Telomere length (TL) is a widely used biomarker of biological aging, and is shortened in people living with HIV (PWH) due to chronic inflammation and immune-activation. The DNA methylation-based estimator of TL (DNAmTL) has emerged as a promising alternative to classical TL measurements, but their comparative performance in PWH remains poorly characterized. We conducted a cross-sectional study comparing DNAmTL and monochrome multiplex quantitative PCR-based TL (qPCR-TL) in two cohorts of PWH: 160 ART-naïve individuals and 143 long-term treated individuals with sustained viral suppression. DNAmTL was derived from blood DNA methylation profiles generated using the Infinium MethylationEPIC arrays, while qPCR-TL was measured using a standardized assay. We evaluated the agreement between methods using correlation, Bland-Altman analysis and Cohen’s Kappa coefficient, and their associations with demographic, clinical and immunological factors were assessed using multivariable linear regression. DNAmTL and qPCR-TL were moderately correlated in both ART-naïve (r = 0.54, p < 0.001) and ART-treated participants (r = 0.46, p < 0.001), showing reasonable to fair agreement across continuous and categorical assessments. Both measures were inversely correlated with chronological age, although associations were stronger for DNAmTL. Compared to qPCR-TL, DNAmTL demonstrated more robust and independent associations with female sex and ethnicity, immunovirological markers of HIV disease severity such as plasma HIV-RNA viral load and the CD4:CD8 ratio, as well as with comorbidities including type 2 diabetes and hepatitis C virus coinfection. DNAmTL provides a more biologically informative measure of aging and disease burden in PWH. Further research is warranted to validate these results and to determine its predictive value for clinical outcomes.

Background:Metformin is increasingly studied as a potential geroprotective agent in the general population. We aimed to test the efficacy and safety of metformin to improve epigenetic age in older, well-controlled, non-diabetic people living with HIV. Methods:METFORAGING was a single-centre, double-blind, randomised, parallel-group, placebo-controlled pilot trial. Non-diabetic participants living with HIV who were aged 50 years or older, virologically suppressed, on a stable antiretroviral regimen with undetectable viral load for at least 12 months, and had CD4+ T-cell counts >500 cells/μL were recruited from the HIV clinic at La Paz University Hospital (Madrid, Spain). Participants were randomly assigned (1:1) to receive 850 mg oral metformin or matching placebo twice a day for 96 weeks. Participants, investigators, and outcome assessors were masked to treatment allocation. Study visits were conducted at baseline and at weeks 4, 8, 24, 48, 72, and 96. Adherence was evaluated at each visit through pill count and self-report. At baseline and week 96, whole-blood samples were used to calculate biological age across 11 epigenetic biomarkers: first-generation epigenetic clocks (Horvath's clock and Hannum's clock), second-generation epigenetic clocks (PhenoAge and GrimAge V2), principal component-derived epigenetic clocks (PC-Horvath, PC-Hannum, PC-PhenoAge and PC-GrimAge), a third-generation clock (DunedinPACE), and the DNA methylation-based estimator of blood telomere length (DNAmTL). The primary outcome was the adjusted between-group difference in epigenetic age acceleration (EAA) change measured by the PhenoAge clock at week 96 in the per-protocol population. Analyses were stratified by age, sex, baseline CD4 count, smoking status, statin treatment, and route of HIV transmission. The trial was registered with EudraCT, 2021-003299-15. Findings:Between March 2, and Oct 2, 2022, 55 individuals were screened, and 40 were randomly assigned to metformin (n = 19) or placebo (n = 21). Enrolment was closed at 40 participants because of slow recruitment, below the pragmatic target of 60 outlined in the study protocol. Median age was 56.4 years (IQR 53.0-60.8), 12 (30%) were female, and 35 (87.5%) self-identified as White. Mean adherence by pill count was 97.5% in both groups. 35 (87.5%) of 40 participants continued treatment to 96 weeks (n = 17 in the metformin group and n = 18 in the placebo group; per-protocol population). At week 96, the adjusted between-group difference (metformin vs. placebo) for PhenoAge EAA was -1.02 years (95% confidence interval [CI] -5.30 to 3.26; p = 0.627). 48 adverse events occurred in 16 (84%) of 19 participants who received metformin and 48 adverse events occurred in 19 (90%) of 21 participants who received placebo. In the metformin group, no serious adverse events were attributed to the medication and no deaths or hospitalisation occurred. Interpretation:Although no significant difference was noted in the primary outcome between groups, these preliminary findings support the feasibility of geroscience-trials in this population and support further investigation of metformin in larger, adequately powered studies to determine whether metformin can modify biological ageing in people with HIV. Funding:Fondo de Investigaciones Sanitarias, Instituto de Salud Carlos III, and the European Union.

Background:Weight gain is a major complication of contemporary antiretroviral treatment (ART). We evaluated the effects of metformin on weight trajectories and metabolic outcomes in non-diabetic, non-obese, older persons with HIV-1 (PWH). Methods:In this double-blind, randomized, placebo-controlled pilot trial (METFORAGING), PWH 50 years or older, virologically suppressed, without diabetes or insulin resistance, were randomly assigned (1:1) to metformin 850 mg or placebo twice daily for 96 weeks at La Paz University Hospital (Madrid, Spain). This prespecified secondary analysis evaluated changes in body weight, body mass index (BMI), glycemic indices, lipid parameters, and serum growth differentiation factor-15 (GDF-15) in the per-protocol population. Results:Forty participants were randomly assigned (metformin n = 19; placebo n = 21); 35 completed treatment through week 96 (per-protocol population). At week 96, mean weight change was -1.5 kg (95% CI -2.9 to -0.03) with metformin and +1.3 kg (-0.7 to 3.3) with placebo (between-group difference -2.8 kg [95% CI -5.2 to -0.4]; P = .025). After discontinuation, weight in the metformin group returned toward baseline (between-group difference from week 96 to 144: 2.2 kg [95% CI 0.05 to 4.5]; P = .045). At week 96, changes in body weight correlated inversely with changes in serum GDF-15 in the metformin group (r = -0.67; P = .003). No serious adverse events were attributed to metformin. Conclusions:Metformin attenuated weight gain in nondiabetic, nonobese PWH receiving contemporary ART, but the effect was not sustained after discontinuation. These findings support larger trials to confirm efficacy and define optimal treatment duration. Trial Registration:EudraCT 2021-003299-15 (https://www.clinicaltrialsregister.eu).

BACKGROUND:Epigenetic age acceleration (EAA) predicts morbidity and mortality in the general population, but its prognostic utility in people with human immunodeficiency virus (HIV) (PWH) on suppressive antiretroviral therapy remains unclear. METHODS:Longitudinal observational cohort of 216 PWH with sustained virological suppression (median 6.5 years) recruited from the DUAL-GESIDA trial and La Paz Aging Cohort (Spain). Baseline blood DNA methylation was assessed to evaluate 8 epigenetic clocks, their principal-component (PC) derivatives, and the telomere length estimator. Over 10 years, we recorded AIDS, serious non-AIDS (SNAEs), and aging-related events. Associations between EAA and outcomes were evaluated by multivariate Cox models. RESULTS:One hundred five participants experienced ≥ 1 event, 162 events recorded in total (1 AIDS event, 65 SNAEs, and 96 aging-related events). A positive EAA by Horvath's clock was associated with a 50% higher risk of aging-related events. Participants with positive EAA according to PC-GrimAge, GrimAge V1-V2, as well as Hannum´s clock experienced more than 2-fold increased risk of SNAEs. Positive EAA by PC-GrimAge and PhenoAge was associated with an increased risk of non-AIDS cancer. Additionally, positive EAA according to GrimAge V1-V2 was linked to a 4-fold increase in all-cause mortality risk. All individuals who died (n = 17) had a baseline DunedinPACE value >1. No associations were found for the composite endpoint or cardiovascular events. CONCLUSIONS:In this preliminary study, GrimAge and PC-GrimAge were associated with increased risk of adverse outcomes in people with well-controlled HIV infection. These findings suggest their potential as long-term health-risk biomarkers, warranting validation in larger cohorts.

Background Epigenetic age acceleration (EAA) predicts morbidity and mortality in the general population, but its prognostic utility in people with human immunodeficiency virus (HIV) (PWH) on suppressive antiretroviral therapy remains unclear.Methods Longitudinal observational cohort of 216 PWH with sustained virological suppression (median 6.5 years) recruited from the DUAL-GESIDA trial and La Paz Aging Cohort (Spain). Baseline blood DNA methylation was assessed to evaluate 8 epigenetic clocks, their principal-component (PC) derivatives, and the telomere length estimator. Over 10 years, we recorded AIDS, serious non-AIDS (SNAEs), and aging-related events. Associations between EAA and outcomes were evaluated by multivariate Cox models.Results One hundred five participants experienced >= 1 event, 162 events recorded in total (1 AIDS event, 65 SNAEs, and 96 aging-related events). A positive EAA by Horvath's clock was associated with a 50% higher risk of aging-related events. Participants with positive EAA according to PC-GrimAge, GrimAge V1-V2, as well as Hannums clock experienced more than 2-fold increased risk of SNAEs. Positive EAA by PC-GrimAge and PhenoAge was associated with an increased risk of non-AIDS cancer. Additionally, positive EAA according to GrimAge V1-V2 was linked to a 4-fold increase in all-cause mortality risk. All individuals who died (n = 17) had a baseline DunedinPACE value >1. No associations were found for the composite endpoint or cardiovascular events.Conclusions In this preliminary study, GrimAge and PC-GrimAge were associated with increased risk of adverse outcomes in people with well-controlled HIV infection. These findings suggest their potential as long-term health-risk biomarkers, warranting validation in larger cohorts.

HIV reservoir latently persists in people with HIV (PWH) despite long-term antiretroviral therapy (ART). Total HIV DNA quantification is commonly employed as a surrogate marker to assess the size of the viral reservoir. We developed a duplex digital PCR assay to easily quantify the total HIV DNA in PWH on the microfluidic automated Absolute Q™ digital PCR platform. We assessed the linearity, specificity, sensitivity, and precision targeting the HIV LTR and the human RPP30 gene. We evaluated the assay in clinical samples from 50 PWH on ART and six ART-naïve PWH. The assay showed good linearity (R² = 0.977, p < 0.0001) and a 95% lower limit of detection of 79.7 HIV DNA copies/10⁶ cells. Repeatability and reproducibility were acceptable for 1,250 copies/10⁶ cells (CV = 8.7% and 10.9%) but higher variability for 150 copies/10⁶ cells (CV = 26.9% and 19.9%). Total HIV reservoir was detected in all PWH samples, with statistically significant differences between ART-treated and ART-naïve PWH (p < 0.0001). Positive strong correlation (rho = 0.868, p < 0.0001) was observed between the HIV reservoir in CD4 + T cells and PBMCs from 15 ART-treated PWH. Our assay expands the range of digital PCR platforms available for accurate HIV reservoir quantification.

ABCA1 methylation and N6-methyladenine DNA have been identified as epigenetic markers in coronary artery disease. We performed a cross-sectional exploratory study to analyze whether global DNA methylation and ABCA1 promoter methylation in monocytes are associated with coronary atherosclerosis in asymptomatic HIV individuals without cardiovascular disease. Coronary atherosclerosis was defined as a coronary plaque in at least one coronary segment. We included 27 individuals, 15 of whom had coronary atherosclerosis. ABCA1 DNA methylation was higher in participants with coronary atherosclerosis, with statistically significant differences at specific loci related to the activator protein-2 (AP-2) binding site. There were no differences in global levels of 5-methylcytosine and N6-methyladenine DNA between individuals with or without coronary atherosclerosis. ABCA1 methylation showed moderate correlations with both the Leaman score and coronary plaque volume, whereas global DNA N6-methyladenine had a negative moderate correlation with the segment severity score and LDL cholesterol. Among participants with coronary atherosclerosis, we found a strong negative correlation between ABCA1 methylation and total atheroma plaque volume, with certain loci (CpG 9 and 12) exhibiting very strong associations.

Background:People with HIV-1 (PWH) age differently than the general population. Blood telomere length (BTL) attrition is a surrogate biomarker of immunosenescence and aging in PWH. BTL is reduced immediately after HIV-1 infection and recovers in PWH with long-term virologic suppression, but the extent of this recovery is unknown. Methods:This prospective 6-year observational study assessed the evolution of BTL in PWH who were virologically suppressed. A cross-sectional analysis additionally compared BTL with age- and sex-matched blood donors and sex-matched persons older than 60 years from a general population cohort. DNA from whole blood was isolated, and relative BTL was determined by monochrome quantitative multiplex polymerase chain reaction assay and expressed as the ratio of telomere to single-copy gene (T/S). Results:A total of 128 PWH were included in the prospective 6-year observational study. These same 128 PWH (median age, 55 years; 27.3% women) were compared cross-sectionally at 6-year follow-up with 128 age- and gender-matched blood donors (median age, 55 years) and 128 gender-matched individuals older than 60 years from a general population cohort (median age, 70 years). An inverse correlation between age and BTL was observed. The median BTL of PWH was shorter than their matched blood donors (T/S, 1.07 [IQR, 0.95-1.17] vs 1.28 [IQR, 1.12-1.48]; P < .001) but longer than the elderly population (T/S, 0.89 [IQR, 0.77-0.98], P < .001). PWH experienced a BTL increase at 6 years of 2.9% (T/S, 1.04 vs 1.07; P = .002). In PWH, age was associated with a shorter BTL (coefficient, -0.007 45, SE = 0.002 04, P = .002) and baseline lower CD4 count with a gain in BTL (coefficient, -0.000 06, SE = 0.000 02, P = .004). Shorter baseline BTL (odds ratio, 0.91 [95% CI, .87-.94]; P < .001) and higher glucose levels (odds ratio, 1.04 [95% CI, 1.02-1.07]; P = .003) were associated with a greater similarity of BTL to the elderly population. Conclusions:PWH with long-term virologic suppression experience a trend toward an increased BTL after 6 years of follow-up. Middle-aged people with long-term controlled HIV-1 have a shorter BTL than expected for their chronologic age but longer than that of people 15 years older in the general population.

BACKGROUND:Previous epigenome-wide association studies have shown that HIV infection can disrupt the host DNA methylation landscape. However, it remains unclear how antiretroviral therapy (ART) affects the HIV-induced epigenetic modifications. METHODS:184 individuals with HIV from the NEAT001/ANRS143 clinical trial (with pre-ART and post-ART samples [96 weeks of follow-up]) and 44 age-and-sex matched individuals without HIV were included. We compared genome-wide DNA methylation profiles in whole blood between groups adjusting for age, sex, batch effects, and DNA methylation-based estimates of leucocyte composition. FINDINGS:We identified 430 differentially methylated positions (DMPs) between HIV+ pre-ART individuals and HIV-uninfected controls. In participants with HIV, ART initiation modified the DNA methylation levels at 845 CpG positions and restored 49.3% of the changes found between HIV+ pre-ART and HIV-uninfected individuals. We only found 15 DMPs when comparing DNA methylation profiles between HIV+ post-ART individuals and participants without HIV. The Gene Ontology enrichment analysis of DMPs associated with untreated HIV infection revealed an enrichment in biological processes regulating the immune system and antiviral responses. In participants with untreated HIV infection, DNA methylation levels at top HIV-related DMPs were associated with CD4/CD8 ratios and viral loads. Changes in DNA methylation levels after ART initiation were weakly correlated with changes in CD4+ cell counts and the CD4/CD8 ratio. INTERPRETATION:Control of HIV viraemia after 96 weeks of ART initiation partly restores the host DNA methylation changes that occurred before antiretroviral treatment of HIV infection. FUNDING:NEAT-ID Foundation and Instituto de Salud Carlos III, co-funded by European Union.

Background Accelerated epigenetic ageing can occur in untreated HIV infection and is partially reversible with effective antiretroviral therapy (ART). We aimed to make a long-term comparison of epigenetic ageing dynamics in people with HIV during untreated HIV infection and during suppressive ART. Methods In this longitudinal study, conducted over 17 years in HIV outpatient clinics in Switzerland, we applied 5 established epigenetic age estimators (epigenetic clocks) in peripheral blood mononuclear cells (PBMCs) in Swiss HIV Cohort Study participants before or during suppressive ART. All participants had a longitudinal set of PBMC samples available at four timepoints (T1-T4). T1 and T2 had to be 3 years or longer apart, as did T3 and T4. We assessed epigenetic age acceleration (EAA) and a novel rate of epigenetic ageing. Findings Between March 13, 1990, and Jan 18, 2018, we recruited 81 people with HIV from the Swiss HIV Cohort Study. We excluded one participant because a sample did not meet quality checks (transmission error). 52 (65%) of 80 patients were men, 76 (95%) were white, and the median patient age was 43 (IQR 37 center dot 5-47) years. Per year of untreated HIV infection (median observation 8 center dot 08 years, IQR 4 center dot 83-11 center dot 09), mean EAA was 0 center dot 47 years (95% CI 0 center dot 37 to 0 center dot 57) for Horvath's clock, 0 center dot 43 years (0 center dot 3 to 0 center dot 57) for Hannum's clock, 0 center dot 36 years (0 center dot 27 to 0 center dot 44) for SkinBlood clock, and 0 center dot 69 years (0 center dot 51 to 0 center dot 86) for PhenoAge. Per year of suppressive ART (median observation 9 center dot 8 years, IQR 7 center dot 2-11), mean EAA was -0 center dot 35 years (95% CI -0 center dot 44 to -0 center dot 27) for Horvath's clock, -0 center dot 39 years (-0 center dot 50 to -0 center dot 27) for Hannum's clock, -0 center dot 26 years (-0 center dot 33 to -0 center dot 18) for SkinBlood clock, and -0 center dot 49 years (-0 center dot 64 to -0 center dot 35) for PhenoAge. Our findings indicate that people with HIV epigenetically aged by a mean of 1 center dot 47 years for Horvath's clock, 1 center dot 43 years for Hannum's clock, 1 center dot 36 years for SkinBlood clock, and 1 center dot 69 years for PhenoAge per year of untreated HIV infection; and 0 center dot 65 years for Horvath's clock, 0 center dot 61 years for Hannum's clock, 0 center dot 74 years for SkinBlood clock, and 0 center dot 51 years for PhenoAge, per year of suppressive ART. GrimAge showed some change in the mean EAA during untreated HIV infection (0 center dot 10 years, 0 center dot 02 to 0 center dot 19) and suppressive ART (-0 center dot 05 years, -0 center dot 12 to 0 center dot 02). We obtained very similar results using the rate of epigenetic ageing. Contribution of multiple HIV-related, antiretroviral, and immunological variables, and of a DNA methylation-associated polygenic risk score to EAA was small. Interpretation In a longitudinal study over more than 17 years, epigenetic ageing accelerated during untreated HIV infection and decelerated during suppressive ART, highlighting the importance of limiting the duration of untreated HIV infection.

We investigated whether blood telomere length (TL), epigenetic age acceleration (EAA), and soluble inflammatory monocyte cytokines are associated with cardiovascular events or diabetes (DM) in people living with HIV (PLHIV). This was a case–control study nested in the Spanish HIV/AIDS Cohort (CoRIS). Cases with myocardial infarction, stroke, sudden death, or diabetes after starting antiretroviral therapy were included with the available samples and controls matched for sex, age, tobacco use, pre-ART CD4 cell count, viral load, and sample time-point. TL (T/S ratio) was analysed by quantitative PCR and EAA with DNA methylation changes by next-generation sequencing using the Weidner formula. Conditional logistic regression was used to explore the association with cardiometabolic events. In total, 180 participants (94 cases (22 myocardial infarction/sudden death, 12 strokes, and 60 DM) and 94 controls) were included. Of these, 84% were male, median (IQR) age 46 years (40–56), 53% were current smokers, and 22% had CD4 count ≤ 200 cells/mm3 and a median (IQR) log viral load of 4.52 (3.77–5.09). TL and EAA were similar in the cases and controls. There were no significant associations between TL, EAA, and monocyte cytokines with cardiometabolic events. TL and EAA were mildly negatively correlated with sCD14 (rho = −0.23; p = 0.01) and CCL2/MCP-1 (rho = −0.17; p = 0.02). We found no associations between TL, EAA, and monocyte cytokines with cardiovascular events or diabetes. Further studies are needed to elucidate the clinical value of epigenetic biomarkers and TL in PLHIV.

The antiretroviral treatment (ART) developed to control HIV infection led to a revolution in the prognosis of people living with HIV (PLWH). PLWH underwent from suffering severe disease and often fatal complications at young ages to having a chronic condition and a life expectancy close to the general population. Nevertheless, chronic age-related diseases increase as PLWH age. The harmful effect of HIV infection on the individual's immune system adds to its deterioration during ageing, exacerbating comorbidities. In addition, PLWH are more exposed to risk factors affecting ageing, such as coinfections or harmful lifestyles. The ART initiation reverses the biological ageing process but only partially, and additionally can have some toxicities that influence ageing. Observational studies suggest premature ageing in PLWH. Therefore, there is considerable interest in the early prediction of unhealthy ageing through validated biomarkers, easy to implement in HIV-clinical settings. The most promising biomarkers are second-generation epigenetic clocks and integrative algorithms.

OBJECTIVES:To evaluate whether the negative impact of tenofovir on telomere length (TL) is due to immune reconstitution interference or inhibition of telomerase.METHODS:One hundred and twenty-eight long-term aviraemic HIV adults treated with tenofovir-containing (n = 79) or tenofovir-sparing regimens (n = 49) were recruited to compare the following: TL in whole blood, PBMCs, CD4+ T cells and CD8+ T cells by quantitative PCR (qPCR); telomerase activity in PBMCs, CD4+ cells and CD8+ T cells using the TRAPeze RT Telomerase Detection Kit; and T cell maturational subset distribution by flow cytometry.RESULTS:In an adjusted analysis, participants treated with tenofovir for at least 4 years had shorter TL in CD8+ T cells (P = 0.04) and lower telomerase activity in CD4+ (P = 0.012) and CD8+ T cells (P = 0.023). Tenofovir treatment was also associated with lower proportions of recent thymic emigrant (RTE) CD4+ cells (P = 0.031) and PD1 marker expression (P = 0.013).CONCLUSIONS:In long-term aviraemic HIV adults, the inhibition of telomerase by tenofovir could explain telomere shortening in CD8+ T cells. There is no telomere shortening in the CD4+ compartment and the decrease in telomerase activity could be explained both by the inhibition by tenofovir and by the lower proportion of RTE CD4+cells.

BACKGROUND:Human immunodeficiency virus (HIV) infection induces epigenetic age acceleration (EAA), but it remains unclear whether epigenetic aging continues to accelerate during successful antiretroviral therapy (ART) and prolonged virological suppression.METHODS:We longitudinally analyzed 63 long-term aviremic HIV-infected adults. Using blood DNA methylation patterns, we calculated EAA measures based on 3 epigenetic clocks (Horvath's clock, PhenoAge, and GrimAge). We recorded the emergence of serious AIDS-related and non-AIDS-related events throughout the study to assess its association with EAA.RESULTS:All participants were on stable ART and were virologically suppressed. After 4 years of follow-up, PhenoAge-EAA and GrimAge-EAA showed no differences, whereas Horvath-EAA slightly decreased (median difference, -0.53 years; P = .015). Longitudinal changes in EAA measures were independent of changes in CD4 cell counts, the ART regimen, or other HIV-related factors. Nineteen percent of participants experienced a serious clinical event during the study. Horvath-EAA was significantly higher at baseline in participants with clinical events (P = .027). After adjusting for confounders, we found a trend toward an association of higher levels of all EAA measures at baseline with serious clinical events.CONCLUSIONS:Epigenetic aging did not accelerate in long-term aviremic HIV-infected adults after 4 years of successful ART. EAA measures deserve further study as potential tools for predicting clinical events.

Background DNA methylation-based estimators of biological age are reliable biomarkers of the ageing process. We aimed to investigate a range of epigenetic ageing biomarkers in a subshrdy of the NEAT001/ANRS143 clinical trial, which compared ritonavir-boosted darunavir with either raltegravir or tenofovir disoproxil fumarate and emtricitabine in antiretroviral therapy (ART)-naive adults. Methods We analysed frozen whole blood samples from 168 ART-naive participants with HIV from the NEAT001/ANRS143 trial, before ART initiation and after 2 years of ART (84 participants on ritonavir-boosted darunavir with raltegravir and 84 participants on ritonavir-boosted darunavir with tenofovir disoproxil fumarate and emtricitabine). We also included 44 participants without HIV with a similar age and sex distribution. We analysed DNA methylation. Epigenetic age estimators (Horvath's clock, Hannum's clock, GrimAge, and PhenoAge) and estimated leucocyte compositions were generated using Horvath's New Online Methylation Age Calculator and Houseman's method. We calculated epigenetic age acceleration measures for each estimator of epigenetic age. The NEAT001/ANRS143 trial is registered with ClinicalTrials.gov, NCT01066962. Findings Compared with the HIV-uninfected group, ART-naive participants with HIV showed higher epigenetic age acceleration (EAA) according to all EAA estimators (mean 2.5 years, 95% CI 1.89-3-22 for Horvath-EAA; 1.4 years, 0.74-1.99 for Hannum-EAA; 2.8 years, 1.97-3-68 for GrimAge-EAA; and 7.3 years, 6.40-8.13 for PhenoAge-EAA), with all differences being statistically significant except for Hannum-EAA (Horvath-EAA p=0.0008; Hannum-EAA p=0.059; GrimAge-EAA p=0.0021; and PhenoAge-EAA p<0.0001). Epigenetic ageing was more pronounced in participants who had C D4 counts less than 200 cells per pL (significant for PhenoAge and Hannum's clock, p=0.0015 and p=0.034, respectively) or viral loads over 100000 copies per mL at baseline (significant for PhenoAge, p=0.017). After 2 years of ART, epigenetic age acceleration was reduced, although PhenoAge and GrimAge remained significantly higher in participants with HIV compared with participants without HIV (mean difference 3.69 years, 95% CI 1.77-5-61; p=0.0002 and 2.2 years, 0.47-3.99; p=0.013, respectively). There were no significant differences in the ART effect on epigenetic ageing between treatment regimens. At baseline, participants with HIV showed dysregulation of DNA methylation-based estimated leucocyte subsets towards more differentiated T-cell phenotypes and proinflammatory leucocytes, which was also partly restored with ART. Interpretation ART initiation partly reversed epigenetic ageing associated with untreated HIV infection. Further studies are needed to understand the long-term dynamics and clinical relevance of epigenetic ageing biomarkers in people with HIV. Copyright (C) 2021 Elsevier Ltd. All rights reserved.

Background: Previously selected lamivudine resistance-associated mutations (RAMs) may remain archived within the proviral HIV-DNA. Objectives: To evaluate the ability of proviral DNA genotyping to detect lamivudine RAMs in HIV-1 virologically suppressed participants; the correlation between Sanger and next generation sequencing (NGS); and predictive factors for detection of lamivudine RAMs in proviral DNA. Methods: Cross-sectional study of participants on stable antiretroviral therapy and suppressed for >= 1 year. Analysis of proviral DNA was performed by Sanger sequencing in whole blood and by NGS in PBMCs. Results: We analysed samples from 102 subjects (52 with and 50 without lamivudine RAMs in historical plasma RNA-genotypes). Among participants with previous lamivudine resistance, Sanger sequencing detected RAMs in 26.9%. Detection rates significantly increased using NGS: 47.9%, 64.6%, 75% and 87.5% with the 20%, 10%, 5% and 1% thresholds, respectively. As for participants without historical lamivudine resistance, Sanger detected the RAMs in 1/49 (2%), and NGS (5% threshold) in 8/45 (17.8%). Multivariate models fitted to the whole population revealed that having a history of lamivudine resistance was a risk factor for detection of lamivudine RAMs by NGS. Among participants with historical lamivudine resistance, multivariate analysis showed that a longer time since HIV diagnosis was associated with persistence of archived mutations by NGS at thresholds of >10% [OR 1.10 (95% CI: 1.00-1.24)] and >5% [OR 1.16 (95% CI: 1.02-1.32)]. Conclusions: Proviral DNA Sanger sequencing does not detect the majority of historical lamivudine RAMs. NGS increases the sensitivity of detection at lower thresholds, although the relevance of these minority populations with lamivudine RAMs needs further evaluation.

OBJECTIVES:Our aim was to investigate factors associated with baseline blood telomere length in participants enrolled in NEAT 001/ANRS 143, a randomized, open-label trial comparing ritonavir-boosted darunavir (DRV/r) plus raltegravir (RAL) with DRV/r plus tenofovir disoproxil fumarate/emtricitabine (TDF/FTC) in antiretroviral therapy (ART)-naïve HIV-positive adults.METHODS:A cross-sectional study of 201 randomly selected participants who had stored samples available was carried out. We measured telomere length (i.e. the relative telomere length, calculated as the telomere to single copy gene ratio) at baseline with monochrome quantitative multiplex polymerase chain reaction (PCR). We used multivariable predictive linear regression to calculate mean differences and 95% confidence intervals (CIs) for the association between baseline telomere length and baseline characteristics.RESULTS:The baseline characteristics of the 201 participants did not differ from those of the 805 participants in the parent trial population: 89% were male, the mean age was 39 years, 83.6% were Caucasian, 93% acquired HIV infection via sexual transmission, the mean estimated time since HIV diagnosis was 2.1 years, the mean HIV-1 RNA load was 4.7 log10 HIV-1 RNA copies/mL, the mean nadir and baseline CD4 counts were 301 and 324 cells/μL, respectively, and the mean CD4:CD8 ratio was 0.4. In the univariate analysis, shorter telomere length was associated with older age (per 10 years) (P < 0.001), HIV-1 RNA ≥ 100 000 copies/mL (P = 0.001), CD4 count < 200 cells/μL (P = 0.037), lower CD4:CD8 ratio (P = 0.018), statin treatment (P = 0.004), and current alcohol consumption (P = 0.035). In the multivariable analysis, older age (P < 0.001) and HIV RNA ≥ 100 000 copies/mL (P = 0.054) were independently associated with shorter telomere length.CONCLUSIONS:Both age and HIV RNA viral load correlated with shorter blood telomere length in untreated persons living with HIV. These results suggest that HIV infection and age have synergistic and independent impacts upon immunosenescence.

AIM:To evaluate the association between DNA methylation and frailty in the HIV-infected population and to investigate the usefulness of assessing frailty as a clinical marker to identify age acceleration.METHODS:Frailty was assessed according to Fried's frailty phenotype. DNA methylation was analyzed in 10 frail patients, and compared with 10 robust control patients, all with HIV. Predicted age was inferred using the Weidner's formula. Age acceleration was assessed using the difference between predicted and chronological age.RESULTS:HIV-infected frail patients had significantly higher biological predicted ages than chronological ages (mean acceleration: 10.3 years; p = 0.012).CONCLUSIONS:We link age acceleration and frailty in an older HIV population. Frailty could be used in this population for implementing specific clinical approaches.

BackgroundWe aimed to describe the most frequently prescribed initial antiretroviral therapy (ART) regimens in recent years in HIV-positive persons in the Cohort of the Spanish HIV/AIDS Research Network (CoRIS) and to investigate factors associated with the choice of each regimen.MethodsWe analyzed initial ART regimens prescribed in adults participating in CoRIS from 2014 to 2017. Only regimens prescribed in >5% of patients were considered. We used multivariable multinomial regression to estimate Relative Risk Ratios (RRRs) for the association between sociodemographic and clinical characteristics and the choice of the initial regimen.ResultsAmong 2874 participants, abacavir(ABC)/lamivudine(3TC)/dolutegavir(DTG) was the most frequently prescribed regimen (32.1%), followed by tenofovir disoproxil fumarate (TDF)/emtricitabine (FTC)/elvitegravir(EVG)/cobicistat(COBI) (14.9%), TDF/FTC/rilpivirine (RPV) (14.0%), tenofovir alafenamide (TAF)/FTC/EVG/COBI (13.7%), TDF/FTC+DTG (10.0%), TDF/FTC+darunavir/ritonavir or darunavir/cobicistat (bDRV) (9.8%) and TDF/FTC+raltegravir (RAL) (5.6%). Compared with ABC/3TC/DTG, starting TDF/FTC/RPV was less likely in patients with CD4<200 cells/μL and HIV-RNA>100.000 copies/mL. TDF/FTC+DTG was more frequent in those with CD4<200 cells/μL and HIV-RNA>100.000 copies/mL. TDF/FTC+RAL and TDF/FTC+bDRV were also more frequent among patients with CD4<200 cells//μL and with transmission categories other than men who have sex with men. Compared with ABC/3TC/DTG, the prescription of other initial ART regimens decreased from 2014-2015 to 2016-2017 with the exception of TDF/FTC+DTG. Differences in the choice of the initial ART regimen were observed by hospitals' location.ConclusionsThe choice of initial ART regimens is consistent with Spanish guidelines' recommendations, but is also clearly influenced by physician's perception based on patient's clinical and sociodemographic variables and by the prescribing hospital location.