Faculty Bibliography

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/CONCLUSIONS: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.

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.IMPORTANCECharacterizing 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.

Co-occurrence of injection drug use (IDU) and hepatitis C virus infection (HCV) is common in people living with HIV (PLWH) and leads to significantly increased mortality. Epigenetic clocks derived from DNA methylation (DNAm) are associated with disease progression and all-cause mortality. In this study, we hypothesized that epigenetic age mediates the relationships between the co-occurrence of IDU and HCV with mortality risk among PLWH. We tested this hypothesis in the Veterans Aging Cohort Study (n = 927) by using four established epigenetic clocks of DNAm age (i.e., Horvath, Hannum, Pheno, Grim). Compared to individuals without IDU and HCV (IDU-HCV-), participants with IDU and HCV (IDU+HCV+) showed a 2.23-fold greater risk of mortality estimated using a Cox proportional hazards model (hazard ratio: 2.23; 95% confidence interval: 1.62-3.09; p = 1.09E-06). IDU+HCV+ was associated with a significantly increased epigenetic age acceleration (EAA) measured by 3 out of 4 epigenetic clocks, adjusting for demographic and clinical variables (Hannum: p = 8.90E-04, Pheno: p = 2.34E-03, Grim: p = 3.33E-11). Furthermore, we found that epigenetic age partially mediated the relationship between IDU+HCV+ and all-cause mortality, up to a 13.67% mediation proportion. Our results suggest that comorbid IDU with HCV increases EAA in PLWH that partially mediates the increased mortality risk.

BACKGROUND:Cervical cancer oncogenesis starts with human papillomavirus (HPV) cell entry after binding to host cell surface receptors; however, the mechanism is not fully known. We examined polymorphisms in receptor genes hypothesized to be necessary for HPV cell entry and assessed their associations with clinical progression to precancer. METHODS:African American women (N = 1,728) from the MACS/WIHS Combined Cohort Study were included. Two case-control study designs were used-cases with histology-based precancer (CIN3+) and controls without; and cases with cytology-based precancer [high-grade squamous intraepithelial lesions (HSIL)] and controls without. SNPs in candidate genes (SDC1, SDC2, SDC3, SDC4, GPC1, GPC2, GPC3, GPC4, GPC5, GPC6, and ITGA6) were genotyped using an Illumina Omni2.5-quad beadchip. Logistic regression was used to assess the associations in all participants and by HPV genotypes, after adjusting for age, human immunodeficiency virus serostatus, CD4 T cells, and three principal components for ancestry. RESULTS:Minor alleles in SNPs rs77122854 (SDC3), rs73971695, rs79336862 (ITGA6), rs57528020, rs201337456, rs11987725 (SDC2), rs115880588, rs115738853, and rs9301825 (GPC5) were associated with increased odds of both CIN3+ and HSIL, whereas, rs35927186 (GPC5) was found to decrease the odds for both outcomes (P value ≤ 0.01). Among those infected with Alpha-9 HPV types, rs722377 (SDC3), rs16860468, rs2356798 (ITGA6), rs11987725 (SDC2), and rs3848051 (GPC5) were associated with increased odds of both precancer outcomes. CONCLUSIONS:Polymorphisms in genes that encode binding receptors for HPV cell entry may play a role in cervical precancer progression. IMPACT:Our findings are hypothesis generating and support further exploration of mechanisms of HPV entry genes that may help prevent progression to cervical precancer.

AIMS/HYPOTHESIS/OBJECTIVE:Our prior analysis of the Diabetes Prevention Program study identified a subset of five miRNAs that predict incident type 2 diabetes. The purpose of this study was to identify mRNAs and biological pathways targeted by these five miRNAs to elucidate potential mechanisms of risk and responses to the tested interventions. METHODS:Using experimentally validated data from miRTarBase version 8.0 and R (2021), we identified mRNAs with strong evidence to be regulated by individual or combinations of the five predictor miRNAs. Overrepresentation of the mRNA targets was assessed in pathways from the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation database. RESULTS:The five miRNAs targeted 167 pathways and 122 mRNAs. Nine of the pathways have known associations with type 2 diabetes: Insulin signaling, Insulin resistance, Diabetic cardiomyopathy, Type 2 diabetes, AGE-RAGE signaling in diabetic complications, HIF-1 signaling, TGF-beta signaling, PI3K/Akt signaling, and Adipocytokine signaling pathways. Vascular endothelial growth factor A (VEGFA) has prior genetic associations with risk for type 2 diabetes and was the most commonly targeted mRNA for this set of miRNAs. CONCLUSIONS/INTERPRETATION/CONCLUSIONS:These findings show that miRNA predictors of incident type 2 diabetes target mRNAs and pathways known to underlie risk for type 2 diabetes. Future studies should evaluate miRNAs as potential therapeutic targets for preventing and treating type 2 diabetes.

OBJECTIVE:MicroRNAs (miRs) are short (i.e., 18-26 nucleotide) regulatory elements of messenger RNA translation to amino acids. The purpose of this study was to assess whether miRs are predictive of incident T2D in the Diabetes Prevention Program (DPP) trial. RESEARCH DESIGN AND METHODS/METHODS:This was a secondary analysis (n = 1,000) of a subset of the DPP cohort that leveraged banked biospecimens to measure miRs. We used random survival forest and Lasso to identify the optimal miR predictors and cox proportional hazards to model time to T2D overall and within intervention arms. RESULTS:We identified five miRs (miR-144, miR-186, miR-203a, miR-205, miR-206) that constituted the optimal predictors of incident T2D after adjustment for covariates (hazards ratio 2.81 (95% confidence interval (CI) 2.05, 3.87); p < 0.001). Predictive risk scores following cross-validation showed the HR for the highest quartile risk group compared to the lowest quartile risk group was 5.91 (95% CI (2.02, 17.3); p < 0.001). There was significant interaction between the intensive lifestyle (HR 3.60, 95% CI (2.50, 5.18); p < 0.001) and the metformin (HR 2.72; 95% CI (1.47, 5.00); p = 0.001) groups compared to placebo. Of the five miRs identified, one targets a gene with prior known associations with risk for T2D. DISCUSSION/CONCLUSIONS:We identified five miRs that are optimal predictors of incident T2D in the DPP cohort. Future directions include validation of this finding in an independent sample in order to determine whether this risk score may have potential clinical utility for risk stratification of individuals with prediabetes, and functional analysis of the potential genes and pathways targeted by the miRs that were included in the risk score.

MicroRNAs (miRs) may contribute to disease etiology by influencing gene expression. Numerous databases are available for miR target prediction and validation, but their functionality is varied, and outputs are not standardized. The purpose of this review is to identify and describe databases for cataloging validated miR targets. Using Tools4miRs and PubMed, we identified databases with experimentally validated targets, human data, and a focus on miR-messenger RNA (mRNA) interactions. Data were extracted about the number of times each database was cited, the number of miRs, the target genes, the interactions per database, experimental methodology and key features of each database. The search yielded 10 databases, which in order of most cited to least were: miRTarBase, starBase/The Encyclopedia of RNA Interactomes, DIANA-TarBase, miRWalk, miRecords, miRGator, miRSystem, miRGate, miRSel and targetHub. Findings from this review suggest that the information presented within miR target validation databases can be enhanced by adding features such as flexibility in performing queries in multiple ways, downloadable data, ongoing updates and integrating tools for further miR-mRNA target interaction analysis. This review is designed to aid researchers, especially those new to miR bioinformatics tools, in database selection and to offer considerations for future development and upkeep of validation tools. Database URL http://mirtarbase.cuhk.edu.cn/.

PURPOSE/OBJECTIVE:To examine if gut microbial taxa abundances and predicted functional pathways correlate with Bristol Stool Form Scale (BSFS) classification at the end of neoadjuvant chemotherapy and radiation therapy (CRT) for rectal cancer. METHODS:tool samples for 16S rRNA gene sequencing. Stool consistency was evaluated using the BSFS. Gut microbiome data were analyzed using QIIME2. Correlation analysis were performed in R. RESULTS: CONCLUSION/CONCLUSIONS:abundance and to mycothiol biosynthesis and sucrose degradation pathways.