Inherited causes of clonal haematopoiesis in 97,691 whole genomes.

Age is the dominant risk factor for most chronic human diseases, but the mechanisms through which ageing confers this risk are largely unknown1. The age-related acquisition of somatic mutations that lead to clonal expansion in regenerating haematopoietic stem cell populations has recently been associated with both haematological cancer2-4 and coronary heart disease5-this phenomenon is termed clonal haematopoiesis of indeterminate potential (CHIP)6. Simultaneous analyses of germline and somatic whole-genome sequences provide the opportunity to identify root causes of CHIP. Here we analyse high-coverage whole-genome sequences from 97,691 participants of diverse ancestries in the National Heart, Lung, and Blood Institute Trans-omics for Precision Medicine (TOPMed) programme, and identify 4,229 individuals with CHIP. We identify associations with blood cell, lipid and inflammatory traits that are specific to different CHIP driver genes. Association of a genome-wide set of germline genetic variants enabled the identification of three genetic loci associated with CHIP status, including one locus at TET2 that was specific to individuals of African ancestry. In silico-informed in vitro evaluation of the TET2 germline locus enabled the identification of a causal variant that disrupts a TET2 distal enhancer, resulting in increased self-renewal of haematopoietic stem cells. Overall, we observe that germline genetic variation shapes haematopoietic stem cell function, leading to CHIP through mechanisms that are specific to clonal haematopoiesis as well as shared mechanisms that lead to somatic mutations across tissues.

Polygenic transcriptome risk scores for COPD and lung function improve cross-ethnic portability of prediction in the NHLBI TOPMed program.

While polygenic risk scores (PRSs) enable early identification of genetic risk for chronic obstructive pulmonary disease (COPD), predictive performance is limited when the discovery and target populations are not well matched. Hypothesizing that the biological mechanisms of disease are shared across ancestry groups, we introduce a PrediXcan-derived polygenic transcriptome risk score (PTRS) to improve cross-ethnic portability of risk prediction. We constructed the PTRS using summary statistics from application of PrediXcan on large-scale GWASs of lung function (forced expiratory volume in 1 s [FEV1] and its ratio to forced vital capacity [FEV1/FVC]) in the UK Biobank. We examined prediction performance and cross-ethnic portability of PTRS through smoking-stratified analyses both on 29,381 multi-ethnic participants from TOPMed population/family-based cohorts and on 11,771 multi-ethnic participants from TOPMed COPD-enriched studies. Analyses were carried out for two dichotomous COPD traits (moderate-to-severe and severe COPD) and two quantitative lung function traits (FEV1 and FEV1/FVC). While the proposed PTRS showed weaker associations with disease than PRS for European ancestry, the PTRS showed stronger association with COPD than PRS for African Americans (e.g., odds ratio [OR] = 1.24 [95% confidence interval [CI]: 1.08-1.43] for PTRS versus 1.10 [0.96-1.26] for PRS among heavy smokers with ≥ 40 pack-years of smoking) for moderate-to-severe COPD. Cross-ethnic portability of the PTRS was significantly higher than the PRS (paired t test p < 2.2 × 10-16 with portability gains ranging from 5% to 28%) for both dichotomous COPD traits and across all smoking strata. Our study demonstrates the value of PTRS for improved cross-ethnic portability compared to PRS in predicting COPD risk.

Dysanapsis Genetic Risk Predicts Lung Function Across the Lifespan.

Rationale Dysanapsis refers to a mismatch between airway tree caliber and lung size arising early in life. Dysanapsis assessed by computed tomography (CT) is evident by early adulthood and associated with chronic obstructive pulmonary disease (COPD) risk later in life. Objective By examining the genetic factors associated with CT-assessed dysanapsis, we aimed to elucidate its molecular underpinnings and physiological significance across the lifespan. Methods We performed a genome-wide association study (GWAS) of CT-assessed dysanapsis in 11,951 adults, including individuals from two population-based and two COPD-enriched studies. We applied colocalization analysis to integrate GWAS and gene expression data from whole blood and lung. Genetic variants associated with dysanapsis were combined into a genetic risk score that was applied to examine association with lung function in children from a population-based birth cohort (n=1,278) and adults from the UK Biobank (n=369,157). Measurements and Main Results CT-assessed dysanapsis was associated with genetic variants from 21 independent signals in 19 gene regions, implicating HHIP, DSP, and NPNT as potential molecular targets based on colocalization of their expression. Higher dysanapsis genetic risk score was associated with obstructive spirometry among 5 year old children and among adults in the 5th, 6th and 7th decades of life. Conclusions CT-assessed dysanapsis is associated with variation in genes previously implicated in lung development and dysanapsis genetic risk is associated with obstructive lung function from early life through older adulthood. Dysanapsis may represent an endo-phenotype link between the genetic variations associated with lung function and COPD.

Mitochondrial DNA copy number variation in asthma risk, severity, and exacerbations.

BACKGROUND: Asthma pathophysiology is associated with mitochondrial dysfunction. Mitochondrial DNA copy number (mtDNA-CN) has been used as a proxy of mitochondrial function, with lower levels indicating mitochondrial dysfunction in population studies of cardiovascular diseases and cancers. OBJECTIVES: We investigated whether lower levels of mtDNA-CN are associated with asthma diagnosis, severity, and exacerbations. METHODS: mtDNA-CN is evaluated in blood from 2 cohorts: UK Biobank (UKB) (asthma, n = 39,147; no asthma, n = 302,302) and Severe Asthma Research Program (SARP) (asthma, n = 1283; nonsevere asthma, n = 703). RESULTS: Individuals with asthma have lower mtDNA-CN compared to individuals without asthma in UKB (beta, -0.006 [95% confidence interval, -0.008 to -0.003], P = 6.23 × 10-6). Lower mtDNA-CN is associated with asthma prevalence, but not severity in UKB or SARP. mtDNA-CN declines with age but is lower in individuals with asthma than in individuals without asthma at all ages. In a 1-year longitudinal study in SARP, mtDNA-CN was associated with risk of exacerbation; those with highest mtDNA-CN had the lowest risk of exacerbation (odds ratio 0.333 [95% confidence interval, 0.173 to 0.542], P = .001). Biomarkers of inflammation and oxidative stress are higher in individuals with asthma than without asthma, but the lower mtDNA-CN in asthma is independent of general inflammation or oxidative stress. Mendelian randomization studies suggest a potential causal relationship between asthma-associated genetic variants and mtDNA-CN. CONCLUSION: mtDNA-CN is lower in asthma than in no asthma and is associated with exacerbations. Low mtDNA-CN in asthma is not mediated through inflammation but is associated with a genetic predisposition to asthma.

Genetic and T2 biomarkers linked to the efficacy of HDM sublingual immunotherapy in asthma.

BACKGROUND: Hypersensitivity to house dust mite (HDM) allergens is a common cause of allergic asthma symptoms and can be effectively treated with allergy immunotherapy (AIT). OBJECTIVE: To investigate whether genetic and type 2 (T2) inflammatory biomarkers correlate with disease severity in subjects with allergic asthma, and whether this can be modified by AIT. METHODS: MITRA (NCT01433523) was a phase III, randomised, double-blind, placebo-controlled trial of HDM sublingual immunotherapy (SLIT)-tablets in adults with HDM allergic asthma. Post hoc analyses of the study population (N=742) evaluated associations between T2 inflammatory (blood eosinophils, eosinophil cationic protein (ECP), total IgE and tryptase) and genetic (single-nucleotide polymorphisms, SNP) biomarkers (n=582) for the primary study endpoint (time to first moderate/severe asthma exacerbation). SNP associations were verified in HDM-positive subgroup from an independent 3-year Severe Asthma Research Programme (SARP3) subject cohort. RESULTS: An increased asthma exacerbation risk in subjects homozygous for SNP rs7216389 (chromosomal locus 17q12-21) was reduced (p=0.037) by treatment with HDM SLIT (HR=0.37 (95% CI 0.22 to 0.64), p<0.001). The associations between exacerbation risk and 17q12-21 SNPs were replicated in the SARP3 HDM-positive subgroup. High levels of T2 biomarkers were associated with increased risk of asthma exacerbations in the placebo group. HDM SLIT-tablet treatment reduced this risk (blood eosinophils: HR=0.50 (95% CI 0.30 to 0.85); ECP: HR=0.45 (95% CI 0.29 to 0.87); tryptase: HR=0.45 (95% CI 0.25 to 0.80)). The treatment effect was higher (p=0.006) for subjects with a higher number of elevated T2 biomarkers. CONCLUSIONS: HDM SLIT-tablet AIT is efficacious in HDM-sensitised asthma subjects with a genetic asthma predisposition and/or an underlying T2 endotype. TRIAL REGISTRATION NUMBER: NCT01433523.

Protein-Protein interactive networks identified in bronchoalveolar lavage of severe compared to nonsevere asthma.

INTRODUCTION: Previous bronchoalveolar lavage fluid (BALF) proteomic analysis has evaluated limited numbers of subjects for only a few proteins of interest, which may differ between asthma and normal controls. Our objective was to examine a more comprehensive inflammatory biomarker panel in quantitative proteomic analysis for a large asthma cohort to identify molecular phenotypes distinguishing severe from nonsevere asthma. METHODS: Bronchoalveolar lavage fluid from 48 severe and 77 nonsevere adult asthma subjects were assessed for 75 inflammatory proteins, normalized to BALF total protein concentration. Validation of BALF differences was sought through equivalent protein analysis of autologous sputum. Subjects' data, stratified by asthma severity, were analysed by standard statistical tests, principal component analysis and 5 machine learning algorithms. RESULTS: The severe group had lower lung function and greater health care utilization. Significantly increased BALF proteins for severe asthma compared to nonsevere asthma were fibroblast growth factor 2 (FGF2), TGFα, IL1Ra, IL2, IL4, CCL8, CCL13 and CXCL7 and significantly decreased were platelet-derived growth factor a-a dimer (PDGFaa), vascular endothelial growth factor (VEGF), interleukin 5 (IL5), CCL17, CCL22, CXCL9 and CXCL10. Four protein differences were replicated in sputum. FGF2, PDGFaa and CXCL7 were independently identified by 5 machine learning algorithms as the most important variables for discriminating severe and nonsevere asthma. Increased and decreased proteins identified for the severe cluster showed significant protein-protein interactions for chemokine and cytokine signalling, growth factor activity, and eosinophil and neutrophil chemotaxis differing between subjects with severe and nonsevere asthma. CONCLUSION: These inflammatory protein results confirm altered airway remodelling and cytokine/chemokine activity recruiting leukocytes into the airways of severe compared to nonsevere asthma as important processes even in stable status.

Low CC16 mRNA Expression Levels in Bronchial Epithelial Cells Are Associated with Asthma Severity.

Rationale: CC16 is a protein mainly produced by nonciliated bronchial epithelial cells (BECs) that participates in host defense. Reduced CC16 protein concentrations in BAL and serum are associated with asthma susceptibility. Objectives: Few studies have investigated the relationship between CC16 and asthma progression, and none has focused on BECs. In this study, we sought to determine if CC16 mRNA expression levels in BECs are associated with asthma severity. Methods: Association analyses between CC16 mRNA expression levels in BECs (242 asthmatics and 69 control subjects) and asthma-related phenotypes in Severe Asthma Research Program were performed using a generalized linear model. Measurements and Main Results: Low CC16 mRNA expression levels in BECs were significantly associated with asthma susceptibility and asthma severity, high systemic corticosteroids use, high retrospective and prospective asthma exacerbations, and low pulmonary function. Low CC16 mRNA expression levels were significantly associated with high T2 inflammation biomarkers (fractional exhaled nitric oxide and sputum eosinophils). CC16 mRNA expression levels were negatively correlated with expression levels of Th2 genes (IL1RL1, POSTN, SERPINB2, CLCA1, NOS2, and MUC5AC) and positively correlated with expression levels of Th1 and inflammation genes (IL12A and MUC5B). A combination of two nontraditional T2 biomarkers (CC16 and IL-6) revealed four asthma endotypes with different characteristics of T2 inflammation, obesity, and asthma severity. Conclusions: Our findings indicate that low CC16 mRNA expression levels in BECs are associated with asthma susceptibility, severity, and exacerbations, partially through immunomodulation of T2 inflammation. CC16 is a potential nontraditional T2 biomarker for asthma development and progression.

A Polygenic Risk Score for Idiopathic Pulmonary Fibrosis and Interstitial Lung Abnormalities.

Rationale: In addition to rare genetic variants and the MUC5B locus, common genetic variants contribute to idiopathic pulmonary fibrosis (IPF) risk. The predictive power of common variants outside the MUC5B locus for IPF and interstitial lung abnormalities (ILAs) is unknown. Objectives: We tested the predictive value of IPF polygenic risk scores (PRSs) with and without the MUC5B region on IPF, ILA, and ILA progression. Methods: We developed PRSs that included (PRS-M5B) and excluded (PRS-NO-M5B) the MUC5B region (500-kb window around rs35705950-T) using an IPF genome-wide association study. We assessed PRS associations with area under the receiver operating characteristic curve (AUC) metrics for IPF, ILA, and ILA progression. Measurements and Main Results: We included 14,650 participants (1,970 IPF; 1,068 ILA) from six multi-ancestry population-based and case-control cohorts. In cases excluded from genome-wide association study, the PRS-M5B (odds ratio [OR] per SD of the score, 3.1; P = 7.1 × 10-95) and PRS-NO-M5B (OR per SD, 2.8; P = 2.5 × 10-87) were associated with IPF. Participants in the top PRS-NO-M5B quintile had ∼sevenfold odds for IPF compared with those in the first quintile. A clinical model predicted IPF (AUC, 0.61); rs35705950-T and PRS-NO-M5B demonstrated higher AUCs (0.73 and 0.7, respectively), and adding both genetic predictors to a clinical model yielded the highest performance (AUC, 0.81). The PRS-NO-M5B was associated with ILA (OR, 1.25) and ILA progression (OR, 1.16) in European ancestry participants. Conclusions: A common genetic variant risk score complements the MUC5B variant to identify individuals at high risk of interstitial lung abnormalities and pulmonary fibrosis.

Bronchial epithelial cell transcriptional responses to inhaled corticosteroids dictate severe asthmatic outcomes.

BACKGROUND: Inhaled corticosteroids (CSs) are the backbone of asthma treatment, improving quality of life, exacerbation rates, and mortality. Although effective for most, a subset of patients with asthma experience CS-resistant disease despite receiving high-dose medication. OBJECTIVE: We sought to investigate the transcriptomic response of bronchial epithelial cells (BECs) to inhaled CSs. METHODS: Independent component analysis was performed on datasets, detailing the transcriptional response of BECs to CS treatment. The expression of these CS-response components was examined in 2 patient cohorts and investigated in relation to clinical parameters. Supervised learning was used to predict BEC CS responses using peripheral blood gene expression. RESULTS: We identified a signature of CS response that was closely correlated with CS use in patients with asthma. Participants could be separated on the basis of CS-response genes into groups with high and low signature expression. Patients with low expression of CS-response genes, particularly those with a severe asthma diagnosis, showed worse lung function and quality of life. These individuals demonstrated enrichment for T-lymphocyte infiltration in endobronchial brushings. Supervised machine learning identified a 7-gene signature from peripheral blood that reliably identified patients with poor CS-response expression in BECs. CONCLUSIONS: Loss of CS transcriptional responses within bronchial epithelium was related to impaired lung function and poor quality of life, particularly in patients with severe asthma. These individuals were identified using minimally invasive blood sampling, suggesting these findings may enable earlier triage to alternative treatments.

CCL5 is a potential bridge between type 1 and type 2 inflammation in asthma.

BACKGROUND: Type 1 (T1) inflammation (marked by IFN-γ expression) is now consistently identified in subsets of asthma cohorts, but how it contributes to disease remains unclear. OBJECTIVE: We sought to understand the role of CCL5 in asthmatic T1 inflammation and how it interacts with both T1 and type 2 (T2) inflammation. METHODS: CCL5, CXCL9, and CXCL10 messenger RNA expression from sputum bulk RNA sequencing, as well as clinical and inflammatory data were obtained from the Severe Asthma Research Program III (SARP III). CCL5 and IFNG expression from bronchoalveolar lavage cell bulk RNA sequencing was obtained from the Immune Mechanisms in Severe Asthma (IMSA) cohort and expression related to previously identified immune cell profiles. The role of CCL5 in tissue-resident memory T-cell (TRM) reactivation was evaluated in a T1high murine severe asthma model. RESULTS: Sputum CCL5 expression strongly correlated with T1 chemokines (P < .001 for CXCL9 and CXCL10), consistent with a role in T1 inflammation. CCL5high participants had greater fractional exhaled nitric oxide (P = .009), blood eosinophils (P < .001), and sputum eosinophils (P = .001) in addition to sputum neutrophils (P = .001). Increased CCL5 bronchoalveolar lavage expression was unique to a previously described T1high/T2variable/lymphocytic patient group in the IMSA cohort, with IFNG trending with worsening lung obstruction only in this group (P = .083). In a murine model, high expression of the CCL5 receptor CCR5 was observed in TRMs and was consistent with a T1 signature. A role for CCL5 in TRM activation was supported by the ability of the CCR5 inhibitor maraviroc to blunt reactivation. CONCLUSION: CCL5 appears to contribute to TRM-related T1 neutrophilic inflammation in asthma while paradoxically also correlating with T2 inflammation and with sputum eosinophilia.