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Transcriptomic profiling facilitates classification of response to influenza challenge

ABSTRACT: Despite increases in vaccination coverage, reductions in influenza-related mortality have not been observed. Better vaccines are therefore required and influenza challenge studies can be used to test the efficacy of new vaccines. However, this requires the accurate post-challenge classification of subjects by outcome, which is limited in current methods that use artificial thresholds to assign “symptomatic” and “asymptomatic” phenotypes. We present data from an influenza challenge study in which 22 healthy adults (11 vaccinated) were inoculated with H3N2 influenza (A/Wisconsin/67/2005). We generated genome-wide gene expression data from peripheral blood taken immediately before the challenge and at 12, 24, and 48 hours post-challenge. Variation in symptomatic scoring was found among those with laboratory confirmed influenza. By combining the dynamic transcriptomic data with the clinical parameters this variability can be reduced. We identified four subjects with severe laboratory confirmed flu that show differential gene expression in 1,103 probes 48 hours post-challenge compared to the remaining subjects. We have further reduced this profile to 6 genes that can be used to define these subjects. We have used this gene set to predict symptomatic infection from an independent study. This analysis gives further insight into host-pathogen interactions during influenza infection. However, the major potential value is in the clinical trial setting by providing a more quantitative method to better classify symptomatic individuals post influenza challenge.

ORGANISM(S): Homo sapiens

PROVIDER: GSE61754 | GEO | 2014/11/06

SECONDARY ACCESSION(S): PRJNA262041

REPOSITORIES: GEO

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Project description:The transcriptional responses of human hosts towards influenza viral pathogens are important for understanding virus-mediated immunopathology. Despite great advances gained through studies using model organisms, the complete temporal host transcriptional responses in a natural human system are poorly understood. In a human challenge study using live influenza (H3N2/Wisconsin) viruses, we conducted a clinically uninformed (unsupervised) factor analysis on gene expression profiles and established an ab initio molecular signature that strongly correlates to symptomatic clinical disease. This is followed by the identification of 42 biomarkers whose expression patterns best differentiate early from late phases of infection. In parallel, a clinically informed (supervised) analysis revealed over-stimulation of multiple viral sensing pathways in symptomatic hosts and linked their temporal trajectory with development of diverse clinical signs and symptoms. The resultant inflammatory cytokine profiles were shown to contribute to the pathogenesis because their significant increase preceded disease manifestation by 36 hours. In subclinical asymptomatic hosts, we discovered strong transcriptional regulation of genes involved in inflammasome activation, genes encoding virus interacting proteins, and evidence of active anti-oxidant and cell-mediated innate immune response. Taken together, our findings offer insights into influenza virus-induced pathogenesis and provide a valuable tool for disease monitoring and management in natural environments. During the challenge study, subjects had peripheral blood taken 24 hours prior to inoculation with virus (baseline), immediately prior to inoculation (pre-challenge) and at set intervals (8 hour intervals) following challenge. RNA was extracted at Expression Analysis (Durham, NC) from whole blood using the PAXgene™ 96 Blood RNA Kit (PreAnalytiX, Valencia, CA) employing the manufacturer's recommended protocol. While whole blood RNA is initially extracted, a secondary procedure (B-globin reduction) was then employed to remove the contribution of red blood cell (RBC) RNA to the total RNA. A set of four peptide nucleic acid (PNA) oligomers whose sequences are complementary to the 3’ portions of the alpha and beta hemoglobin RNA transcripts were added to reduce globin RNA transcription due to RBC. The inhibition of globin cDNA synthesis dramatically reduces the relative amount of anti-sense, biotin-labeled cRNA corresponding to the hemoglobin transcripts. Hybridization and microarray data collection was performed using the Human Genome U133A 2.0 Array (Affymetrix, Santa Clara, CA) and expression profiles were pre-processed using robust multi-array (RMA) method

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Transcriptomic profiling facilitates classification of response to influenza challenge

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