Targeted sequencing of lung function loci in chronic obstructive pulmonary disease cases and controls

Chronic obstructive pulmonary disease (COPD) is the third leading cause of death worldwide; smoking is the main risk factor for COPD, but genetic factors are also relevant contributors. Genome-wide association studies (GWAS) of the lung function measures used in the diagnosis of COPD have identified a number of loci, however association signals are often broad and collectively these loci only explain a small proportion of the heritability. In order to examine the association with COPD risk of genetic variants down to low allele frequencies, to aid fine-mapping of association signals and to explain more of the missing heritability, we undertook a targeted sequencing study in 300 COPD cases and 300 smoking controls for 26 loci previously reported to be associated with lung function. We used a pooled sequencing approach, with 12 pools of 25 individuals each, enabling high depth (30x) coverage per sample to be achieved. This pooled design maximised sample size and therefore power, but led to challenges during variant-calling since sequencing error rates and minor allele frequencies for rare variants can be very similar. For this reason we employed a rigorous quality control pipeline for variant detection which included the use of 3 independent calling algorithms. In order to avoid false positive associations we also developed tests to detect variants with potential batch effects and removed them before undertaking association testing. We tested for the effects of single variants and the combined effect of rare variants within a locus. We followed up the top signals with data available (only 67% of collapsing methods signals) in 4,249 COPD cases and 11,916 smoking controls from UK Biobank. We provide suggestive evidence for the combined effect of rare variants on COPD risk in TNXB and in sliding windows within MECOM and upstream of HHIP. These findings can lead to an improved understanding of the molecular pathways involved in the development of COPD