The Altered DNA Methylome as a Determinant of Variable Disease Progression in IPF

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DESCRIPTION (provided by applicant): Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease with no known effective therapy. Despite the dismal median survival, the prognosis of individual patients is variable and unpredictable, with some patients remaining stable and others deteriorating rapidly. The mechanisms that account for the variability in patients' clinical disease course are unknown. Fibroblasts are the main effector cell in fibrosis, and their relentless and exuberant activity is felt to contribute to the progressive nature of IPF. IPF fibroblasts exhibit gene expression changes that persist in culture even after multiple cell passages, suggesting that epigenetic modifications may be responsible for some of these gene expression differences. Alterations in DNA methylation of select genes have been identified in IPF fibroblasts, but a global assessment of the DNA methylomic abnormalities in IPF and their contribution to disease progression has never been assessed. The objectives of this project are to explore how genome-wide DNA methylomic and transcriptomic changes in IPF cells contribute to the clinical variability in IPF and determine how specific anti- and profibrotic mediators, prostaglandin E2 (PGE2) and transforming growth factor (TGF)-�1 respectively, contribute to altered DNA methylation patterns in IPF. The central hypothesis is that fibroblasts of IPF patients acquire distinct alterations in the DNA methylome in response to multiple extracellular signals, and that these distinct patterns of methylation contribute to both the heterogeneous gene expression profiles of fibroblasts and variable rates of clinical progression in IPF patients. This project has two specific aims: 1) examine how DNA methylomic changes in IPF fibroblasts contribute to both variable fibroblast gene expression patterns and clinical disease progression, and 2) understand the mechanisms by which PGE2 and TGF-�1 regulate DNA methyltransferases (DNMTs) and DNA methylation patterns and how they may contribute to differences in DNA methylation and gene expression of IPF fibroblasts. Fibroblasts from IPF and nonfibrotic lungs will be examined for methylation differences using the Illumina HumanMethylation450 Bead-Chip Array. Differentially methylated genes will be correlated with differential gene expression and longitudinal clinical disease progression. In Aim 2, the effects o PGE2 and TGF-�1 on DNA methylation and DNA methylation machinery will be examined in normal and IPF fibroblasts. The approach is innovative because it will utilize nonbiased, "next-generation" technologies to correlate whole genomic DNA methylation patterns with longitudinal clinical data from IPF patients and delineate mechanisms that regulate DNA methylation machinery and specific DNA methylation patterns. This project is significant because it will establish DNA methylation as a mechanism that contributes to differential gene expression, altered fibroblast function, and ultimately clinical disease progression, offer insight into how DN methylation patterns may be regulated, and identify novel pathways that may be candidates for future therapeutic targeting.