Lori J. Silveira

Association Between the MUC5B Promoter Polymorphism and Survival in Patients With Idiopathic Pulmonary Fibrosis

IMPORTANCE Current prediction models of mortality in idiopathic pulmonary fibrosis (IPF), which are based on clinical and physiological parameters, have modest value in predicting which patients will progress. In addition to the potential for improving prognostic models, identifying genetic and molecular features that are associated with IPF mortality may provide insight into the underlying mechanisms of disease and inform clinical trials. OBJECTIVE To determine whether the MUC5B promoter polymorphism (rs35705950), previously reported to be associated with the development of pulmonary fibrosis, is associated with survival in IPF. DESIGN, SETTING, AND PARTICIPANTS Retrospective study of survival in 2 independent cohorts of patients with IPF: the INSPIRE cohort, consisting of patients enrolled in the interferon-γ1b trial (n = 438; December 15, 2003-May 2, 2009; 81 centers in 7 European countries, the United States, and Canada), and the Chicago cohort, consisting of IPF participants recruited from the Interstitial Lung Disease Clinic at the University of Chicago (n = 148; 2007-2010). The INSPIRE cohort was used to model the association of the MUC5B genotype with survival, accounting for the effect of matrix metalloproteinase 7 (MMP-7) blood concentration and other demographic and clinical covariates. The Chicago cohort was used for replication of findings. MAIN OUTCOMES AND MEASURES The primary end point was all-cause mortality. RESULTS The numbers of patients in the GG, GT, and TT genotype groups were 148 (34%), 259 (59%), and 31 (7%), respectively, in the INSPIRE cohort and 41 (28%), 98 (66%), and 9 (6%), respectively, in the Chicago cohort. The median follow-up period was 1.6 years for INSPIRE and 2.1 years for Chicago. During follow-up, there were 73 deaths (36 GG, 35 GT, and 2 TT) among INSPIRE patients and 64 deaths (26 GG, 36 GT, and 2 TT) among Chicago patients. The unadjusted 2-year cumulative incidence of death was lower among patients carrying 1 or more copies of the IPF risk allele (T) in both the INSPIRE cohort (0.25 [95% CI, 0.17-0.32] for GG, 0.17 [95% CI, 0.11-0.23] for GT, and 0.03 [95% CI, 0.00-0.09] for TT) and the Chicago cohort (0.50 [95% CI, 0.31-0.63] for GG, 0.22 [95% CI, 0.13-0.31] for GT, and 0.11 [95% CI, 0.00-0.28] for TT). In the INSPIRE cohort, the TT and GT genotypes (risk for IPF) were associated with improved survival compared with GG (hazard ratios, 0.23 [95% CI, 0.10-0.52] and 0.48 [95% CI, 0.31-0.72], respectively; P < .001). This finding was replicated in the Chicago cohort (hazard ratios, 0.15 [95% CI, 0.05-0.49] and 0.39 [95% CI, 0.21-0.70], respectively; P < .002). The observed association of MUC5B with survival was independent of age, sex, forced vital capacity, diffusing capacity of carbon monoxide, MMP-7, and treatment status. The addition of the MUC5B genotype to the survival models significantly improved the predictive accuracy of the model in both the INSPIRE cohort (C = 0.71 [95% CI, 0.64-0.75] vs C = 0.68 [95% CI, 0.61-0.73]; P < .001) and the Chicago cohort (C = 0.73 [95% CI, 0.62-0.78] vs C = 0.69 [95% CI, 0.59-0.75]; P = .01). CONCLUSIONS AND RELEVANCE Among patients with IPF, a common risk polymorphism in MUC5B was significantly associated with improved survival. Further research is necessary to refine the risk estimates and to determine the clinical implications of these findings.

Response to sublingual immunotherapy with grass pollen extract: Monotherapy versus combination in a multiallergen extract

BACKGROUND To date, there have been no randomized, double-blind studies showing the effectiveness of sublingual immunotherapy with multiple allergens. OBJECTIVE The purpose of this study was to examine whether the efficacy of sublingual immunotherapy (SLIT) with standardized timothy extract was reduced by combination with other allergen extracts. METHODS A single-center, randomized, double-blind, placebo-controlled trial with SLIT was conducted. After an observational grass season, SLIT was administered for 10 months to 54 patients randomized to 1 of 3 treatment arms: placebo, timothy extract (19 microg Phl p 5 daily) as monotherapy, or the same dose of timothy extract plus 9 additional pollen extracts. Symptom and medication scores were collected and titrated nasal challenges, titrated skin prick tests, specific IgE, IgG4 and cytokines release by timothy-stimulated lymphocyte proliferation were performed. RESULTS Perhaps because of a very low grass pollen season in 2008, there were no significant differences in medication or symptom scores in either treatment group compared with placebo. Compared with placebo, in the timothy monotherapy group, thresholds for titrated nasal challenge and skin prick tests (P = .03 and P = .001, respectively), and serum-specific IgG4 levels (P = .005) significantly increased, and IFN- gamma levels decreased (P = .02), whereas in the multiallergen group, there was significant improvement only in the titrated skin prick tests (P = .04) which was less than in the monotherapy group. There were no significant differences between the 2 active groups in any outcome measure, and both active groups experienced more adverse events than placebo. There were no systemic reactions. CONCLUSION Improvement in multiple relevant outcomes strongly suggests that SLIT with timothy extract alone was effective; however, the results for symptom and medication scores were not significant. The differences between multiple allergen SLIT and placebo only in skin sensitivity to timothy suggest a reduction in SLIT efficacy in this group. However, further studies are required to confirm these observations.

Influence of MHC CLASS II in Susceptibility to Beryllium Sensitization and Chronic Beryllium Disease

A glutamic acid at residue 69(Glu69) in the HLA-DPB1 gene (Glu69) is associated with chronic beryllium disease (CBD) and possibly beryllium sensitization (BeS). This study tested the hypothesis that MHC class II polymorphisms are important in susceptibility to BeS and CBD and that the Glu69 variant is related to markers of disease severity. Genomic DNA was obtained from BeS (n = 50), CBD (n = 104), and beryllium-exposed nondiseased (Be-nondiseased) (n = 125) subjects. HLA-DPB1, -DRB1, and -DQB1 genotypes were determined by (sequence-specific primers) PCR. Disease severity was assessed by pulmonary function and exercise testing. A higher frequency of the DPB1 Glu69 gene was found in CBD and BeS compared with the Be-nondiseased subjects, with odds ratios of 10.1 for CBD vs Be-nondiseased and 9.5 for BeS vs Be-nondiseased. The majority of BeS and CBD subjects displayed non-0201 Glu69 alleles. Glu69 homozygosity was higher in the CBD subjects, while BeS subjects were intermediate and Be-nondiseased lowest. DRB1*01 and DQB1*05 phenotypes were reduced in CBD vs Be-nondiseased subjects, while DRB1*13 and DQB1*06 were associated with CBD in the absence of Glu69. Markers of disease severity, including a lower forced vital capacity, diffusion capacity for carbon monoxide, PaO2 at rest, maximum workload on exercise testing, and a higher arterial-alveolar gradient at rest, were associated with Glu69 homozygosity. We conclude that DPB1 Glu69 is a marker of sensitization and not specific for disease. Glu69 homozygosity acts as a functional marker associated with markers of CBD severity.

Tumour necrosis factor gene polymorphisms are associated with COPD

Tumour necrosis factor (TNF)-α has been shown to be an important factor in animal models of chronic obstructive pulmonary disease (COPD). However, human studies of TNF polymorphisms in COPD have been equivocal. Six TNF single nucleotide polymorphisms (-1031C/T, -863C/A, -857C/T, -237G/A, -308G/A and +487G/A) and their haplotypes were investigated in 423 Caucasian smokers (298 patients with spirometric evidence of COPD and 125 without airflow obstruction). The -308 minor allele (A) had a higher odds ratio (OR) of being associated with COPD in multivariate analysis (controlling for age, sex, pack-yrs; OR 1.9, 95% confidence interval (CI) 1.1–3.2) and was also associated with worse forced expiratory volume in one second/forced vital capacity. The -237 minor allele (A) had a lower OR of being associated with COPD (OR 0.40, 95% CI 0.19–0.86). In COPD patients, the -857 minor allele (T) had a lower OR of being associated with severe stages of COPD (Global Initiative for Obstructive Lung Disease stage III and IV versus stage I and II, OR 0.46, 95% CI 0.24–0.88). Other TNF single nucleotide polymorphisms were not associated with COPD but the -1031/-863 haplotype CC/TC had a lower OR in COPD patients versus smoking controls (OR 0.22, 95% CI 0.05–0.97). The present study adds further evidence that tumour necrosis factor genotypes play a role in susceptibility to cigarette smoke.

Urinary leukotriene E4 levels identify children with tobacco smoke exposure at risk for asthma exacerbation

BACKGROUND Children with asthma exposed to secondhand smoke (SHS) might be at higher risk for severe exacerbations, but biomarkers of susceptibility to SHS exposure have not been previously reported. OBJECTIVES We sought to assess the usefulness of urinary leukotriene E(4) (uLTE₄) levels in the prediction of increased risk of severe asthma exacerbations requiring emergency department (ED) or urgent care (UC) visits. METHODS Forty-four schoolchildren with moderate-to-severe asthma receiving inhaled corticosteroids were followed for 5 months with repeated measurements of uLTE₄ and monitoring of ED and UC visits. SHS exposure status was determined by using prestudy questionnaires and repeated measurements of urinary cotinine during the study. RESULTS Nine (45%) of 20 children with SHS exposure experienced a severe exacerbation requiring an ED or UC visit compared with 3 (12.5%) of 24 children without significant SHS exposure (relative risk, 3.6; 95% CI, 1.1-11.5; P = .02). The uLTE₄ level was a significant predictor of exacerbation risk in children exposed to SHS (area under the curve, 0.85; P = .003). Other predictors, such as nighttime symptom frequency, prebronchodilator and postbronchodilator lung function, and exhaled nitric oxide levels, were not related to exacerbations in this group. uLTE₄ levels at or greater than 106 pg/mg achieved 67% (6/9) sensitivity and 100% (11/11) specificity for predicting children with SHS exposure who required an ED or UC visit. CONCLUSIONS Children exposed to SHS are at increased risk for severe asthma exacerbations, despite use of inhaled corticosteroids. uLTE₄ levels identify children exposed to SHS at high risk for asthma exacerbations.

The Response of Children with Asthma to Ambient Particulate Is Modified by Tobacco Smoke Exposure

RATIONALE Ambient particulate matter concentrations have been positively associated with urinary leukotriene E(4) (LTE(4)) levels and albuterol usage in children with asthma but interactions with environmental tobacco smoke (ETS) exposure have not been demonstrated despite obvious exposure to both pollutants in an urban setting. OBJECTIVES To assess the health effects of concurrent ETS and ambient particulate matter exposure in children with asthma. METHODS Albuterol usage and LTE(4) levels were monitored in 82 urban schoolchildren with asthma over three consecutive fall to spring school periods. Concentrations of morning maximum ambient particulate matter <2.5 μm in aerodynamic diameter (mmPM(2.5)) and urine cotinine levels were also measured daily. MEASUREMENTS AND MAIN RESULTS Albuterol usage and LTE(4) were related to mmPM(2.5) concentrations on days when urine cotinine levels were low (<10 ng/ml/mg creatinine); on these days, mean albuterol usage and LTE(4) increased up to 5 or 6% per 10 μg/m(3) increase in mmPM(2.5). In contrast, no significant relationship was observed when cotinine was high, although mean albuterol usage and LTE(4) levels were greater in this case. Model fits for LTE(4) levels as a function of mmPM(2.5) concentrations were improved when mmPM(2.5) concentrations were logged, suggesting a nonlinear dose-response relationship between particulate matter exposure concentrations and airway mediators of asthma, for which the relationship tends to flatten at higher concentrations. CONCLUSIONS This study suggests that ETS modifies the acute effects of low-level ambient PM(2.5) exposure on childhood asthma. This negative interaction, the smaller effect of particulate matter exposure in children exposed to higher ETS, may be related to a nonlinear dose-response relationship between asthma mediators and particulate exposures.

TGF-β1 Variants in Chronic Beryllium Disease and Sarcoidosis

Evidence suggests a genetic predisposition to chronic beryllium disease (CBD) and sarcoidosis, which are clinically and pathologically similar granulomatous lung diseases. TGF-β1, a cytokine involved in mediating the fibrotic/Th1 response, has several genetic variants which might predispose individuals to these lung diseases. We examined whether certain TGF-β1 variants and haplotypes are found at higher rates in CBD and sarcoidosis cases compared with controls and are associated with disease severity indicators for both diseases. Using DNA from sarcoidosis cases/controls from A Case Control Etiologic Study of Sarcoidosis Group (ACCESS) and CBD cases/controls, TGF-β1 variants were analyzed by sequence-specific primer PCR. No significant differences were found between cases and controls for either disease in the TGF-β1 variants or haplotypes. The −509C and codon 10T were significantly associated with disease severity indicators in both CBD and sarcoidosis. Haplotypes that included the −509C and codon 10T were also associated with more severe disease, whereas one or more copies of the haplotype containing the −509T and codon 10C was protective against severe disease for both sarcoidosis and CBD. These studies suggest that the −509C and codon 10T, implicated in lower levels of TGF-β1 protein production, are shared susceptibility factors associated with more severe granulomatous disease in sarcoidosis and CBD. This association may be due to lack of down-regulation by TGF-β1, although future studies will be needed to correlate TGF-β1 protein levels with known TGF-β1 genotypes and assess whether there is a shared mechanisms for TGF-β1 in these two granulomatous diseases.

Risk of Chronic Beryllium Disease by HLA-DPB1 E69 Genotype and Beryllium Exposure in Nuclear Workers

RATIONALE Beryllium sensitization (BeS) and chronic beryllium disease (CBD) are determined by at least one genetic factor, a glutamic acid at position 69 (E69) of the HLA-DPB1 gene, and by exposure to beryllium. The relationship between exposure and the E69 genotype has not been well characterized. OBJECTIVES The study goal was to define the relationship between beryllium exposure and E69 for CBD and BeS. METHODS Workers (n = 386) from a U.S. nuclear weapons facility were enrolled into a case-control study (70 BeS, 61 CBD, and 255 control subjects). HLA-DPB1 genotypes were determined by sequence-specific primer-polymerase chain reaction. Beryllium exposures were reconstructed on the basis of worker interviews and historical exposure measurements. MEASUREMENTS AND MAIN RESULTS Any E69 carriage increased odds for CBD (odds ratio [OR], 7.61; 95% confidence interval [CI], 3.66-15.84) and each unit increase in lifetime weighted average exposure increased the odds for CBD (OR, 2.27; 95% CI, 1.26-4.09). Compared with E69-negative genotypes, a single E69-positive *02 allele increased the odds for BeS (OR, 12.01; 95% CI, 4.28-33.71) and CBD (OR, 3.46; 95% CI, 1.42-8.43). A single non-*02 E69 allele further increased the odds for BeS (OR, 29.54; 95% CI, 10.33-84.53) and CBD (OR, 11.97; 95% CI, 5.12-28.00) and two E69 allele copies conferred the highest odds for BeS (OR, 55.68; 95% CI, 14.80-209.40) and CBD (OR, 22.54; 95% CI, 7.00-72.62). CONCLUSIONS E69 and beryllium exposure both contribute to the odds of CBD. The increased odds for CBD and BeS due to E69 appear to be differentially distributed by genotype, with non-*02 E69 carriers and E69 homozygotes at higher odds than those with *02 genotypes.

Beryllium medical surveillance at a former nuclear weapons facility during cleanup operations.

Despite increasing need to remediate beryllium-contaminated buildings in industry, little is known about the magnitude of risk associated with beryllium abatement or the merits of beryllium medical surveillance for cleanup workers. We examined beryllium lymphocyte proliferation tests and reviewed medical evaluations on workers at a nuclear weapons facility during the process of decontamination and decommissioning. Of 2221 workers, 19 (0.8%) were beryllium sensitized based on two or more abnormal beryllium lymphocyte proliferation tests. Eight of 19 sensitized individuals underwent full clinical evaluation, of whom two were diagnosed with chronic beryllium disease (CBD). Notably, seven beryllium sensitized and CBD cases were hired after the start of cleanup operations. Beryllium medical surveillance detects sensitization and CBD in cleanup workers. Exposure controls and medical surveillance need to be ‘broad-based’ to include all cleanup workers involved in beryllium-contaminated building remediation.

Gene-environment interactions influence airways function in laboratory animal workers

BACKGROUND Most diseases, including asthma, result from the interaction between environmental exposures and genetic variants. Functional variants of CD14 negatively affect lung function in farm workers and children exposed to animal allergens and endotoxin. OBJECTIVE We hypothesized that CD14 polymorphisms interact with inhaled endotoxin, mouse allergen, or both to decrease airways function in laboratory animal workers. METHODS Three hundred sixty-nine Caucasian workers completed a symptom and work exposure questionnaire, skin prick testing, and spirometry. Individual exposure estimates for endotoxin and murine allergen were calculated by weighting task-based breathing zone concentrations by time reported for each task and length of time in the current job. Real-time PCR was used to assess CD14/-1619, -550, and -159 alleles. Multiple linear regression predicting airways function included an interaction term between genotype and exposure. RESULTS Workers at the highest quartile of the natural log-transformed cumulative endotoxin exposure and with the endotoxin-responsive CD14/-1619 G allele had significantly lower FEV(1) and forced expiratory flow, midexpiratory phase (FEF(25-75)) percent predicted compared with workers with an AA genotype, with no significant differences noted at lower endotoxin levels for either genotype. The gene-environment effect was marked for atopic workers. Laboratory animal allergy, murine allergen exposure, CD14/-159 or -550 genotype, and a gene-exposure interaction term for these genotypes and exposures did not predict changes in lung function. CONCLUSIONS A significant gene-environment interaction affects airways function in laboratory animal workers. More highly endotoxin-exposed workers with CD14/-1619G alleles have significantly lower FEV(1) and FEF(25-75) percent predicted than those with CD14/-1619AA alleles. Atopic workers are particularly affected by cumulative endotoxin exposures.