Bradley E. Aouizerat

Abstract

Oral squamous cell carcinoma (OSCC) biomarker studies rarely employ multi-omic biomarker strategies and pertinent clinicopathologic characteristics to predict mortality. In this study we determine for the first time a combined epigenetic, gene expression, and histology signature that differentiates between patients with different tobacco use history (heavy tobacco use with ≥10 pack years vs. no tobacco use). Using The Cancer Genome Atlas (TCGA) cohort (n = 257) and an internal cohort (n = 40), we identify 3 epigenetic markers (GPR15, GNG12, GDNF) and 13 expression markers (IGHA2, SCG5, RPL3L, NTRK1, CD96, BMP6, TFPI2, EFEMP2, RYR3, DMTN, GPD2, BAALC, and FMO3), which are dysregulated in OSCC patients who were never smokers vs. those who have a ≥ 10 pack year history. While mortality risk prediction based on smoking status and clinicopathologic covariates alone is inaccurate (c-statistic = 0.57), the combined epigenetic/expression and histologic signature has a c-statistic = 0.9409 in predicting 5-year mortality in OSCC patients.

Abstract

Substance use disparities among sexual and gender minority (SGM) people are attributed to minority stress, but few studies have examined minority stress and cannabis use over time or investigated differences in cannabis use trajectories by less-studied gender subgroups. We examined if longitudinal cannabis use trajectories are related to baseline minority stressors and if gender differences persisted after accounting for minority stress. Cannabis use risk was measured annually over four years (2017–2021) within a longitudinal cohort study of SGM adults in the United States (N = 11,813). Discrimination and victimization, internalized stigma, disclosure and concealment, and safety and acceptance comprised minority stress (n = 5,673). Latent class growth curve mixture models identified five cannabis use trajectories: ‘low or no risk’, ‘low moderate risk’, ‘high moderate risk’, ‘steep risk increase’, and ‘highest risk’. Participants who reported past-year discrimination and/or victimization at baseline had greater odds of membership in any cannabis risk category compared to the ‘low risk’ category (odds ratios [OR] 1.17–1.33). Internalized stigma was related to ‘high moderate’ and ‘highest risk’ cannabis use (ORs 1.27–1.38). After accounting for minority stress, compared to cisgender men, gender expansive people and transgender men had higher odds of ‘low moderate risk’ (ORs 1.61, 1.67) or ‘high moderate risk’ (ORs 2.09, 1.99), and transgender men had higher odds of ‘highest risk’ (OR 2.36) cannabis use. This study indicates minority stress is related to prospective cannabis use risk trajectories among SGM people, and transgender men and gender expansive people have greater odds of trajectories reflecting cannabis use risk.

Abstract

Background Epigenome-wide association studies (EWAS) have identified CpG sites associated with HIV infection in blood cells in bulk, which offer limited knowledge of cell-type specific methylation patterns associated with HIV infection. In this study, we aim to identify differentially methylated CpG sites for HIV infection in immune cell types: CD4+ T-cells, CD8+ T-cells, B cells, Natural Killer (NK) cells, and monocytes. Methods Applying a computational deconvolution method, we performed a cell-type based EWAS for HIV infection in three independent cohorts (Ntotal = 1,382). DNA methylation in blood or in peripheral blood mononuclear cells (PBMCs) was profiled by an array-based method and then deconvoluted by Tensor Composition Analysis (TCA). The TCA-computed CpG methylation in each cell type was first benchmarked by bisulfite DNA methylation capture sequencing in a subset of the samples. Cell-type EWAS of HIV infection was performed in each cohort separately and a meta-EWAS was conducted followed by gene set enrichment analysis. Results The meta-analysis unveiled a total of 2,021 cell-type unique significant CpG sites for five inferred cell types. Among these inferred cell-type unique CpG sites, the concordance rate in the three cohorts ranged from 96% to 100% in each cell type. Cell-type level meta-EWAS unveiled distinct patterns of HIV-associated differential CpG methylation, where 74% of CpG sites were unique to individual cell types (false discovery rate, FDR <0.05). CD4+ T-cells had the largest number of unique HIV-associated CpG sites (N = 1,624) compared to any other cell type. Genes harboring significant CpG sites are involved in immunity and HIV pathogenesis (e.g. CD4+ T-cells: NLRC5, CX3CR1, B cells: IFI44L, NK cells: IL12R, monocytes: IRF7), and in oncogenesis (e.g. CD4+ T-cells: BCL family, PRDM16, monocytes: PRDM16, PDCD1LG2). HIV-associated CpG sites were enriched among genes involved in HIV pathogenesis and oncogenesis that were enriched among interferon-α and -γ, TNF-α, inflammatory response, and apoptotic pathways. Conclusion Our findings uncovered computationally inferred cell-type specific modifications in the host epigenome for people with HIV that contribute to the growing body of evidence regarding HIV pathogenesis.

Abstract

Telomere length (TL) is an important indicator of cellular aging. Shorter TL is associated with several age-related diseases including coronary heart disease, heart failure, diabetes, osteoporosis, and cancer. Recently, a DNA methylation-based TL (DNAmTL) estimator has been developed as an alternative method for directly measuring TL. In this study, we examined the association of DNAmTL with cancer prevalence and mortality risk among people with and without HIV in the Veterans Aging Cohort Study Biomarker Cohort (VACS, N = 1917) and Women's Interagency HIV Study Cohort (WIHS, N = 481). We profiled DNAm in whole blood (VACS) or in peripheral blood mononuclear cells (WIHS) using an array-based method. Cancer prevalence was estimated from electronic medical records and cancer registry data. The VACS Index was used as a measure of physiologic frailty. Models were adjusted for self-reported race and ethnicity, batch, smoking status, alcohol consumption, and five cell types (CD4, CD8, NK, B cell, and monocyte). We found that people with HIV had shorter average DNAmTL than those without HIV infection [beta = −0.25, 95% confidence interval (−0.32, −0.18), p = 1.48E-12]. Greater value of VACS Index [beta = −0.002 (−0.003, −0.001), p = 2.82E-05] and higher cancer prevalence [beta = −0.07 (−0.10, −0.03), p = 1.37E-04 without adjusting age] were associated with shortened DNAmTL. In addition, one kilobase decrease in DNAmTL was associated with a 40% increase in mortality risk [hazard ratio: 0.60 (0.44, 0.82), p = 1.42E-03]. In summary, HIV infection, physiologic frailty, and cancer are associated with shortening DNAmTL, contributing to an increased risk of all-cause mortality.

Abstract

Aims: The purpose of this study was to assess the levels of cardiovascular health (CVH) of Black and Latino adults with type 2 diabetes (T2D) and examine the association of individual and microsystem level factors with their CVH score. Methods: This was a cross-sectional design in 60 Black and Latino Adults aged 18–40 with T2D. Data were collected on sociodemographic, individual (sociodemographic, diabetes self-management, sleep disturbance, depressive symptoms, quality of life, and the inflammatory biomarkers IL-6 and hs-CRP) and microsystem factors (family functioning), and American Heart Association’s Life’s Simple 7 metrics of CVH. Factors significantly associated with the CVH score in the bivariate analyses were entered into a linear regression model. Results: The sample had a mean age 34 ± 5 years and was primarily female (75%) with a mean CVH score was 8.6 ± 2.2 (possible range of 0–14). The sample achieved these CVH factors at ideal levels: body mass index <25 kg/m2 (8%); blood pressure <120/80 (42%); hemoglobin A1c < 7% (57%); total cholesterol <200 mg/dL (83%); healthy diet (18%); never or former smoker > one year (95%); and physical activity (150 moderate-to-vigorous minutes/week; 45%). In the multivariable model, two factors were significantly associated with cardiovascular health: hs-CRP (B = −0.11621, p <.0001) and the general health scale (B = 0.45127, p =.0013). Conclusions: This sample had an intermediate level of CVH, with inflammation and general health associated with overall CVH score.

Abstract

Problem: Bacterial vaginosis (BV) disproportionally impacts Black and Hispanic women, placing them at risk for HIV, sexually transmitted infections and preterm birth. It is unknown whether there are differences by genetic ancestry in BV risk or whether polymorphisms associated with BV risk differ by ancestry. Methods: Women's Interagency HIV Study (WIHS) participants with longitudinal Nugent scores were dichotomized as having (n = 319, Nugent 7–10) or not having BV (n = 367, Nugent 0–3). Genetic ancestry was defined by clustering of principal components from ancestry informative markers and further stratified by BV status. 627 single nucleotide polymorphisms (SNPs) across 41 genes important in mucosal defense were identified in the WIHS GWAS. A logistic regression analysis was adjusted for nongenetic predictors of BV and self-reported race/ethnicity to assess associations between genetic ancestry and genotype. Results: Self-reported race and genetic ancestry were associated with BV risk after adjustment for behavioral factors. Polymorphisms in mucosal defense genes including syndecans, cytokines and toll-like receptors (TLRs) were associated with BV in all ancestral groups. Conclusions: The common association of syndecan, cytokine and TLR genes and the importance of immune function and inflammatory pathways in BV, suggests these should be targeted for further research on BV pathogenesis and therapeutics.

Abstract

The Diabetes Prevention Program (DPP) randomized controlled trial demonstrated that metformin treatment reduced progression to type 2 diabetes (T2D) by 31% compared to placebo in adults with prediabetes. Circulating micro-ribonucleic acids (miRs) are promising biomarkers of T2D risk, but little is known about their associations with metformin regimens for T2D risk reduction. We compared the change in 24 circulating miRs from baseline to 2 years in a subset from DPP metformin intervention (n = 50) and placebo (n = 50) groups using Wilcoxon signed rank tests. Spearman correlations were used to evaluate associations between miR change and baseline clinical characteristics. Multiple linear regression was used to adjust for covariates. The sample was 73% female, 17% Black, 13% Hispanic, and 50 ± 11 years. Participants were obese, normotensive, prediabetic, and dyslipidemic. Change in 12 miR levels from baseline to 2 years was significantly different in the metformin group compared with placebo after adjusting for multiple comparisons: six (let-7c-5p, miR-151a-3p, miR-17-5p, miR-20b-5p, miR-29b-3p, and miR-93-5p) were significantly upregulated and six (miR-130b-3p, miR-22-3p, miR-222-3p, miR-320a-3p, miR-320c, miR-92a-3p) were significantly downregulated in the metformin group. These miRs help to explain how metformin is linked to T2D risk reduction, which may lead to novel biomarkers, therapeutics, and precision health strategies.

Abstract

Objective: This epigenomics sub-study embedded within a randomized controlled trial examined whether an evidenced-based behavioral intervention model that decreased stimulant use altered leukocyte DNA methylation (DNAm). Methods: Sexual minority men with HIV who use methamphetamine were randomized to a five-session positive affect intervention (n = 32) or an attention-control condition (n = 21), both delivered during three months of contingency management for stimulant abstinence. All participants exhibited sustained HIV virologic control – an HIV viral load less than 40 copies/mL at baseline and six months post-randomization. The Illumina EPIC BeadChip measured leukocyte methylation of cytosine-phosphate-guanosine (CpG) sites mapping onto five a priori candidate genes of interest (i.e., ADRB2, BDNF, FKBP5, NR3C1, OXTR). Functional DNAm pathways and soluble markers of immune dysfunction were secondary outcomes. Results: Compared to the attention-control condition, the positive affect intervention significantly decreased methylation of CpG sites on genes that regulate β2 adrenergic and oxytocin receptors. There was an inconsistent pattern for the direction of the intervention effects on methylation of CpG sites on genes for glucocorticoid receptors and brain-derived neurotrophic factor. Pathway analyses adjusting for the false discovery rate (padj < 0.05) revealed significant intervention-related alterations in DNAm of Reactome pathways corresponding to neural function as well as dopamine, glutamate, and serotonin release. Positive affect intervention effects on DNAm were accompanied by significant reductions in the self-reported frequency of stimulant use. Conclusions: There is an epigenetic signature of an evidence-based behavioral intervention model that reduced stimulant use, which will guide the identification of biomarkers for treatment responses.

Abstract

Within-host HIV populations continually diversify during untreated infection, and this diversity persists within infected cell reservoirs during antiretroviral therapy (ART). Achieving a better understanding of on-ART proviral evolutionary dynamics, and a better appreciation of how the overall persisting pool of (largely genetically defective) proviruses differs from the much smaller replication-competent HIV reservoir, is critical to HIV cure efforts. We reconstructed within-host HIV evolutionary histories in blood from seven participants of the Women’s Interagency HIV Study who experienced HIV seroconversion, and used these data to characterize the diversity, lineage origins, and ages of proviral env-gp120 sequences sampled longitudinally up to 12 years on ART. We also studied HIV sequences emerging from the reservoir in two participants. We observed that proviral clonality generally increased over time on ART, with clones frequently persisting long term. While on-ART proviral integration dates generally spanned the duration of untreated infection, HIV emerging in plasma was exclusively younger (i.e., dated to the years immediately pre-ART). The genetic and age distributions of distinct proviral sequences remained stable during ART in all but one participant, in whom there was evidence that younger proviruses had been preferentially eliminated after 12 years on ART. Analysis of the gag region in three participants corroborated our env-gp120-based observations, indicating that our observations are not influenced by the HIV region studied. Our results underscore the remarkable genetic stability of the distinct proviral sequences that persist in blood during ART. Our results also suggest that the replication-competent HIV reservoir is a genetically restricted, younger subset of this overall proviral pool. IMPORTANCE Characterizing the genetically diverse HIV sequences that persist in the reservoir despite antiretroviral therapy (ART) is critical to cure efforts. Our observations confirm that proviruses persisting in blood on ART, which are largely genetically defective, broadly reflect the extent of within-host HIV evolution pre-ART. Moreover, on-ART clonal expansion is not appreciably accompanied by the loss of distinct proviral lineages. In fact, on-ART proviral genetic composition remained stable in all but one participant, in whom, after 12 years on ART, proviruses dating to around near ART initiation had been preferentially eliminated. We also identified recombinant proviruses between parental sequence fragments of different ages. Though rare, such sequences suggest that reservoir cells can be superinfected with HIV from another infection era. Overall, our finding that the replication-competent reservoir in blood is a genetically restricted, younger subset of all persisting proviruses suggests that HIV cure strategies will need to eliminate a reservoir that differs in key respects from the overall proviral pool.

Abstract

Objectives: To determine the dysregulated signaling pathways of head and neck squamous cell carcinoma associated with circulating tumor cells (CTCs) via single-cell molecular characterization. Introduction: Head and neck squamous cell carcinoma (HNSCC) has a significant global burden and is a disease with poor survival. Despite trials exploring new treatment modalities to improve disease control rates, the 5 year survival rate remains low at only 60%. Most cancer malignancies are reported to progress to a fatal phase due to the metastatic activity derived from treatment-resistant cancer cells, regarded as one of the most significant obstacles to develope effective cancer treatment options. However, the molecular profiles of cancer cells have not been thoroughly studied. Methods: Here, we examined in-situ HNSCC tumors and pairwisely followed up with the downstream circulating tumor cells (CTCs)-based on the surrogate biomarkers to detect metastasis that is established in other cancers – not yet being fully adopted in HNSCC treatment algorithms. Results: Specifically, we revealed metastatic HNSCC patients have complex CTCs that could be defined through gene expression and mutational gene profiling derived from completed single-cell RNASeq (scRNASeq) that served to confirm molecular pathways inherent in these CTCs. To enhance the reliability of our findings, we cross-validated those molecular profiles with results from previously published studies. Conclusion: Thus, we identified 5 dysregulated signaling pathways in CTCs to derive HNSCC biomarker panels for screening HNSCC in situ tumors.