifloxacin, clofazimine, bedaquiline, pretomanid). No patient was on prothionamide, and the measured levels for ethionamide were close to its LLOQ (with further work instead examining the suitability of ethionamide's metabolite for monitoring exposure). In summary, we describe the development of a multi-analyte panel for DR-TB drugs in hair as a technique for therapeutic drug monitoring during drug-resistant TB treatment.


Assuntos
Antituberculosos/análise , Cromatografia Líquida/métodos , Cabelo/química , Espectrometria de Massas em Tandem/métodos , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Calibragem , Terapia Diretamente Observada , Humanos , Limite de Detecção , Padrões de Referência , Tuberculose Resistente a Múltiplos Medicamentos/sangue
9.
JMIR Res Protoc ; 9(4): e15029, 2020 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-32238341

RESUMO

BACKGROUND: The worldwide expansion of preexposure prophylaxis (PrEP) with oral tenofovir-disoproxil-fumarate/emtricitabine will be critical to ending the HIV epidemic. However, maintaining daily adherence to PrEP can be difficult, and the accuracy of self-reported adherence is often limited by social desirability bias. Pharmacologic adherence monitoring (measuring drug levels in a biomatrix) has been critical to interpreting PrEP trials, but testing usually requires expensive equipment and skilled personnel. We have recently developed a point-of-care (POC) immunoassay to measure tenofovir in urine, allowing real-time adherence monitoring for the first time. OBJECTIVE: The goal of this study is to examine a point-of-care adherence metric in PrEP to support and increase adherence via a randomized controlled trial. METHODS: The paper describes the protocol for a pilot randomized controlled trial to test the acceptability, feasibility, and impact on long-term adherence of implementing a POC urine test to provide real-time adherence feedback among women on PrEP. Eligible women (n=100) will be HIV-negative, ≥18 years old, and recruited from a clinic in Kenya that provides PrEP. Participants will be randomized 1:1 to the intervention of providing real-time feedback via the assay versus standard of care adherence counseling. Acceptability by participants will be assessed by a quantitative survey, as well as by qualitative data collected via in-depth interviews (n=20) and focus group discussions (n=4 groups, 5-10 women each). Feasibility will be assessed by the proportion of women retained in the study, the mean number of missed visits, the proportion of planned urine assessments completed, and messages delivered, while in-depth interviews with providers (n=8) will explore the ease of administering the urine test. Tenofovir levels in hair will serve as long-term adherence metrics. A linear mixed-effects model will estimate the effect of the intervention versus standard of care on logarithmically transformed levels of tenofovir in hair. RESULTS: This study has been funded by the National Institute of Health, approved by the Kenya Medical Research Institute Institutional Review Board, and will commence in June 2020. CONCLUSIONS: A novel urine assay to measure and deliver information on adherence to PrEP in real-time will be tested for the first time in this trial planned among women on PrEP in Kenya. Study findings will inform a larger-scale trial assessing the impact of real-time adherence monitoring/feedback on HIV prevention. Improving adherence to PrEP will have long-term implications for efforts to end the HIV epidemic worldwide. TRIAL REGISTRATION: ClinicalTrials.gov NCT03935464; https://clinicaltrials.gov/ct2/show/NCT03935464. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): PRR1-10.2196/15029.

10.
JCI Insight ; 5(4)2020 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-32045386

RESUMO

BACKGROUNDThe relative stabilities of the intact and defective HIV genomes over time during effective antiretroviral therapy (ART) have not been fully characterized.METHODSWe used the intact proviral DNA assay (IPDA) to estimate the rate of change of intact and defective proviruses in HIV-infected adults on ART. We used linear spline models with a knot at seven years and a random intercept and slope up to the knot. We estimated the influence of covariates on rates of change.RESULTSWe studied 81 individuals for a median of 7.3 (IQR 5.9-9.6) years. Intact genomes declined more rapidly from initial suppression through seven years (15.7% per year decline; 95% CI -22.8%, -8.0%) and more slowly after seven years (3.6% per year; 95% CI -8.1%, +1.1%). The estimated half-life of the reservoir was 4.0 years (95% CI 2.7-8.3) until year seven and 18.7 years (95% CI 8.2-infinite) thereafter. There was substantial variability between individuals in the rate of decline until year seven. Intact provirus declined more rapidly than defective provirus (P < 0.001) and showed a faster decline in individuals with higher CD4+ T cell nadirs.CONCLUSIONThe biology of the replication-competent (intact) reservoir differs from that of the replication-incompetent (non-intact) pool of proviruses. The IPDA will likely be informative when investigating the impact of interventions targeting the reservoir.FUNDINGDelaney AIDS Research Enterprise, UCSF/Gladstone Institute of Virology & Immunology CFAR, CFAR Network of Integrated Systems, amfAR Institute for HIV Cure Research, I4C and Beat-HIV Collaboratories, Howard Hughes Medical Institute, Gilead Sciences, Bill and Melinda Gates Foundation.


Assuntos
Fármacos Anti-HIV/uso terapêutico , DNA Viral/metabolismo , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Provírus/genética , Adulto , Contagem de Linfócito CD4 , Relação CD4-CD8 , Linfócitos T CD4-Positivos/virologia , Estudos de Coortes , Reservatórios de Doenças , Feminino , Infecções por HIV/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase/métodos , Latência Viral
12.
J Virol ; 94(3)2020 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-31723024

RESUMO

Understanding the impact of antiretroviral therapy (ART) duration on HIV-infected cells is critical for developing successful curative strategies. To address this issue, we conducted a cross-sectional/inter-participant genetic characterization of HIV-1 RNA from pre- and on-therapy plasmas and HIV-1 DNA from CD4+ T cell subsets derived from peripheral blood (PB), lymph node (LN), and gut tissues of 26 participants after 3 to 17.8 years of ART. Our studies revealed in four acute/early participants who had paired PB and LN samples a substantial reduction in the proportion of HIV-infected cells per year on therapy within the LN. Extrapolation to all 12 acute/early participants estimated a much smaller reduction in the proportion of HIV-1-infected cells within LNs per year on therapy that was similar to that in the participants treated during chronic infection. LN-derived effector memory T (TEM) cells contained HIV-1 DNA that was genetically identical to viral sequences derived from pre- and on-therapy plasma samples. The proportion of identical HIV-1 DNA sequences increased within PB-derived TEM cells. However, the infection frequency of TEM cells in PB was stable, indicating that cellular proliferation that compensates for T cell loss over time contributes to HIV-1 persistence. This study suggests that ART reduces HIV-infected T cells and that clonal expansion of HIV-infected cells maintains viral persistence. Importantly, LN-derived TEM cells are a probable source of HIV-1 genomes capable of producing infectious HIV-1 and should be targeted by future curative strategies.IMPORTANCE HIV-1 persists as an integrated genome in CD4+ memory T cells during effective therapy, and cessation of current treatments results in resumption of viral replication. To date, the impact of antiretroviral therapy duration on HIV-infected CD4+ T cells and the mechanisms of viral persistence in different anatomic sites is not clearly elucidated. In the current study, we found that treatment duration was associated with a reduction in HIV-infected T cells. Our genetic analyses revealed that CD4+ effector memory T (TEM) cells derived from the lymph node appeared to contain provirus that was genetically identical to plasma-derived virions. Moreover, we found that cellular proliferation counterbalanced the decay of HIV-infected cells throughout therapy. The contribution of cellular proliferation to viral persistence is particularly significant in TEM cells. Our study emphasizes the importance of HIV-1 intervention and provides new insights into the location of memory T cells infected with HIV-1 DNA, which is capable of contributing to viremia.


Assuntos
Antirretrovirais/uso terapêutico , Duração da Terapia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , HIV-1/imunologia , Adolescente , Linfócitos T CD4-Positivos/virologia , Criança , Pré-Escolar , Estudos Transversais , DNA Viral , HIV-1/genética , Humanos , Linfonodos , Provírus/genética , Subpopulações de Linfócitos T/virologia , Carga Viral , Viremia/virologia , Replicação Viral/efeitos dos fármacos
14.
AIDS ; 34(2): 255-260, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31634188

RESUMO

OBJECTIVE: HIV prevention and treatment studies demonstrate that pharmacologic adherence metrics are more accurate than self-report. Currently available metrics use liquid-chromatography/tandem-mass-spectrometry (LC-MS/MS), which is expensive and laboratory-based. We developed a specific and sensitive antibody against tenofovir, the backbone of treatment and prevention, but conversion to a lateral flow assay (LFA) - analogous to a urine pregnancy test - is required for point-of-care testing. We describe the development of the first LFA to measure antiretroviral adherence in real-time. METHODS: Previous work in a directly observed therapy study of providing tenofovir disoproxil fumarate (TDF) to HIV-noninfected volunteers at various simulated adherence patterns defined the appropriate cut-off for the LFA (1500 ng tenofovir/ml urine). We developed the LFA using a sample pad for urine; a conjugate pad coated with TFV-specific antibodies conjugated to colloidal gold nanoparticles; a nitrocellulose membrane striped with tenofovir-antigen (test line) and a control line; with an absorbent pad to draw urine across the reaction membrane. RESULTS: We tested 300 urine samples collected from the directly observed therapy study by this LFA and the gold-standard method of LC-MS/MS. The LFA demonstrated 97% specificity (95% CI 93-99%) and 99% sensitivity (94-100%) compared with LC-MS/MS. The LFA accurately classified 98% of patients who took a dose within 24 h as adherent. CONCLUSION: We describe the development and validation of the first point-of-care assay to measure short-term adherence to HIV prevention and treatment in routine settings. The assay is low-cost, easy-to-perform and measures the breakdown product (tenofovir) of both TDF and tenofovir alafenamide (TAF). This assay has the potential to improve HIV and PrEP outcomes worldwide by triggering differentiated service delivery with further study merited.


Assuntos
Fármacos Anti-HIV/urina , Adesão à Medicação/estatística & dados numéricos , Testes Imediatos , Profilaxia Pré-Exposição/métodos , Tenofovir/urina , Fármacos Anti-HIV/uso terapêutico , Cromatografia Líquida , Ouro/urina , Infecções por HIV/tratamento farmacológico , Infecções por HIV/prevenção & controle , Humanos , Nanopartículas Metálicas , Profilaxia Pré-Exposição/estatística & dados numéricos , Espectrometria de Massas em Tandem , Tenofovir/uso terapêutico
15.
Clin Infect Dis ; 71(6): 1517-1523, 2020 09 12.
Artigo em Inglês | MEDLINE | ID: mdl-31608363

RESUMO

BACKGROUND: Food insecurity is a well-established determinant of suboptimal, self-reported antiretroviral therapy (ART) adherence, but few studies have investigated this association using objective adherence measures. We examined the association of food insecurity with levels of ART concentrations in hair among women living with human immunodeficiency virus (WLHIV) in the United States. METHODS: We analyzed longitudinal data collected semiannually from 2013 through 2015 from the Women's Interagency HIV Study, a multisite, prospective, cohort study of WLHIV and controls not living with HIV. Our sample comprised 1944 person-visits from 677 WLHIV. Food insecurity was measured using the US Household Food Security Survey Module. ART concentrations in hair, an objective and validated measure of drug adherence and exposure, were measured using high-performance liquid chromatography with mass spectrometry detection for regimens that included darunavir, atazanavir, raltegravir, or dolutegravir. We conducted multiple 3-level linear regressions that accounted for repeated measures and the ART medication(s) taken at each visit, adjusting for sociodemographic and clinical characteristics. RESULTS: At baseline, 67% of participants were virally suppressed and 35% reported food insecurity. In the base multivariable model, each 3-point increase in food insecurity was associated with 0.94-fold lower ART concentration in hair (95% confidence interval, 0.89 to 0.99). This effect remained unchanged after adjusting for self-reported adherence. CONCLUSIONS: Food insecurity was associated with lower ART concentrations in hair, suggesting that food insecurity may be associated with suboptimal ART adherence and/or drug absorption. Interventions seeking to improve ART adherence among WLHIV should consider and address the role of food insecurity.


Assuntos
Fármacos Anti-HIV , Infecções por HIV , Preparações Farmacêuticas , Fármacos Anti-HIV/uso terapêutico , Estudos de Coortes , Feminino , Insegurança Alimentar , Abastecimento de Alimentos , HIV , Infecções por HIV/tratamento farmacológico , Infecções por HIV/epidemiologia , Humanos , Adesão à Medicação , Estudos Prospectivos , Estados Unidos/epidemiologia
16.
Clin Infect Dis ; 70(10): 2143-2151, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-31314073

RESUMO

BACKGROUND: Direct measurement of tenofovir (TFV) in urine could be an objective measure to monitor adherence to preexposure prophylaxis (PrEP) or TFV-based antiretroviral therapy (ART). METHODS: We conducted a 3-arm randomized, pharmacokinetic study of tenofovir disoproxil fumarate (TDF) 300 mg/emtricitabine (FTC) 200 mg among adults living with human immunodeficiency virus. Participants were randomized to receive controlled TDF/FTC dosing as (1) "perfect" adherence (daily); (2) "moderate" adherence (4 doses/week); or (3) "low" adherence (2 doses/week). We obtained trough spot urine and plasma samples during a 6-week directly observed therapy period and a 4-week washout period. TFV concentrations were compared between adherence arms using 1-way analysis of variance. RESULTS: Among 28 participants, the median age was 33 years and 16 (57%) were male. Correlation between TFV plasma and urine concentrations was strong (ρ = 0.78; P < .0001). Median (interquartile range) steady-state trough TFV concentrations (ng/mL) for perfect, moderate, and low TDF adherence were 41 (26-52), 16 (14-19), and 4 (3-5) in plasma; and 6480 (3940-14 300), 3405 (2210-5020), and 448 (228-675) in urine. Trough TFV concentrations at steady state were significantly different between the 3 adherence arms for plasma (P < .0001) and urine (P = .0002). Following drug cessation, TFV concentrations persisted longer in urine than plasma samples. Washout urine TFV concentrations and time to undetectable concentrations did not differ between the 3 randomized adherence groups. CONCLUSIONS: Urine TFV concentrations can inform interpretation of novel point-of-care urine-based TFV assays to assess recent TDF adherence. CLINICAL TRIALS REGISTRATION: NCT03012607


Assuntos
Fármacos Anti-HIV , Infecções por HIV , Preparações Farmacêuticas , Profilaxia Pré-Exposição , Tenofovir , Adulto , Fármacos Anti-HIV/uso terapêutico , Emtricitabina/uso terapêutico , Infecções por HIV/tratamento farmacológico , Humanos , Masculino , Plasma , Tenofovir/uso terapêutico
17.
J Infect Dis ; 221(5): 744-755, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-31796951

RESUMO

BACKGROUND: Identifying where human immunodeficiency virus (HIV) persists in people living with HIV and receiving antiretroviral therapy is critical to develop cure strategies. We assessed the relationship of HIV persistence to expression of chemokine receptors and their chemokines in blood (n = 48) and in rectal (n = 20) and lymph node (LN; n = 8) tissue collected from people living with HIV who were receiving suppressive antiretroviral therapy. METHODS: Cell-associated integrated HIV DNA, unspliced HIV RNA, and chemokine messenger RNA were quantified by quantitative polymerase chain reaction. Chemokine receptor expression on CD4+ T cells was determined using flow cytometry. RESULTS: Integrated HIV DNA levels in CD4+ T cells, CCR6+CXCR3+ memory CD4+ T-cell frequency, and CCL20 expression (ligand for CCR6) were highest in rectal tissue, where HIV-infected CCR6+ T cells accounted for nearly all infected cells (median, 89.7%). Conversely in LN tissue, CCR6+ T cells were infrequent, and there was a statistically significant association of cell-associated HIV DNA and RNA with CCL19, CCL21, and CXCL13 chemokines. CONCLUSIONS: HIV-infected CCR6+ CD4+ T cells accounted for the majority of infected cells in rectal tissue. The different relationships between HIV persistence and T-cell subsets and chemokines in rectal and LN tissue suggest that different tissue-specific strategies may be required to eliminate HIV persistence and that assessment of biomarkers for HIV persistence may not be generalizable between blood and other tissues.


Assuntos
Antirretrovirais/uso terapêutico , Linfócitos T CD4-Positivos/virologia , Infecções por HIV/tratamento farmacológico , HIV/genética , Receptores CCR6/metabolismo , Reto/imunologia , Quimiocinas/metabolismo , DNA Viral/sangue , DNA Viral/genética , Feminino , Infecções por HIV/sangue , Infecções por HIV/virologia , Humanos , Linfonodos/imunologia , Linfonodos/virologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , RNA Viral/sangue , RNA Viral/genética , Reto/virologia
18.
J Virus Erad ; 5(3): 167-173, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31700665

RESUMO

Observational and interventional studies for HIV cure research often use single-copy assays to quantify rare entities in blood or tissue samples. Statistical analysis of such measurements presents challenges due to tissue sampling variability and frequent findings of 0 copies in the sample analysed. We examined four approaches to analysing such studies, reflecting different ways of handling observations of 0 copies: (A) replace observations of 0 copies with 1 copy; (B) add 1 to all observed numbers of copies; (C) treat observations of 0 copies as left-censored at 1 copy; and (D) leave the data unaltered and apply a method for count data, negative binomial regression. Because research seeks to estimate general patterns rather than individuals' values, we argue that unaltered use of 0 copies is suitable for research purposes and that altering those observations can introduce bias. When applied to a simulated study comparing preintervention to postintervention measurements within 12 participants, methods A-C showed more attenuation than method D in the estimated intervention effect, less chance of finding P < 0.05 for the intervention effect and a lower chance of including the true intervention effect within the 95% confidence interval. Application of the methods to actual data from a study comparing multiply-spliced HIV RNA among men and women estimated smaller differences by methods A-C than by method D. We recommend that negative binomial regression, which is readily available in many statistical software packages, be considered for analysis of studies of rare entities that are measured by single-copy assays.

19.
J Perinatol ; 39(12): 1696, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31601948

RESUMO

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

20.
Front Microbiol ; 10: 2214, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31611857

RESUMO

To date, most assays for measuring the human immunodeficiency virus (HIV-1) reservoir do not include memory CD4+ T-cells expressing the activation marker, human leukocyte antigen-antigen D related (HLA-DR). However, little is known concerning the role these cells play in maintaining persistent HIV-1 during effective antiretroviral therapy (ART). To address this issue, we examined, cellular activation/exhaustion markers (Ki67, CCR5, PD-1, Lag-3 and Tim-3) and viral gag-pol DNA sequences within HLA-DR- and HLA-DR+ memory CD4+ T-cell subsets longitudinally from the peripheral blood of six participants over 3 to ≥15 years of effective therapy. HLA-DR expression was readily detected during the study period in all participants. The average expression levels of CCR5, PD-1 and Tim-3 were higher on the HLA-DR+ T-cell subset whereas the average of LAG-3 expression was higher on their HLA-DR- counterpart. The proportion of HIV-infected cells increased within the HLA-DR+ subset by an average of 18% per year of ART whereas the frequency of infected HLA-DR- T-cells slightly decreased over time (5% per year). We observed that 20-33% of HIV-DNA sequences from the early time points were genetically identical to viral sequences from the last time point within the same cell subset during ART. This indicates that a fraction of proviruses persists within HLA-DR+ and HLA-DR- T-cell subsets during prolonged ART. Our HIV-DNA sequence analyses also revealed that cells transitioned between the HLA-DR+ and HLA-DR- phenotypes. The Ki67 expression, a marker for cellular proliferation, and the combined markers of Ki67/PD-1 averaged 19-fold and 22-fold higher on the HLA-DR+ T-cell subset compared to their HLA-DR- counterpart. Moreover, cellular proliferation, as reflected by the proportion of genetically identical HIV-DNA sequences, increased within both T-cell subsets over the study period; however, this increase was greater within the HLA-DR+ T-cells. Our research revealed that cellular transition and proliferation contribute to the persistence of HIV in HLA-DR+ and HLA-DR- T-cell subsets during prolonged therapy. As such, the HIV reservoir expands during effective ART when both the HLA-DR+ and HLA-DR- cell subsets are included, and therapeutic interventions aimed at reducing the HIV-1 reservoir should target HLA-DR+ and HLA-DR- T-cells.

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