Background: Even during long-term combination antiretroviral therapy (cART), people living with HIV (PLHIV) have a dysregulated immune system, characterized by persistent immune activation, accelerated immune ageing and increased risk of non-AIDS comorbidities. A multi-omics approach is applied to a large cohort of PLHIV to understand pathways underlying these dysregulations in order to identify new biomarkers and novel genetically validated therapeutic drugs targets.Methods: The 2000HIV study is a prospective longitudinal cohort study of PLHIV on cART. In addition, untreated HIV spontaneous controllers were recruited. In-depth multi-omics characterization will be performed, including genomics, epigenomics, transcriptomics, proteomics, metabolomics and metagenomics, functional immunological assays and extensive immunophenotyping. Furthermore, the latent viral reservoir will be assessed through cell associated HIV-1 RNA and DNA, and full-length individual proviral sequencing on a subset. Clinical measurements include an ECG, carotid intima-media thickness and plaque measurement, hepatic steatosis and fibrosis measurement as well as psychological symptoms and recreational drug questionnaires. Additionally, considering the developing pandemic, COVID-19 history and vaccination was recorded. Participants return for a two-year follow-up visit. The 2000HIV study consists of a discovery and validation cohort collected at separate sites to immediately validate any finding in an independent cohort.Results: Overall, 1895 PLHIV from four sites were included for analysis, 1559 in the discovery and 336 in the validation cohort. The study population was representative of a Western European HIV population, including 288 (15.2%) cis-women, 463 (24.4%) non-whites, and 1360 (71.8%) MSM (Men who have Sex with Men). Extreme phenotypes included 114 spontaneous controllers, 81 rapid progressors and 162 immunological non-responders. According to the Framingham score 321 (16.9%) had a cardiovascular risk of >20% in the next 10 years. COVID-19 infection was documented in 234 (12.3%) participants and 474 (25.0%) individuals had received a COVID-19 vaccine.Conclusion: The 2000HIV study established a cohort of 1895 PLHIV that employs multi-omics to discover new biological pathways and biomarkers to unravel non-AIDS comorbidities, extreme phenotypes and the latent viral reservoir that impact the health of PLHIV. The ultimate goal is to contribute to a more personalized approach to the best standard of care and a potential cure for PLHIV.

People living with HIV (PLHIV) have an increased susceptibility to non-AIDS comorbidities. In this study, we systematically profiled 1,342 PLHIV across five omics layers and immune function. We found latent factors, resulting from integrating epigenomics, transcriptomics, proteomics, metabolomics and immune responses, linked to cardiovascular diseases, the presence of carotid plaque and chronic obstructive pulmonary disease in PLHIV. Mapping four omics layers to genetic variation identified 5,962 molecular quantitative trait loci, illustrating a common genetic regulation in PLHIV compared to healthy individuals. By performing Mendelian randomization, we uncovered host genetic-driven changes in baseline molecules causally related to immune responses upon stimulation with inactivated pathogens. Lastly, we uncovered that the inflammasome, genetically regulated by the NLRP12 locus, contributes to systemic inflammation across multiple molecular layers. This study offers a unique catalog of genetic and molecular determinants of immune function in PLHIV and elucidates molecular pathways driving inter-individual variation in immune response and comorbidities.

BACKGROUND: People living with HIV (PLHIV) receiving antiretroviral therapy (ART) exhibit persistent immune dysregulation and microbial dysbiosis, leading to development of cardiovascular diseases (CVDs). We initially compared plasma proteomic profiles between 205 PLHIV and 120 healthy control participants (HCs) and validated the results in an independent cohort of 639 PLHIV and 99 HCs. Differentially expressed proteins (DEPs) were then associated to microbiome data. Finally, we assessed which proteins were linked with CVD development in PLHIV. METHODS: Proximity extension assay technology was used to measure 1,472 plasma proteins. Markers of systemic inflammation (C-reactive protein, D-dimer, IL-6, soluble CD14, and soluble CD163) and microbial translocation (IFABP) were measured by ELISA, and gut bacterial species were identified using shotgun metagenomic sequencing. Baseline CVD data were available for all PLHIV, and 205 PLHIV were recorded for development of CVD during a 5-year follow-up. RESULTS: PLHIV receiving ART had systemic dysregulation of protein concentrations, compared with HCs. Most of the DEPs originated from the intestine and lymphoid tissues and were enriched in immune- and lipid metabolism–related pathways. DEPs originating from the intestine were associated with specific gut bacterial species. Finally, we identified upregulated proteins in PLHIV (GDF15, PLAUR, RELT, NEFL, COL6A3, and EDA2R), unlike most markers of systemic inflammation, associated with the presence and risk of developing CVD during 5-year follow-up. CONCLUSION: Our findings suggest a systemic dysregulation of protein concentrations in PLHIV; some proteins were associated with CVD development. Most DEPs originated from the gut and were related to specific gut bacterial species. TRIAL REGISTRATION: ClinicalTrials.gov NCT03994835. FUNDING: AIDS-fonds (P-29001), ViiV healthcare grant (A18-1052), Spinoza Prize (NWO SPI94-212), European Research Council (ERC) Advanced grant (grant 833247), and Indonesia Endowment Fund for Education.

Background: Cardiovascular disease is a major cause of morbidity and mortality in people living with HIV, who are at higher risk than the general population. We assessed, in a large cohort of people living with HIV, which cardiovascular, HIV‐specific, and lipoproteomic markers were associated with carotid intima‐media thickness (cIMT) and carotid plaque presence. We also studied guideline adherence on lipid‐lowering medication in individuals with high and very high risk for cardiovascular disease. Methods and Results: In 1814 individuals with a median (interquartile range) age of 53 (44–60) years, we found a carotid plaque in 909 (50.1%) and a median (interquartile range) intima‐media thickness of 0.66 (0.57–0.76) mm. Ultrasonography was used for the assessment of cIMT and plaque presence. Univariable and multivariable regression models were used for associations with cIMT and presence of plaques. Age, Black race, body mass index, type 2 diabetes, and smoking (pack years) were all positively associated with higher cIMT. Levels of high‐density lipoprotein cholesterol, specifically medium and large high‐density lipoprotein subclasses, were negatively associated with higher cIMT. Only age and prior myocardial infarction were positively related to the presence of a carotid plaque. Lipid‐lowering treatment was prescribed in one‐third of people living with HIV, who are at high and very high risk for cardiovascular disease. Conclusions: Traditional cardiovascular risk factors were significantly associated with higher cIMT but not with carotid plaques, except for age. HIV‐specific factors were not associated with both ultrasound measurements. Future studies are needed to elucidate which factors contribute to plaque formation. Improvement of guideline adherence on prescription of lipid‐lowering treatment in high‐ and very high‐risk patients for cardiovascular disease is recommended. Registration: URL: https://www.clinicaltrials.gov ; Unique identifier: NCT03994835.

In people living with HIV (PLHIV), integrase strand transfer inhibitors (INSTIs) are part of the first-line combination antiretroviral therapy (cART), while non-nucleoside reverse transcriptase inhibitor (NNRTI)-based regimens are alternatives. Distinct cART regimens may variably influence the risk for non-AIDS comorbidities. We aimed to compare the metabolome and lipidome of INSTI and NNRTI-based regimens. The 2000HIV study includes asymptomatic PLHIV (n = 1646) on long-term cART, separated into a discovery cohort with 730 INSTI and 617 NNRTI users, and a validation cohort encompassing 209 INSTI and 90 NNRTI users. Baseline plasma samples from INSTI and NNRTI users were compared using mass spectrometry-based untargeted metabolomic (n = 500) analysis. Perturbed metabolic pathways were identified using MetaboAnalyst software. Subsequently, nuclear magnetic resonance spectroscopy was used for targeted lipoprotein and lipid (n = 141) analysis. Metabolome homogeneity was observed between the different types of INSTI and NNRTI. In contrast, higher and lower levels of 59 and 45 metabolites, respectively, were found in the INSTI group compared to NNRTI users, of which 77.9% (81/104) had consistent directionality in the validation cohort. Annotated metabolites belonged mainly to ‘lipid and lipid-like molecules’, ‘organic acids and derivatives’ and ‘organoheterocyclic compounds’. In pathway analysis, perturbed ‘vitamin B1 (thiamin) metabolism’, ‘de novo fatty acid biosynthesis’, ‘bile acid biosynthesis’ and ‘pentose phosphate pathway’ were detected, among others. Lipoprotein and lipid levels in NNRTIs were heterogeneous and could not be compared as a group. INSTIs compared to individual NNRTI types showed that HDL cholesterol was lower in INSTIs compared to nevirapine but higher in INSTIs compared to doravirine. In addition, LDL size was lower in INSTIs and nevirapine compared to doravirine. NNRTIs show more heterogeneous cardiometabolic effects than INSTIs, which hampers the comparison between these two classes of drugs. Targeted lipoproteomic and lipid NMR spectroscopy showed that INSTI use was associated with a more unfavorable lipid profile compared to nevirapine, which was shifted to a more favorable profile for INSTI when substituting nevirapine for doravirine, with evidently higher fold changes. The cardiovascular disease risk profile seems more favorable in INSTIs compared to NNRTIs in untargeted metabolomic analysis using mass-spectrometry.

Background: We aimed to compare the non-AIDS event (nADE) risk between normal progressors using antiretroviral therapy (NP-ART) and people with human immunodeficiency virus (HIV, PWH) who naturally control HIV infection (HIV controllers), as well as the risk of nADE following ART in HIV controllers. Methods: The primary end point was the composite of cardiovascular disease, non-AIDS malignancy, or all-cause mortality, whichever came first. The role of ART in HIV controllers was assessed as a time-varying covariate. Results: We included 1007 ART-naive HIV controllers (60 of them were elite controllers), 1510 early-ART (<6 months after negative HIV test), and 15437 NP-ART (reference group), contributing 3813, 11 060, and 160 050 years of follow-up, respectively. HIV controllers had lower risk of the primary end point (hazard ratio [HR], 0.55; 95% confidence interval [CI]: .38–.81; P = .0023), all-cause mortality (adjusted HR [aHR], 0.45; 95% CI: .25–.79; P = .0054), and cardiovascular disease (aHR, 0.47; 95% CI: .22–.99; P = .046), but not non-AIDS malignancy (aHR, 0.74; 95% CI: .41–1.35; P = .33), compared with NP-ART. Among HIV controllers, each log10 lower baseline viral load further decreased the risk of a nADE (aHR, 0.54; 95% CI: .29–.99; P = .045). ART in HIV controllers did not reduce the risk of any nADE (aHR, 1.22; 95% CI: .66–2.29; P = .53). Conclusions: HIV controllers had a lower n ADE risk than NP-ART, especially in those with low plasma viral loads. ART did not alter the nADE risk in HIV controllers. Our findings help clinicians to decide on prescribing ART in HIV controllers.