The Peripheral Blood Transcriptome Identifies the Presence and Extent of Disease in Idiopathic Pulmonary Fibrosis

<div><h3>Rationale</h3><p>Peripheral blood biomarkers are needed to identify and determine the extent of idiopathic pulmonary fibrosis (IPF). Current physiologic and radiographic prognostic indicators diagnose IPF too late in the course of disease. We hypothesize that peripheral blood biomarkers will identify disease in its early stages, and facilitate monitoring for disease progression.</p> <h3>Methods</h3><p>Gene expression profiles of peripheral blood RNA from 130 IPF patients were collected on Agilent microarrays. Significance analysis of microarrays (SAM) with a false discovery rate (FDR) of 1% was utilized to identify genes that were differentially-expressed in samples categorized based on percent predicted D_LCO and FVC.</p> <h3>Main Measurements and Results</h3><p>At 1% FDR, 1428 genes were differentially-expressed in mild IPF (D_LCO >65%) compared to controls and 2790 transcripts were differentially- expressed in severe IPF (D_LCO >35%) compared to controls. When categorized by percent predicted D_LCO, SAM demonstrated 13 differentially-expressed transcripts between mild and severe IPF (< 5% FDR). These include CAMP, CEACAM6, CTSG, DEFA3 and A4, OLFM4, HLTF, PACSIN1, GABBR1, IGHM, and 3 unknown genes. Principal component analysis (PCA) was performed to determine outliers based on severity of disease, and demonstrated 1 mild case to be clinically misclassified as a severe case of IPF. No differentially-expressed transcripts were identified between mild and severe IPF when categorized by percent predicted FVC.</p> <h3>Conclusions</h3><p>These results demonstrate that the peripheral blood transcriptome has the potential to distinguish normal individuals from patients with IPF, as well as extent of disease when samples were classified by percent predicted D_LCO, but not FVC.</p> </div

Genome-wide imputation study identifies novel HLA locus for pulmonary fibrosis and potential role for auto-immunity in fibrotic idiopathic interstitial pneumonia.

To access publisher's full text version of this article, please click on the hyperlink in Additional Links field or click on the hyperlink at the top of the page marked Files. This article is open access.Fibrotic idiopathic interstitial pneumonias (fIIP) are a group of fatal lung diseases with largely unknown etiology and without definitive treatment other than lung transplant to prolong life. There is strong evidence for the importance of both rare and common genetic risk alleles in familial and sporadic disease. We have previously used genome-wide single nucleotide polymorphism data to identify 10 risk loci for fIIP. Here we extend that work to imputed genome-wide genotypes and conduct new RNA sequencing studies of lung tissue to identify and characterize new fIIP risk loci.We performed genome-wide genotype imputation association analyses in 1616 non-Hispanic white (NHW) cases and 4683 NHW controls followed by validation and replication (878 cases, 2017 controls) genotyping and targeted gene expression in lung tissue. Following meta-analysis of the discovery and replication populations, we identified a novel fIIP locus in the HLA region of chromosome 6 (rs7887 P meta  = 3.7 × 10(-09)). Imputation of classic HLA alleles identified two in high linkage disequilibrium that are associated with fIIP (DRB1*15:01 P = 1.3 × 10(-7) and DQB1*06:02 P = 6.1 × 10(-8)). Targeted RNA-sequencing of the HLA locus identified 21 genes differentially expressed between fibrotic and control lung tissue (Q < 0.001), many of which are involved in immune and inflammatory response regulation. In addition, the putative risk alleles, DRB1*15:01 and DQB1*06:02, are associated with expression of the DQB1 gene among fIIP cases (Q < 1 × 10(-16)).We have identified a genome-wide significant association between the HLA region and fIIP. Two HLA alleles are associated with fIIP and affect expression of HLA genes in lung tissue, indicating that the potential genetic risk due to HLA alleles may involve gene regulation in addition to altered protein structure. These studies reveal the importance of the HLA region for risk of fIIP and a basis for the potential etiologic role of auto-immunity in fIIP.National Heart, Lung and Blood Institute R01-HL095393 R01-HL097163 P01-HL092870 RC2-HL101715 U01-HL089897 U01-HL089856 U01-HL108642 P50-HL089493

Overlaid networks and associated pathway analysis

<p>. Solid lines (direct relationship); Dashed lines (indirect relationship); Red filled (up-regulation); and Green filled (down-regulation).</p

Principal component analysis of severe IPF (D_LCO ≤35% predicted).

<p>Red spheres: definite IPF; Orange spheres: probable IPF; Yellow spheres: unclassifiable fibrosis; Green spheres: healthy controls. Axis labels: white-first principal component; blue-second principal component; lavender- third principal component. The majority of severe cases, both probable and definite, cluster along the first principal component. Three cases of unclassifiable fibrosis distribute with IPF cases.</p

Differentially-expressed genes that distinguish mild IPF from control.

‡<p>Significance analysis of microarrays (SAM) of IPF samples when categorized by percent predicted D_LCO ≥65% [N = 16]. Differentially- expressed transcripts with <1% false discovery rate and > 2-fold change in expression are represented. Fold changes are expressed as log₂ ratio. See supplementary tables for a complete list of differentially-expressed genes and corresponding accession numbers.</p

Clinical and demographic IPF variables categorized by D_LCO.

<p>Abbreviations: Idiopathic Pulmonary Fibrosis (IPF); Not Reported (NR); Diffusing Capacity for Carbon Monoxide (D_LCO); Forced Vital Capacity (FVC).</p

Clinical and demographic IPF variables categorized by FVC.

<p>Abbreviations: Idiopathic Pulmonary Fibrosis (IPF); Not Reported (NR); Diffusing Capacity for Carbon Monoxide (D_LCO); Forced Vital Capacity (FVC).</p