Autism-associated neuroligin-3 mutations commonly impair striatal circuits to boost repetitive behaviors.

In humans, neuroligin-3 mutations are associated with autism, whereas in mice, the corresponding mutations produce robust synaptic and behavioral changes. However, different neuroligin-3 mutations cause largely distinct phenotypes in mice, and no causal relationship links a specific synaptic dysfunction to a behavioral change. Using rotarod motor learning as a proxy for acquired repetitive behaviors in mice, we found that different neuroligin-3 mutations uniformly enhanced formation of repetitive motor routines. Surprisingly, neuroligin-3 mutations caused this phenotype not via changes in the cerebellum or dorsal striatum but via a selective synaptic impairment in the nucleus accumbens/ventral striatum. Here, neuroligin-3 mutations increased rotarod learning by specifically impeding synaptic inhibition onto D1-dopamine receptor-expressing but not D2-dopamine receptor-expressing medium spiny neurons. Our data thus suggest that different autism-associated neuroligin-3 mutations cause a common increase in acquired repetitive behaviors by impairing a specific striatal synapse and thereby provide a plausible circuit substrate for autism pathophysiology.

Fulfilling the Promise of Breathomics: Considerations for the Discovery and Validation of Exhaled Volatile Biomarkers.

The exhaled breath represents an ideal matrix for non-invasive biomarker discovery, and exhaled metabolomics have the potential to be clinically useful in the era of precision medicine. In this concise translational review we will specifically address volatile organic compounds in the breath, with a view towards fulfilling the promise of these as actionable biomarkers, in particular for lung diseases. We review the literature paying attention to seminal work linked to key milestones in breath research; discuss potential applications for breath biomarkers across disease areas and healthcare systems, including the perspectives of industry; and outline critical aspects of study design that will need to be considered for any pivotal research going forward, if breath analysis is to provide robust validated biomarkers that meet the requirements for future clinical implementation.

Exhaled Volatile Organic Compounds for Asthma Control Classification in Children with Moderate to Severe Asthma: Results from the SysPharmPediA Study.

RATIONALE: Early identification of children with poorly controlled asthma is imperative for optimizing treatment strategies. The analysis of exhaled volatile organic compounds (VOCs) is an emerging approach to identify prognostic and diagnostic biomarkers in pediatric asthma. OBJECTIVES: To assess the accuracy of gas chromatography-mass spectrometry based exhaled metabolite analysis to differentiate between controlled and uncontrolled pediatric asthma. METHODS: This study encompassed a discovery (SysPharmPediA) and validation phase (U-BIOPRED, PANDA). Firstly, exhaled VOCs that discriminated asthma control levels were identified. Subsequently, outcomes were validated in two independent cohorts. Patients were classified as controlled or uncontrolled, based on asthma control test scores and number of severe attacks in the past year. Additionally, potential of VOCs in predicting two or more future severe asthma attacks in SysPharmPediA was evaluated. MEASUREMENTS AND MAIN RESULTS: Complete data were available for 196 children (SysPharmPediA=100, U-BIOPRED=49, PANDA=47). In SysPharmPediA, after randomly splitting the population into training (n=51) and test sets (n=49), three compounds (acetophenone, ethylbenzene, and styrene) distinguished between uncontrolled and controlled asthmatics. The area under the receiver operating characteristic curve (AUROCC) for training and test sets were respectively: 0.83 (95% CI: 0.65-1.00) and 0.77 (95% CI: 0.58-0.96). Combinations of these VOCs resulted in AUROCCs of 0.74 ±0.06 (UBIOPRED) and 0.68 ±0.05 (PANDA). Attacks prediction tests, resulted in AUROCCs of 0.71 (95% CI 0.51-0.91) and 0.71 (95% CI 0.52-0.90) for training and test sets. CONCLUSIONS: Exhaled metabolites analysis might enable asthma control classification in children. This should stimulate further development of exhaled metabolites-based point-of-care tests in asthma.

Biomarker Predictors of Clinical Efficacy of the Anti-IgE Biologic Omalizumab in Severe Asthma in Adults: Results of the SoMOSA Study.

Background: The anti-IgE monoclonal antibody omalizumab is widely used for severe asthma. This study aimed to identify biomarkers that predict clinical improvement during 1 year of omalizumab treatment. Methods: One-year open-label Study of Mechanisms of action of Omalizumab in Severe Asthma (SoMOSA) involving 216 patients with severe (Global Initiative for Asthma step 4/5) uncontrolled atopic asthma (at least two severe exacerbations in the previous year) taking high-dose inhaled corticosteroids and long-acting ß-agonists with or without maintenance oral corticosteroids. It had two phases: 0-16 weeks, to assess early clinical improvement by Global Evaluation of Therapeutic Effectiveness (GETE); and 16-52 weeks, to assess late responses based on ⩾50% reduction in exacerbations or mOCS dose. All participants provided samples (exhaled breath, blood, sputum, urine) before and after 16 weeks of omalizumab treatment. Measurements and Main Results: A total of 191 patients completed phase 1; 63% had early improvement. Of 173 who completed phase 2, 69% had reduced exacerbations by ⩾50% and 57% (37 of 65) taking mOCSs had reduced their dose by ⩾50%. The primary outcomes 2,3-dinor-11-ß-PGF2α, GETE score, and standard clinical biomarkers (blood and sputum eosinophils, exhaled nitric oxide, serum IgE) did not predict either clinical response. Five volatile organic compounds and five plasma lipid biomarkers strongly predicted the ⩾50% reduction in exacerbations (receiver operating characteristic areas under the curve of 0.780 and 0.922, respectively) and early responses (areas under the curve of 0.835 and 0.949, respectively). In an independent cohort, gas chromatography/mass spectrometry biomarkers differentiated between severe and mild asthma. Conclusions: This is the first discovery of omics biomarkers that predict improvement in asthma with biologic agent treatment. Prospective validation and development for clinical use is justified.

Physiologically Based Pharmacokinetic Modeling to Predict the Impact of Liver Cirrhosis on Glucuronidation via UGT1A4 and UGT2B7/2B4-A Case Study with Midazolam.

Hepatic impairment, due to liver cirrhosis, decreases the activity of cytochrome P450 enzymes (CYPs). The use of physiologically based pharmacokinetic (PBPK) modeling to predict this effect for CYP substrates has been well-established, but the effect of cirrhosis on uridine-glucuronosyltransferase (UGT) activities is less studied and few PBPK models have been reported. UGT enzymes are involved in primary N-glucuronidation of midazolam and glucuronidation of 1'-OH-midazolam following CYP3A hydroxylation. In this study, Simcyp was used to establish PBPK models for midazolam, its primary metabolites midazolam-N-glucuronide (UGT1A4) and 1'-OH midazolam (CYP3A4/3A5), and the secondary metabolite 1'-OH-midazolam-O-glucuronide (UGT2B7/2B4), allowing to simulate the impact of liver cirrhosis on the primary and secondary glucuronidation of midazolam. The model was verified in noncirrhotic subjects before extrapolation to cirrhotic patients of Child-Pugh (CP) classes A, B, and C. Our model successfully predicted the exposures of midazolam and its metabolites in noncirrhotic and cirrhotic patients, with 86% of observed plasma concentrations within 5th-95th percentiles of predictions and observed geometrical mean of area under the plasma concentration curve between 0 hours to infinity and maximal plasma concentration within 0.7- to 1.43-fold of predictions. The simulated metabolic ratio defined as the ratio of the glucuronide metabolite AUC over the parent compound AUC (AUCglucuronide/AUCparent, metabolic ratio [MR]), was calculated for midazolam-N-glucuronide to midazolam (indicative of UGT1A4 activity) and decreased by 40% (CP A), 48% (CP B), and 75% (CP C). For 1'-OH-midazolam-O-glucuronide to 1'-OH-midazolam, the MR (indicative of UGT2B7/2B4 activity) dropped by 35% (CP A), 51% (CP B), and 64% (CP C). These predicted MRs were corroborated by the observed data. This work thus increases confidence in Simcyp predictions of the effect of liver cirrhosis on the pharmacokinetics of UGT1A4 and UGT2B7/UGT2B4 substrates. SIGNIFICANCE STATEMENT: This article presents a physiologically based pharmacokinetic model for midazolam and its metabolites and verifies the accurate simulation of pharmacokinetic profiles when using the Simcyp hepatic impairment population models. Exposure changes of midazolam-N-glucuronide and 1'-OH-midazolam-O-glucuronide reflect the impact of decreases in UGT1A4 and UGT2B7/2B4 glucuronidation activity in cirrhotic patients. The approach used in this study may be extended to verify the modeling of other uridine glucuronosyltransferase enzymes affected by liver cirrhosis.

Expiratory flow limitation development index (ELDI): a novel method of assessing respiratory mechanics in COPD.

BACKGROUND: Expiratory flow limitation (EFL) can be detected using oscillometric reactance and is associated with a worse clinical presentation in chronic obstructive pulmonary disease (COPD). Reactance can show negative swings upon exhalation, which may develop at different rates between patients. We propose a new method to quantify the rate of EFL development; the EFL Development Index (ELDI). METHODS: A retrospective analysis of data from 124 COPD patients was performed. Data included lung function tests, Impulse Oscillometry (IOS), St Georges Respiratory Questionnaire (SGRQ), modified Medical Research Council (mMRC) scale and COPD Assessment Test (CAT) score. Fifty four patients had repeat data after 6 months. Twenty two patients had data recorded after 5 days of treatment with long acting bronchodilator therapy. EDLI was calculated as the mean expiratory reactance divided by the minimum expiratory reactance. RESULTS: The mean ELDI was used to categorise patients with rapid onset of EFL (> 0.63; n = 29) or gradual onset (≤ 0.63; n = 34). Those with rapid development had worse airflow obstruction, lower quality of life scores, and greater resting hyperinflation, compared to those with gradual development. In patients with EFL, ELDI correlated with symptoms scores, airflow obstruction, lung volumes and gas diffusion. Both EFL and ELDI were stable over 6 months. EFL and EDLI improved with bronchodilator treatment. CONCLUSIONS: COPD patients with rapid EFL development (determined by ELDI) had worse clinical characteristics than those with gradual EFL development. The rate of EFL development appears to be associated with clinical and physiological characteristics.

Volatilomes of human infection.

The human volatilome comprises a vast mixture of volatile emissions produced by the human body and its microbiomes. Following infection, the human volatilome undergoes significant shifts, and presents a unique medium for non-invasive biomarker discovery. In this review, we examine how the onset of infection impacts the production of volatile metabolites that reflects dysbiosis by pathogenic microbes. We describe key analytical workflows applied across both microbial and clinical volatilomics and emphasize the value in linking microbial studies to clinical investigations to robustly elucidate the metabolic species and pathways leading to the observed volatile signatures. We review the current state of the art across microbial and clinical volatilomics, outlining common objectives and successes of microbial-clinical volatilomic workflows. Finally, we propose key challenges, as well as our perspectives on emerging opportunities for developing clinically useful and targeted workflows that could significantly enhance and expedite current practices in infection diagnosis and monitoring.

Illness perceptions, symptom severity and psychosocial outcomes in adults with dysfunctional breathing.

BACKGROUND: People with dysfunctional breathing (DB) experience symptoms such as air hunger and breathing pattern irregularities. The condition is often comorbid with other respiratory conditions, as well as anxiety and depression. Illness perceptions, the beliefs an individual has of an illness may explain health and wellbeing outcomes. METHODS: In this cross-sectional study we examined the illness perceptions of those diagnosed with DB, symptom severity, and psychosocial outcomes of depression, anxiety, and impact on daily living. Data were analyzed using tests of comparison and regression analysis. RESULTS: 82 people diagnosed with DB completed the brief illness perception questionnaire, the Nijmegen symptoms questionnaire, and questionnaires measuring mood and impact on daily living. The illness perceptions of those with DB were overall negative. There was a positive correlation between illness perceptions and mood, indicating that the stronger the beliefs that individuals had that DB is a serious condition, the more negative their mood. Illness perceptions significantly predicted psychosocial outcomes, even when controlling for demographic factors and symptom severity (depression: adj. R2=.352, F(10,51)=4.32, p<.001; anxiety: adj. R2=.40, F(11,47)=4.55, p<.001; impact on daily living: adj. R2= .33, F(8,53)=4.79, p<.001). CONCLUSIONS: This is the first study to examine illness perceptions held by those diagnosed with DB. Our study found significant relationships between illness perceptions and psychosocial outcomes. It is possible that psychological interventions that target illness perceptions may also improve outcomes.

Asthma innovations from the second International Collaborative Asthma Network (ICAN) forum.

Asthma continues to cause morbidity and mortality despite advances in treatment that include biologics targeting Type 2 inflammation. The International Collaborative Asthma Network (ICAN) forum was developed with the primary goal of promoting innovative, collaborative research that focuses on mechanisms and treatment for asthma that does not respond or that responds poorly to currently available treatments. The mission of ICAN is innovation, collaboration, translation, and increasing high quality research. At the second ICAN meeting, presenters covered a broad scope and depth of asthma-related topics in the categories of complex data, novel therapeutics and diagnostics, breath analysis and microbiome, disease mechanisms, systemic effects, and circadian rhythm. Key actionable needs and research topics were identified during the group discussions. The presentations and discussions that occurred at the second ICAN had an immediate impact on asthma research in the form of new collaborations and implementation of new research ideas and techniques. The forum also served to connect early-stage investigators with investigators who are well established, thereby fostering innovation, translation, and collaboration well into the future. A third ICAN meeting is planned for 2025 to further the innovations and collaborations that will translate into novel therapies and diagnostics to improve the lives of patients with asthma.

Oropharyngeal Microbiota Clusters in Children with Asthma or Wheeze Associate with Allergy, Blood Transcriptomic Immune Pathways, and Exacerbation Risk.

Rationale: Children with preschool wheezing or school-age asthma are reported to have airway microbial imbalances. Objectives: To identify clusters in children with asthma or wheezing using oropharyngeal microbiota profiles. Methods: Oropharyngeal swabs from the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) pediatric asthma or wheezing cohort were characterized using 16S ribosomal RNA gene sequencing, and unsupervised hierarchical clustering was performed on the Bray-Curtis ß-diversity. Enrichment scores of the Molecular Signatures Database hallmark gene sets were computed from the blood transcriptome using gene set variation analysis. Children with severe asthma or severe wheezing were followed up for 12-18 months, with assessment of the frequency of exacerbations. Measurements and Main Results: Oropharyngeal samples from 241 children (age range, 1-17 years; 40% female) revealed four taxa-driven clusters dominated by Streptococcus, Veillonella, Rothia, and Haemophilus. The clusters showed significant differences in atopic dermatitis, grass pollen sensitization, FEV1% predicted after salbutamol, and annual asthma exacerbation frequency during follow-up. The Veillonella cluster was the most allergic and included the highest percentage of children with two or more exacerbations per year during follow-up. The oropharyngeal clusters were different in the enrichment scores of TGF-ß (transforming growth factor-ß) (highest in the Veillonella cluster) and Wnt/ß-catenin signaling (highest in the Haemophilus cluster) transcriptomic pathways in blood (all q values <0.05). Conclusions: Analysis of the oropharyngeal microbiota of children with asthma or wheezing identified four clusters with distinct clinical characteristics (phenotypes) that associate with risk for exacerbation and transcriptomic pathways involved in airway remodeling. This suggests that further exploration of the oropharyngeal microbiota may lead to novel pathophysiologic insights and potentially new treatment approaches.

Protocol for a scoping review to map health outcomes in individuals with inducible laryngeal obstruction.

BACKGROUND: Inducible laryngeal obstruction causes narrowing of the laryngeal aperture in response to external triggers. Outcomes are measured in inducible laryngeal obstruction to monitor changes in health status over time. METHODS: This study is a scoping review based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) methodology. The review will be guided by the following research question: 'What health outcomes are measured in studies including people with inducible laryngeal obstruction?' The research question was validated using the Population-Concept-Context framework according to the methodology for Joanna Briggs Institution Scoping Reviews. Relevant peer-reviewed studies and grey literature conducted over the last 40 years will be identified from electronic databases including AMED, CINAHL, Embase, EMCARE, MEDLINE, OVID, PubMed and PsycINFO. The search strings 'inducible laryngeal obstruction', 'ILO', 'vocal cord dysfunction', 'VCD', 'paradoxical vocal fold motion', 'PVFM', 'outcome', 'measure', 'measurement instrument', 'assessment', 'scale', 'questionnaire' will be combined using Boolean logic. An independent reviewer will conduct title screening; two independent reviewers will conduct abstract and full article screening, followed by data extraction by two reviewers. Analyses will be conducted appropriate to the findings. DISCUSSION: The review will document evidence of health outcomes measured in inducible laryngeal obstruction, identifying measurement characteristics and potential utility. Collating studies may identify gaps in coverage, the need for novel tools, and for standardisation for clinical and research purposes. WHAT THIS PAPER ADDS: What is already known on the subject Inducible laryngeal obstruction causes narrowing of the laryngeal aperture in response to external triggers. Outcomes are measured in inducible laryngeal obstruction to monitor changes in health status over time. Currently, there are no standardised outcome measures for measuring the effects of interventions in inducible laryngeal obstruction (ILO). What this paper adds to existing knowledge Assessment of health can be measured in a variety of ways. Physiological, radiological and biochemical measurements of impairment are more common historically but there are a lot of outcomes of other factors now including subjective measures of functional status and health-related quality of life, with data collected directly from patients. This study will allow us to scope the literature to see the health outcomes being measured in ILO to attempt to standardise and develop future health outcomes. What are the potential or actual clinical implications of this work? The review will document evidence of health outcomes measured in inducible laryngeal obstruction, identifying measurement characteristics and potential utility. Collating studies may identify gaps in coverage, the need for novel tools and for standardisation for clinical and research purposes.

Stratification of asthma by lipidomic profiling of induced sputum supernatant.

BACKGROUND: Asthma is a chronic respiratory disease with significant heterogeneity in its clinical presentation and pathobiology. There is need for improved understanding of respiratory lipid metabolism in asthma patients and its relation to observable clinical features. OBJECTIVE: We performed a comprehensive, prospective, cross-sectional analysis of the lipid composition of induced sputum supernatant obtained from asthma patients with a range of disease severities, as well as from healthy controls. METHODS: Induced sputum supernatant was collected from 211 adults with asthma and 41 healthy individuals enrolled onto the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) study. Sputum lipidomes were characterized by semiquantitative shotgun mass spectrometry and clustered using topologic data analysis to identify lipid phenotypes. RESULTS: Shotgun lipidomics of induced sputum supernatant revealed a spectrum of 9 molecular phenotypes, highlighting not just significant differences between the sputum lipidomes of asthma patients and healthy controls, but also within the asthma patient population. Matching clinical, pathobiologic, proteomic, and transcriptomic data helped inform the underlying disease processes. Sputum lipid phenotypes with higher levels of nonendogenous, cell-derived lipids were associated with significantly worse asthma severity, worse lung function, and elevated granulocyte counts. CONCLUSION: We propose a novel mechanism of increased lipid loading in the epithelial lining fluid of asthma patients resulting from the secretion of extracellular vesicles by granulocytic inflammatory cells, which could reduce the ability of pulmonary surfactant to lower surface tension in asthmatic small airways, as well as compromise its role as an immune regulator.

Microbial Volatiles as Diagnostic Biomarkers of Bacterial Lung Infection in Mechanically Ventilated Patients.

BACKGROUND: Early and accurate recognition of respiratory pathogens is crucial to prevent increased risk of mortality in critically ill patients. Microbial-derived volatile organic compounds (mVOCs) in exhaled breath could be used as noninvasive biomarkers of infection to support clinical diagnosis. METHODS: In this study, we investigated the diagnostic potential of in vitro-confirmed mVOCs in the exhaled breath of patients under mechanical ventilation from the BreathDx study. Samples were analyzed by thermal desorption-gas chromatography-mass spectrometry. RESULTS: Pathogens from bronchoalveolar lavage (BAL) cultures were identified in 45 of 89 patients and Staphylococcus aureus was the most commonly identified pathogen (n = 15). Of 19 mVOCs detected in the in vitro culture headspace of 4 common respiratory pathogens (S. aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae, and Escherichia coli), 14 were found in exhaled breath samples. Higher concentrations of 2 mVOCs were found in the exhaled breath of patients infected with S. aureus compared to those without (3-methylbutanal: P < .01, area under the receiver operating characteristic curve [AUROC] = 0.81-0.87; and 3-methylbutanoic acid: P = .01, AUROC = 0.79-0.80). In addition, bacteria identified from BAL cultures that are known to metabolize tryptophan (E. coli, Klebsiella oxytoca, and Haemophilus influenzae) were grouped and found to produce higher concentrations of indole compared to breath samples with culture-negative (P = .034) and other pathogen-positive (P = .049) samples. CONCLUSIONS: This study demonstrates the capability of using mVOCs to detect the presence of specific pathogen groups with potential to support clinical diagnosis. Although not all mVOCs were found in patient samples within this small pilot study, further targeted and qualitative investigation is warranted using multicenter clinical studies.

Changes in exhaled volatile organic compounds following indirect bronchial challenge in suspected asthma.

BACKGROUND: Inhaled mannitol provokes bronchoconstriction via mediators released during osmotic degranulation of inflammatory cells, and, hence represents a useful diagnostic test for asthma and model for acute attacks. We hypothesised that the mannitol challenge would trigger changes in exhaled volatile organic compounds (VOCs), generating both candidate biomarkers and novel insights into their origin. METHODS: Participants with a clinical diagnosis of asthma, or undergoing investigation for suspected asthma, were recruited. Inhaled mannitol challenges were performed, followed by a sham challenge after 2 weeks in participants with bronchial hyper-responsiveness (BHR). VOCs were collected before and after challenges and analysed using gas chromatography-mass spectrometry. RESULTS: Forty-six patients (mean (SD) age 52 (16) years) completed a mannitol challenge, of which 16 (35%) were positive, and 15 of these completed a sham challenge. Quantities of 16 of 51 identified VOCs changed following mannitol challenge (p<0.05), of which 11 contributed to a multivariate sparse partial least square discriminative analysis model, with a classification error rate of 13.8%. Five of these 16 VOCs also changed (p<0.05) in quantity following the sham challenge, along with four further VOCs. In patients with BHR to mannitol distinct postchallenge VOC signatures were observed compared with post-sham challenge. CONCLUSION: Inhalation of mannitol was associated with changes in breath VOCs, and in people with BHR resulted in a distinct exhaled breath profile when compared with a sham challenge. These differentially expressed VOCs are likely associated with acute airway inflammation and/or bronchoconstriction and merit further investigation as potential biomarkers in asthma.

Development of Core Outcome Measures sets for paediatric and adult Severe Asthma (COMSA).

BACKGROUND: Effectiveness studies with biological therapies for asthma lack standardised outcome measures. The COMSA (Core Outcome Measures sets for paediatric and adult Severe Asthma) Working Group sought to develop Core Outcome Measures (COM) sets to facilitate better synthesis of data and appraisal of biologics in paediatric and adult asthma clinical studies. METHODS: COMSA utilised a multi-stakeholder consensus process among patients with severe asthma, adult and paediatric clinicians, pharmaceutical representatives, and health regulators from across Europe. Evidence included a systematic review of development, validity and reliability of selected outcome measures plus a narrative review and a pan-European survey to better understand patients' and carers' views about outcome measures. It was discussed using a modified GRADE (Grading of Recommendations Assessment, Development and Evaluation) Evidence to Decision framework. Anonymous voting was conducted using predefined consensus criteria. RESULTS: Both adult and paediatric COM sets include forced expiratory volume in 1 s (FEV1) as z-scores, annual frequency of severe exacerbations and maintenance oral corticosteroid use. Additionally, the paediatric COM set includes the Paediatric Asthma Quality of Life Questionnaire and Asthma Control Test or Childhood Asthma Control Test, while the adult COM set includes the Severe Asthma Questionnaire and Asthma Control Questionnaire-6 (symptoms and rescue medication use reported separately). CONCLUSIONS: This patient-centred collaboration has produced two COM sets for paediatric and adult severe asthma. It is expected that they will inform the methodology of future clinical trials, enhance comparability of efficacy and effectiveness of biological therapies, and help assess their socioeconomic value. COMSA will inform definitions of non-response and response to biological therapy for severe asthma.

Contribution of UGT Enzymes to Human Drug Metabolism Stereoselectivity: A Case Study of Medetomidine, RO5263397, Propranolol, and Testosterone.

The enantiomeric forms of chiral compounds have identical physical properties but may vary greatly in their metabolism by individual enzymes. Enantioselectivity in UDP-glucuronosyl transferase (UGT) metabolism has been reported for a number of compounds and with different UGT isoforms involved. However, the impact of such individual enzyme results on overall clearance stereoselectivity is often not clear. The enantiomers of medetomidine, RO5263397, and propranolol and the epimers testosterone and epitestosterone exhibit more than a 10-fold difference in glucuronidation rates by individual UGT enzymes. In this study, we examined the translation of human UGT stereoselectivity to hepatic drug clearance considering the combination of multiple UGTs to overall glucuronidation, the contribution of other metabolic enzymes such as cytochrome P450s (P450s), and the potential for differences in protein binding and blood/plasma partitioning. For medetomidine and RO5263397, the high individual enzyme (UGT2B10) enantioselectivity translated into ∼3- to >10-fold differences in predicted human hepatic in vivo clearance. For propranolol, the UGT enantioselectivity was irrelevant in the context of high P450 metabolism. For testosterone, a complex picture emerged due to differential epimeric selectivity of various contributing enzymes and potential for extrahepatic metabolism. Quite different patterns of P450- and UGT-mediated metabolism were observed across species, as well as differences in stereoselectivity, indicating that extrapolation from human enzyme and tissue data are essential when predicting human clearance enantioselectivity. SIGNIFICANCE STATEMENT: Individual enzyme stereoselectivity illustrates the importance of three-dimensional drug-metabolizing enzyme-substrate interactions and is essential when considering the clearance of racemic drugs. However, translation from in vitro to in vivo can be challenging as contributions from multiple enzymes and enzyme classes must be combined with protein binding and blood/plasma partitioning data to estimate the net intrinsic clearance for each enantiomer. Preclinical species may be misleading as enzyme involvement and metabolism stereoselectivity can differ substantially.

PBPK Modelling for Drugs Cleared by Non-CYP Enzymes: State-of-the-Art and Future Perspectives.

Physiologically-based pharmacokinetic (PBPK) modeling has become the established method for predicting human pharmacokinetics (PK) and drug-drug interactions (DDI). The number of drugs cleared by non-CYP enzyme metabolism has increased steadily and to date, there is no consolidated overview of PBPK modeling for drugs cleared by non-CYP enzymes. This review aims to describe the state-of-the-art for PBPK modeling for drugs cleared via non-CYP enzymes, to identify successful strategies, to describe gaps and to provide suggestion to overcome them. To this end, we conducted a detailed literature search and found 58 articles published before the 1st of January 2023 containing 95 examples of clinical PBPK models for 62 non-CYP enzyme substrates. Reviewed articles covered the drug clearance by uridine 5'-diphospho-glucuronosyltransferases (UGTs), aldehyde oxidase (AO), flavin-containing monooxygenases (FMOs), sulfotransferases (SULTs) and carboxylesterases (CES), with UGT2B7, UGT1A9, CES1, FMO3 and AO being the enzymes most frequently involved. In vitro-in vivo extrapolation (IVIVE) of intrinsic clearance and the bottom-up PBPK modeling involving non-CYP enzymes remains challenging. We observed that the middle-out modeling approach was applied in 80% of the cases, with metabolism parameters optimized in 73% of the models. Our review could not identify a standardized approach used for model optimization based on clinical data, with manual optimization employed most frequently. Successful development of models for UGT2B7, UGT1A9, CES1, and FMO3 substrates provides a foundation for other drugs metabolized by these enzymes and guides the way forward in creating PBPK models for other enzymes in these families. Significance Statement Our review charts the rise of PBPK modeling for drugs cleared by non-CYP enzymes. Analyzing 58 articles and 62 non-CYP enzyme substrates, we found that UGTs, AO, FMOs, SULTs, and CES were the main enzyme families involved and that UGT2B7, UGT1A9, CES1, FMO3 and AO are the individual enzymes with the strongest PBPK modeling precedents. Approaches established for these enzymes can now be extended to additional substrates and to drugs metabolized by enzymes that are similarly well characterized.

The role of inflammation in anxiety and depression in the European U-BIOPRED asthma cohorts.

BACKGROUND: Growing evidence indicates high comorbid anxiety and depression in patients with asthma. However, the mechanisms underlying this comorbid condition remain unclear. The aim of this study was to investigate the role of inflammation in comorbid anxiety and depression in three asthma patient cohorts of the Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes (U-BIOPRED) project. METHODS: U-BIOPRED was conducted by a European Union consortium of 16 academic institutions in 11 European countries. A subset dataset from subjects with valid anxiety and depression measures and a large blood biomarker dataset were analysed, including 198 non-smoking patients with severe asthma (SAn), 65 smoking patients with severe asthma (SAs), 61 non-smoking patients with mild-to-moderate asthma (MMA), and 20 healthy non-smokers (HC). The Hospital Anxiety and Depression Scale was used to measure anxiety and depression and a series of inflammatory markers were analysed by the SomaScan v3 platform (SomaLogic, Boulder, Colo). ANOVA and the Kruskal-Wallis test were used for multiple-group comparisons as appropriate. RESULTS: There were significant group effects on anxiety and depression among the four cohort groups (p < 0.05). Anxiety and depression of SAn and SAs groups were significantly higher than that of MMA and HC groups (p < 0.05. There were significant differences in serum IL6, MCP1, CCL18, CCL17, IL8, and Eotaxin among the four groups (p < 0.05). Depression was significantly associated with IL6, MCP1, CCL18 level, and CCL17; whereas anxiety was associated with CCL17 only (p < 0.05). CONCLUSIONS: The current study suggests that severe asthma patients are associated with higher levels of anxiety and depression, and inflammatory responses may underlie this comorbid condition.

Aspergillus sensitisation detection using point-of-care lateral flow assay in moderate to severe asthma.

Allergic fungal airway diseases are associated with asthma exacerbations and poor control. However, the early identification of allergic Aspergillus airway diseases can be challenging, especially in resource-poor countries. We aimed to evaluate the clinical utility of the point-of-care Aspergillus IgG-IgM lateral flow assay in diagnosing Aspergillus airway diseases in patients with moderate-severe asthma. Patients with moderate-severe asthma, severe asthma with fungal sensitisation (SAFS) and allergic bronchopulmonary aspergillosis (ABPA) were recruited. Clinical information was extracted from clinical records. Blood samples were collected for serological tests. Serum samples were evaluated using Aspergillus immunochromatographic test (ICT). A total of 65 patients were recruited into the study, of whom 23.1% had clinical diagnosis of ABPA, 18.5% had SAFS and 58.5% had moderate-to-severe asthma who did not fit ABPA or SAFS criteria. The ICT test gave a sensitivity of 69 [95% confidence interval: 51-88]% and a specificity of 77 [60-88]% in predicting a positive Aspergillus IgG test. The sensitivity and specificity for a positive Aspergillus IgE were 77 [59-88]% and 86 [71-94]%, respectively. The majority (sensitivity: 87 [62-96]%) of patients with ABPA had positive ICT results, with a specificity of 70%. The negative predictive value was high (95 [82-99]%) with a low negative likelihood ratio (< 0.2), making it potentially useful in ruling out ABPA. The ICT assay may be valuable in ruling out ABPA in resource-limited countries where serological investigations are less feasible. The ICT assay may be particularly useful in ruling out ABPA and warrants further validation.

Allergic Aspergillus diseases can make patients with asthma more unwell, but this is difficult to diagnose, especially in developing countries where tests are not widely available. We have found that a cheap, easy-to-use and fast test may be useful in diagnosing allergic Aspergillus diseases.

Volatile metabolites differentiate air-liquid interface cultures after infection with Staphylococcus aureus.

Early detection of lung infection is critical to clinical diagnosis, treatment, and monitoring. Measuring volatile organic compounds (VOCs) in exhaled breath has shown promise as a rapid and accurate method of evaluating disease metabolism and phenotype. However, further investigations of the role and function of VOCs in bacterial-host-stress response is required and this can only be realised through representative in vitro models. In this study we sampled VOCs from the headspace of A549 cells at an air-liquid interface (ALI). We hypothesised VOC sampling from ALI cultures could be used to profile potential biomarkers of S. aureus lung infection. VOCs were collected using thin film microextraction (TFME) and were analysed by thermal desorption-gas chromatography-mass spectrometry. After optimising ALI cultures, we observed seven VOCs changed between A549 and media control samples. After infecting cells with S. aureus, supervised principal component-discriminant function analysis revealed 22 VOCs were found to be significantly changed in infected cells compared to uninfected cells (p < 0.05), five of which were also found in parallel axenic S. aureus cultures. We have demonstrated VOCs that could be used to identify S. aureus in ALI cultures, supporting further investigation of VOC analysis as a highly sensitive and specific test for S. aureus lung infection.