Citations

Abstract RATIONALE: Chronic Obstructive Pulmonary Disease (COPD) is a leading cause of mortality worldwide. We previously published the polygenic transcriptome risk score (PTRS) which uses the cumulative effect of predicted gene expression to construct genetic predictors for complex diseases, and we demonstrated the value of PTRS for COPD built on Genotype-Tissue Expression (GTEx) Lung tissue with significantly improved cross-ancestry portability. However, the postmortem collection and lack of lung disease status on GTEx samples likely affect results. We hypothesized that the performance of PTRS will be improved by using disease-relevant expression quantitative trait loci (eQTLs) to construct the score. Here, we aimed to improve performance of PTRS by leveraging disease-relevant eQTLs. METHODS: We constructed two transcriptome prediction models, Elastic Net (EN) and Prediction Using Models Informed by Chromatin conformation and Epigenomics (PUMICE), using TOPMed lung RNA-seq data from the Lung Tissue Research Consortium (LTRC), which included COPD patients undergoing lung surgery. We compared their performance with two published models built on RNA-seq from GTEx Lung tissue, GTEx-EN and GTEx-PUMICE. We first integrated each of four transcriptome models with multi-ancestry GWAS of FEV1/FVC ratio (Shrine et al. 2023) to generate transcriptome-wide association study (TWAS) results, which produced trait-associated genes. Pathway analysis was then conducted on significant TWAS genes. Finally, the PTRS was computed on TWAS genes that were included in the top 20 significantly enriched pathways. We tested performance of PTRS in race-stratified analysis for moderate-to-severe and severe COPD in COPDGene. RESULTS: Our models produced more significant TWAS genes (FDR<0.05) and had more overlaps with genes identified for FEV1/FVC ratio from Shrine et al. 2023 compared to two GTEx models (Fig. A). Additionally, our models had significantly higher AUC for predicting both moderate-to-severe and severe COPD in race-stratified analysis. The mean AUC from LTRC-EN for predicting severe COPD was 0.57 and 0.55 respectively for EUR and AFA, which was significantly higher than mean AUC of 0.54 and 0.51 from GTEx-EN (Delong p-value=8.15x10-10 for EUR and 1.42x10-3 for AFA) (Fig. B). Similarly, the PTRS derived from our models produced higher Odds Ratios (OR) per standard deviation. The mean OR from LTRC-PUMICE for moderate-to-severe COPD was 1.25 and 1.18 respectively for EUR and AFA, while it was 1.17 and 1.12 from GTEx-PUMICE (Fig. C). CONCLUSIONS: Our study demonstrates the value of leveraging disease-relevant RNA-seq to construct transcriptome prediction models and their improvement on identifying lung function genes and predicting COPD across ancestries.

Abstract Rationale: The club cell secretory protein (CC16), encoded by the SCGB1A1 gene, has an anti-inflammatory role in airways diseases, including asthma (Li, AJRCCM 2023; Voraphani, AJRCCM 2023). We hypothesize that additional pathway genes regulated or co-expressed with CC16 have cumulative effects on asthma severity outcomes by modulating airway inflammation and maintaining lung homeostasis. Methods: As published previously, four genes were downregulated in CC16 knockout experiments and correlated with CC16 levels: BPIFA1, SFTPD, LTF, and LYZ (Iannuzo, Front. Immunol 2023). We developed a four-gene biomarker score in 94 patients with asthma from the NHLBI-sponsored Severe Asthma Research Program (SARP3) who underwent research bronchoscopy for lower airway epithelial brushings, RNA isolation, and bulk RNA sequencing. We log-transformed the raw values of each transcript; obtained standardized residuals from regression models that included sex, age, and batch effects; and summed the standardized residuals of all four proteins to estimate a standardized sum score (Zhai, JACI Allergy 2024). Logistic regression models tested the four-gene score, three-gene score without LTF, and individual transcripts for associations with exacerbations during 12-month follow-up and zero-inflated negative binomial regression for number of exacerbations. Results: 30.85% percent of 94 patients experienced at least one exacerbation at the end of follow-up. The CC16-associated four-gene biomarker score showed protective effects on asthma exacerbation risk with a 54% percent reduction in exacerbation risk for every one-fold higher score (OR=0.46, 95% CI: 0.27-0.78, p =0.0040, Figure 1). Transcript expression of BPIFA1 (OR=0.87, 95% CI:0.66-1.16, p=0.337), SFTPD (OR=0.33, 95% CI:0.04-2.52, p=0.283), LYZ (OR=0.62, 95%CI:0.28-1.39, p=0.248), and LTF (OR=0.36, 95% CI:0.18-0.74, p=0.0053) all indicated protective directionality with respect to exacerbation risk. However, only LTF demonstrated a statistically significant association. When we excluded LTF, the remaining three-gene score showed a trend toward association with risk (OR=0.64, 95% CI:0.40-1.03, p=0.068) and was significantly associated with number of exacerbations (RR=0.49, 95% CI:0.26-0.95, p=0.034). Compared to LTF alone (OR=0.48, PPV=0.50, NPV=0.71, Sensitivity=0.14, AUC=0.65), the combined four-gene score (OR=0.46, PPV=0.60, NPV= 0.73, Sensitivity=0.21, AUC=0.68) showed improved predictive performance. Conclusions: We identified a CC16-associated transcriptomic predictive score for asthma exacerbations, with LTF showing the strongest effect. Although the other three genes showed individually weak and non-significant associations with exacerbations, the three gene score was significant for number of exacerbations.We demonstrate the potential of this transcriptomic score as a predictive biomarker for asthma severity-related outcomes. Further research investigating these genes and their interaction with other CC16-associated immune pathways could provide insight into therapeutic targets.

Abstract RATIONALE: Albuterol-budesonide 180/160 µg is indicated in the US for as-needed treatment or prevention of bronchoconstriction and to reduce risk of exacerbations in patients ≥18 years old with asthma. The objective of the ALbuterol-budesonide Treatment in Acute airway obstruction (ALTA) study (NCT05555290) is to assess comparative efficacy, safety, and mechanisms of action of albuterol-budesonide 180/160 µg versus albuterol 180 µg in treating and preventing bronchoconstriction. This report describes the clinical profiles of patients meeting criteria for randomization into the trial. METHODS: ALTA is a phase 3b, multicenter, randomized, double-blind, 2-period, cross-over study evaluating the effects of repetitive dosing of albuterol-budesonide 180/160 µg versus albuterol 180 µg for treating adults treated as intermittent asthma and experiencing acute airway obstruction induced by repeated mannitol challenges. Inclusion criteria evaluated during screening include asthma diagnosis ≥6 months, prescribed and utilizing albuterol as the only asthma treatment for ≥4 weeks, pre-bronchodilator FEV1 ≥1.50 L and FEV1 ≥60% to <90% predicted, and positive response to a mannitol challenge (decrease in FEV1≥15% at ≤635 mg cumulative mannitol dose). Exclusion criteria include clinically significant lung disease other than asthma; any smoking in the prior 6 months or ≥10 pack-year smoking history; or a clinical encounter for asthma worsening within the past 4 weeks. RESULTS: Thus far, 84 patients have met the criteria for randomization into ALTA. Mean(SD) age=44.9(13.7) years, 56.0% female, 40.5% African American, 9.5% Hispanic/Latino. Clinical characteristics include 10.7% with 1 prior-year severe exacerbation; mean(SD) fractional exhaled nitric oxide=37.0(37.4) ppb; mean(SD) baseline pre-bronchodilator FEV1=2.464(0.538) L; mean(SD) baseline percent predicted FEV1=76.7(7.9)%; mean(SD) % fall in FEV1 with mannitol challenge=22.1(7.3)% at a mean(SD) cumulative mannitol dose of 146.4(176.3) mg. Despite being treated for intermittent asthma, based upon symptoms, lung function, and exacerbation history, no patients met NAEPP 2007 criteria for intermittent disease, and 13.1% fell into the mild, 78.6% moderate, and 8.3% severe persistent asthma categories (Figure). With respect to asthma control, as measured by Asthma Impairment and Risk Questionnaire scores, 60.7% had well-controlled, 20.2% not well-controlled, and 19.1% very poorly controlled disease. CONCLUSION: The baseline characteristics of patients in the ALTA study reveal that patients treated as intermittent asthma exhibit heterogeneity in clinical characteristics, airway hyper-reactivity, T2 inflammation, and disease severity and control. Additional genomic and biomarker data are forthcoming and altogether will provide an opportunity to address the mechanism of albuterol and budesonide interactions in a broad spectrum of asthma phenotypes.

ImportanceMetastatic cutaneous Crohn disease (MCD) is a rare condition for which there are currently no published diagnostic criteria. Formal MCD diagnostic criteria will expand clinical care and research by enabling future diagnostic code validation, crystallizing a more uniform disease entity for the purposes of translational research, and allowing the development of more formalized outcome measures aimed at treatment response.ObjectiveTo define a set of criteria for the diagnosis of MCD.Evidence ReviewIn this consensus statement, a panel of experts on MCD and related conditions was assembled. Their opinions were gathered regarding the clinical and histologic findings that are sufficient, necessary, and highly suggestive of MCD through serial survey. Subsequent surveys were continued until consensus was achieved. Sufficient was defined as a feature that, if noted, would enable diagnosis of MCD without any other signs, symptoms, or testing. Necessary was defined as a feature that must be present to establish a diagnosis of MCD. Highly suggestive referred to any other features that were indicative of MCD but did not qualify as sufficient or necessary to diagnose MCD.FindingsOf 24 experts invited, 19 agreed to participate, with a 79% to 100% response rate across 5 rounds conducted. Five patients were also invited to participate, but none opted to participate, citing lack of clinical expertise and time concerns. The experts were all from the US; 5 (26%) have practiced medicine for 0 to 5 years, 5 (26%) practiced for 6 to 10 years, 6 (32%) practiced for 11 to 15 years, and 3 (16%) practiced for more than 20 years. A total of 18 experts (95%) worked in an academic practice setting. There was consensus agreement that MCD could present as oral disease, genital disease, or, rarely, in another location. Consensus was achieved on 2 major and 5 minor criteria for genital/other subtypes of MCD and 5 minor criteria for oral MCD. There was consensus that, while there are some pathological findings that are highly suggestive of MCD, histologic criteria are not necessary for diagnosis.Conclusions and RelevanceA panel of experts reached consensus on the diagnostic criteria for MCD described in this study. These criteria include major and minor clinical criteria, along with histopathological criteria for the diagnosis of genital and oral MCD. This consensus is a crucial step in the diagnosis of and future treatment development for MCD, a rare and clinically understudied condition.

Background: Despite the availability of effective vaccines and a recent decrease in annual deaths, coronavirus disease-2019 (COVID-19) remains a leading cause of death. Serological studies of anti-nucleocapsid (N) antibody response to infection provide insights into host immunobiology of adaptive immune response, which holds promise for identifying high-risk individuals for adverse acute and chronic COVID-19 outcomes. Hypothesis: Among participants previously infected with SARS-CoV-2, traditional vascular disease risk factors are associated with higher anti-N antibodies while vaccination is associated with lower anti-N antibodies. Methods: Among previously infected participants, anti-N antibodies were measured from dried blood spots collected between February 2021-February 2023 among 1,419 Collaborative Cohort of Cohorts for COVID-19 Research (C4R) participants with prior SARS-CoV-2 infection. We measured anti-N IgG antibodies reported as median fluorescence intensity (MFI) units; reactivity (i.e., seropositivity) was defined based on MFI above a validated threshold. Vascular disease risk factors were assessed via pre-pandemic in-person examination, questionnaires, and medical record review. Multivariable generalized linear models regressed anti-N reactivity (modified Poisson) or LN transformed MFI (linear). Results: Among 1,419 participants with prior infection, mean age (standard deviation) was 65.8(12.1) years, 61% were women, and 42.8% were self-reported from a race/ethnicity minority group. Participant reactivity to nucleocapsid peaked at 69% by 4 months post-infection and waned to only 44% ≥12 months after infection. After multivariable adjustment, higher anti-N antibody response was associated with older age, Hispanic (vs. White) or American Indian (vs. White) race/ethnicity, lower income and education, former smoking, and higher anti-spike antibody levels. Asian race (vs. White) and vaccination ( even after infection ) were associated with lower nucleocapsid reactivity; common cardiometabolic co-morbidities were not associated with anti-N reactivity or levels (Figures 1 and 2). Conclusions: Among participants previously infected with SARS-CoV-2, select sociodemographic and behavioral risk factors were associated with higher anti-N antibody levels while vaccination was associated with lower anti-N antibody levels. The observation that vaccination, even after infection, is related to lower anti-N antibody levels merits further investigation.

Abstract Rationale Increased risk of coronavirus disease (COVID-19) hospitalization and death has been reported among patients with clinical lung disease. Objectives To test the association of objective measures of prepandemic lung function and structure with COVID-19 outcomes in U.S. adults. Methods Prepandemic obstruction (FEV1/FVC < 0.70) and restriction (FEV1/FVC ⩾ 0.7, FVC < 80%) were defined based on the most recent spirometry exam conducted in 11 prospective U.S. general population–based cohorts. Severe obstruction was classified by FEV1 < 50%. Percentage emphysema, percentage high-attenuation areas, and interstitial lung abnormalities were defined on computed tomography in a subset. Incident COVID-19 was ascertained via questionnaires, serosurvey, and medical records from 2020 to 2023 and classified as severe (hospitalized or fatal) or nonsevere. Cause-specific hazard models were adjusted for sociodemographics, anthropometry, smoking, comorbidities, and COVID-19 vaccination status. Measurements and Main Results Among 29,323 participants (mean age, 67 yr), there were 748 severe incident COVID-19 cases over median follow-up of 17.3 months from March 1, 2020. Greater hazards of severe COVID-19 were associated with severe obstruction (vs. normal; adjusted hazard ratio [aHR], 2.11; 95% confidence interval [CI], 1.02–1.27), restriction (vs. normal; aHR, 1.40; 95% CI, 1.12–1.76), and percentage emphysema (highest vs. lowest quartile; aHR, 1.64; 95% CI, 1.03–2.61), but not greater high-attenuation areas or interstitial lung abnormalities. COVID-19 vaccination provided greater absolute risk reduction in these groups. Results were similar in participants without smoking, obesity, or clinical cardiopulmonary disease. Conclusions Prepandemic severe spirometric obstruction, spirometric restriction, and greater percentage emphysema lung on computed tomography were associated with risk of severe COVID-19. These findings support enhanced COVID-19 risk mitigation for individuals with impaired lung health and warrant further mechanistic studies on interactions of lung function, structure, and vulnerability to acute respiratory illnesses.

Objectives/Goals: Aspiration causes or aggravates lung diseases. While bedside swallow evaluations are not sensitive/specific, gold standard tests for aspiration are invasive, uncomfortable, expose patients to radiation, and are resource intensive. We propose the development and validation of an AI model that analyzes voice to noninvasively predict aspiration. Methods/Study Population: Retrospectively recorded [i] phonations from 163 unique ENT patients were analyzed for acoustic features including jitter, shimmer, harmonic to noise ratio (HNR), etc. Patients were classified into three groups: aspirators (Penetration-Aspiration Scale, PAS 6–8), probable (PAS 3–5), and non-aspirators (PAS 1–2) based on video fluoroscopic swallow (VFSS) findings. Multivariate analysis evaluated patient demographics, history of head and neck surgery, radiation, neurological illness, obstructive sleep apnea, esophageal disease, body mass index, and vocal cord dysfunction. Supervised machine learning using five folds cross-validated neural additive network modelling (NAM) was performed on the phonations of aspirator versus non-aspirators. The model was then validated using an independent, external database. Results/Anticipated Results: Aspirators were found to have quantifiably worse quality of sound with higher jitter and shimmer but lower harmonics noise ratio. NAM modeling classified aspirators and non-aspirators as distinct groups (aspirator NAM risk score 0.528+0.2478 (mean + std) vs. non-aspirator (control) risk score of 0.252+0.241 (mean + std); p Discussion/Significance of Impact: We report the use of voice as a novel, noninvasive biomarker to detect aspiration risk using machine learning techniques. This tool has the potential to be used for the safe and early detection of aspiration in a variety of clinical settings including intensive care units, wards, outpatient clinics, and remote monitoring.

Objectives/Goals: Primary sclerosing cholangitis (PSC) manifests with an inflammatory milieu that leads to fibrotic scarring of the liver. Human PSC liver bile ducts are enriched with neutrophils; however, their infiltration and functional role is unexplored. Our goal is to investigate the mechanism and impact of peribiliary neutrophil infiltration observed in PSC. Methods/Study Population: Primary cholangiocytes (bile duct cells) isolated from WT and mouse models of PSC (3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)-fed mice and Mdr2-/- mice) were analyzed by RNA-sequencing. Immunofluorescence (IF) was performed on liver tissues from PSC patients and mouse models of PSC for markers of bile ducts and neutrophils (KRT19 and MPO). Intrahepatic leukocytes (IHL) isolated from mice livers were evaluated for neutrophil abundance and activation state. Anti-Ly6G antibody-mediated neutrophil depletion in Mdr2-/- mice was analyzed by IF, histology, and cytometry by time-of-flight (CyTOF). Cholangiocytes stimulated with TNFα (to induce an inflammatory phenotype) were analyzed for neutrophil chemoattractants with genetic and pharmacological interventions. Results/Anticipated Results: RNA-seq analysis of primary cholangiocytes from PSC mouse models revealed enrichment in inflammatory and neutrophil degranulation pathways. Flow cytometry and RT-PCR analysis revealed an increase in the neutrophil population in PSC mice with activated phenotype. Peripheral depletion of neutrophils in Mdr2-/- mice alleviated liver injury and inflammation, along with a reduction in peribiliary neutrophil infiltration and attenuated bridging fibrosis. CyTOF analysis showed a significant reduction in CD8+ T cells upon neutrophil depletion, implying neutrophils sustain CD8+ T cells in PSC liver. Mechanistically in cholangiocytes, TNFα mediates expression of neutrophil chemoattractants, CXCL1 and CXCL8, through the cyclic GMP-AMP synthase stimulator of interferon genes (cGAS-STING) pathway. Discussion/Significance of Impact: Our findings suggest that activation of the STING pathway in cholangiocytes in cholestatic liver disease triggers an immune response resulting in peri-portal neutrophil infiltration via CXC chemokines. The sustained presence of these activated neutrophils engages the adaptive immune system to perpetuate the inflammation and fibrosis seen in PSC.

Abstract Despite the availability of effective vaccines and a recent decrease in annual deaths, COVID-19 remains a leading cause of death. Serological studies provide insights into host immunobiology of adaptive immune response to infection, which holds promise for identifying high-risk individuals for adverse COVID-19 outcomes. We investigated correlates of anti-nucleocapsid antibody responses following SARS-CoV-2 infection in a US population-based meta-cohort of adults participating in longstanding National Institutes of Health–funded cohort studies. Anti-nucleocapsid antibodies were measured from dried blood spots collected between February 2021 and February 2023. Among 1419 Collaborative Cohort of Cohorts for COVID-19 Research participants with prior SARS-CoV-2 infection, the mean age (standard deviation) was 65.8 (12.1), 61% were women, and 42.8% self-reported membership in a race/ethnicity minority group. The proportion of participants reactive to nucleocapsid peaked at 69% by 4 months after infection and waned to only 44% ≥12 months after infection. Higher anti-nucleocapsid antibody response was associated with older age, Hispanic or American Indian Alaskan Native (vs White) race/ethnicity, lower income, lower education, former smoking, and higher anti-spike antibody levels. Asian race (vs White) and vaccination (even after infection) were associated with lower nucleocapsid reactivity. Neither vaccine manufacturer nor common cardiometabolic comorbidities were not associated with anti-nucleocapsid response. These findings inform the underlying immunobiology of adaptive immune response to infection, as well as the potential utility of anti-nucleocapsid antibody response for clinical practice and COVID-19 serosurveillance.