Genetic and Epigenetic Determinants of Pediatric Obesity-Associated Asthma

Obesity-associated asthma is a distinct entity characterized by worse airflow obstruction and suboptimal response to conventional asthma management that disproportionately affects urban children. However, its pathogenesis is poorly understood. In contrast to atopic T helper (Th) type 2 inflammation found in ?classic? childhood asthma, studies from our lab indicate that obesity-associated asthma is associated with non-atopic Th1 inflammation that correlates with lower airway obstruction. Additionally, we found decreased promoter methylation of genes associated with T cell signaling suggesting that epigenetic mechanisms may mediate the distinct asthma phenotype in obese children. However, molecular mechanisms that underlie non-atopic inflammation in obese asthmatics are not known. Asthma and obesity are multifactorial diseases, determined by genetic susceptibility, including that related to ancestry, epigenetic modification by environmental exposures, and non-genetic environmental factors that influence gene expression and thus clinical phenotype. Hence, investigation of the association between gene expression, genetic polymorphisms, and epigenetic modulation may identify a unifying molecular mechanism to explain the obese asthma phenotype observed in urban children. We have collected 200 samples from children in the Bronx during the past 5 years of this award to test the hypothesis that Th-cell gene expression in obese asthmatics differs from that in normal-weight asthmatics, is determined by genetic polymorphisms (eQTLs and ancestry-specific haplotypes), and is influenced by DNA methylation. We proposed the following specific aims to address these hypotheses: (1) To quantify Th-cell gene expression differences between obese asthmatic and normal-weight asthmatic children. (2) To determine if epigenetic mechanisms mediate the effect of obesity on asthma. (3) To investigate if genetic polymorphisms, including ancestry-specific haplotypes, determine susceptibility to obesity-associated asthma. Th cells from 100 obese asthmatic and 100 normal-weight asthmatic children matched for age and gender have been isolated. We have simultaneously conducted genome-wide assays of a) gene expression by Directional Transcriptome Sequencing (RNA-seq) b) DNA methylation by HELP-tagging and c) genetic polymorphisms and single nucleotide polymorphisms (SNPs) informative of ancestry by array-based genotyping on these cells. Association of gene expression with genetic polymorphisms has identified variants predicting genetic susceptibility, which may be used to screen at-risk individuals. Verification studies of gene expression, DNA methylation, and genetic polymorphisms will now allow us to confirm the mechanistic molecular pathways specific to the obese asthma phenotype and improve our understanding of the disease pathogenesis. We will identify key molecules associated with non-atopic inflammation that could be novel targets for therapeutic intervention. Together, our study will provide fundamental insights into the pathogenesis of obesity-associated asthma among high-risk ethnicities. As a pediatric pulmonologist, my goal is to become an independent translational investigator in the field of pediatric asthma with a focus on the pulmonary effects of obesity. This K23 Mentored Patient Oriented- Research Career Development Award has allowed an in-depth study of genetic and epigenetic factors influencing Th cell mediated inflammation in obese asthmatic minority children. Findings from this proposal have provided the foundation for my R01 funding, which is focused on further investigation of the CDC42 pathway, identified as the top differentially expressed pathway in this K23 award. As our future goals, we plan to a) validate the key differentially expressed and/or methylated molecules in a separate cohort of children with poorly controlled disease b) confirm the role of key molecules in in-vitro cell culture systems with molecule- specific inhibitors and/or methylation modulators c) conduct a prospective investigation into links between obesity onset, rapidity of weight gain, development of asthma and Th cell differentiation, given the young age at which obesity-associated asthma afflicts urban children. The career development plan proposed in this application incorporated a) mentorship from established researchers in the field of epigenomics, obesity-related diseases, T cell biology, asthma, and mucosal and systemic immunity, b) coursework in genetics and epigenomics, and hand on training in novel methods to investigate gene expression, genotyping and DNA methylation, and c) participation and presentation at local, regional and national meetings. Together, the research proposed in this K23 award laid the foundation for my development into an independent investigator in the field of pediatric obesity-associated asthma. I am now focused on building on this foundation by verifying our findings to subsequently use these results as preliminary data for further investigation of the contribution of genetic susceptibility to childhood obesity- associated asthma.