Protein Structure Inspired Drug Discovery.

Drug discovery starts with known function, either of a compound or a protein, in-turn prompting investigations to probe 3D structure of the compound-protein interface. As protein structure determines function, we hypothesized that unique 3D structural motifs represent primary information denoting unique function that can drive discovery of novel agents. Using a physics-based protein structure analysis platform developed by us, designed to conduct computationally intensive analysis at supercomputing speeds, we probed a high-resolution protein x-ray crystallographic library developed by us. We selected 3D structural motifs whose function was not otherwise established, that offered environments supporting binding of drug-like chemicals and were present on proteins that were not established therapeutic targets. For each of eight potential binding pockets on six different proteins we accessed a 60 million compound library and used our analysis platform to evaluate binding. Using eight-day colony formation assays acquired compounds were screened for efficacy against human breast, prostate, colon and lung cancer cells and toxicity against human bone marrow stem cells. Compounds selectively inhibiting cancer growth segregated to two pockets on separate proteins. The compound, Dxr2-017, exhibited selective activity against human melanoma cells in the NCI-60 cell line screen, had an IC50 of 19 nM against human melanoma M14 cells in our eight-day assay, while over 2100-fold higher concentrations inhibited stem cells by less than 30%. We show that Dxr2-017 induces anoikis, a unique form of programmed cell death in need of targeted therapeutics. The predicted target protein for Dxr2-017 is expressed in bacteria, not in humans. This supports our strategy of focusing on unique 3D structural motifs. It is known that functionally important 3D structures are evolutionarily conserved. Here we demonstrate proof-of-concept that protein structure represents high value primary data to support discovery of novel therapeutics. This approach is widely applicable.

Association of Ground Glass Opacities with Systemic Inflammation and Progression of Emphysema.

RATIONAL: Ground glass opacities (GGO) in the absence of interstitial lung disease are understudied. OBJECTIVE: To assess the association of GGO with white blood cells (WBCs) and progression of quantified chest CT emphysema. METHODS: We analyzed data of participants in the Subpopulations and Intermediate Outcome Measures In COPD Study (SPIROMICS). Chest radiologists and pulmonologists labeled regions of the lung as GGO and adaptive multiple feature method (AMFM) trained the computer to assign those labels to image voxels and quantify the volume of the lung with GGO (%GGOAMFM). We used multivariable linear regression, zero-inflated negative binomial, and proportional hazards regression models to assess the association of %GGOAMFM with WBC, changes in %emphysema, and clinical outcomes. MEASUREMENTS AND MAIN RESULTS: Among 2,714 participants, 1,680 had COPD and 1,034 had normal spirometry. Among COPD participants, based on the multivariable analysis, current smoking and chronic productive cough was associated with higher %GGOAMFM. Higher %GGOAMFM was cross-sectionally associated with higher WBCs and neutrophils levels. Higher %GGOAMFM per interquartile range at visit 1 (baseline) was associated with an increase in emphysema at one-year follow visit by 11.7% (Relative increase; 95%CI 7.5-16.1%;P<0.001). We found no association between %GGOAMFM and one-year FEV1 decline but %GGOAMFM was associated with exacerbations and all-cause mortality during a median follow-up time of 1,544 days (Interquartile Interval=1,118-2,059). Among normal spirometry participants, we found similar results except that %GGOAMFM was associated with progression to COPD at one-year follow-up. CONCLUSIONS: Our findings suggest that GGOAMFM is associated with increased systemic inflammation and emphysema progression.

Drug release profile of a novel exenatide long-term drug delivery system (OKV-119) administered to cats.

Beneficial weight-loss properties of glucagon-like peptide-1 receptor agonists (GLP-1RA) in obese people, with corresponding improvements in cardiometabolic risk factors, are well established. OKV-119 is an investigational drug delivery system that is being developed for the long-term delivery of the GLP-1RA exenatide to feline patients. The purpose of this study was to evaluate the drug release characteristics of subcutaneous OKV-119 implants configured to release exenatide for 84 days. Following a 7-day acclimation period, five purpose-bred cats were implanted with OKV-119 protypes and observed for a 112-day study period. Food intake, weekly plasma exenatide concentrations and body weight were measured. Exenatide plasma concentrations were detected at the first measured timepoint (Day 7) and maintained above baseline for over 84 Days. Over the first 28 days, reduced caloric intake and a reduction in body weight were observed in four of five cats. In these cats, a body weight reduction of at least 5% was maintained throughout the 112-day study period. This study demonstrates that a single OKV-119 implant can deliver the GLP-1RA exenatide for a months long duration. Results suggest that exposure to exenatide plasma concentrations ranging from 1.5 ng/ml to 4 ng/ml are sufficient for inducing weight loss in cats.

From TgO/GABA-AT, GABA, and T-263 Mutant to Conception of Toxoplasma.

Toxoplasma gondii causes morbidity, mortality, and disseminates widely via cat sexual stages. Here, we find T. gondii ornithine aminotransferase (OAT) is conserved across phyla. We solve TgO/GABA-AT structures with bound inactivators at 1.55 Å and identify an inactivator selective for TgO/GABA-AT over human OAT and GABA-AT. However, abrogating TgO/GABA-AT genetically does not diminish replication, virulence, cyst-formation, or eliminate cat's oocyst shedding. Increased sporozoite/merozoite TgO/GABA-AT expression led to our study of a mutagenized clone with oocyst formation blocked, arresting after forming male and female gametes, with "Rosetta stone"-like mutations in genes expressed in merozoites. Mutations are similar to those in organisms from plants to mammals, causing defects in conception and zygote formation, affecting merozoite capacitation, pH/ionicity/sodium-GABA concentrations, drawing attention to cyclic AMP/PKA, and genes enhancing energy or substrate formation in TgO/GABA-AT-related-pathways. These candidates potentially influence merozoite's capacity to make gametes that fuse to become zygotes, thereby contaminating environments and causing disease.

Null effects of temporal prediction on recognition memory but evidence for differential neural activity at encoding. A registered report.

Previous research has demonstrated that rhythmic presentation of stimuli during encoding boosts subsequent recognition and is associated with distinct neural activity compared with when stimuli are presented in an arrhythmic manner. However, it is unclear whether the effect is driven by automatic entrainment to rhythm or non-rhythmic temporal prediction. This registered report presents an Electroencephalographic (EEG) study aimed at establishing the cognitive and neural mechanisms of the effect of temporal prediction on recognition. In a blocked design, stimulus onset during encoding was systematically manipulated in four conditions prior to recognition testing: rhythmic fixed (RF), rhythmic variable (RV), arrhythmic fixed (AF), and arrhythmic variable (AV). By orthogonally varying rhythm and temporal position we were able to assess their independent contributions to recognition enhancement. Our behavioural results did not replicate previous findings that show a difference in recognition memory based on temporal predictability at encoding. However, event-related potential (ERP) component analysis did show an early (N1) interaction effect of temporal position and rhythm, and later (N2 and Dm) effects driven by temporal position only. Taken together, we observed effects of temporal prediction at encoding, but these differences did not translate to later effects of memory, suggesting that effects of temporal prediction on recognition are less robust than previously thought.

A high-throughput structural system biology approach to increase structure representation of proteins from Clostridioides difficile.

Clostridioides difficile causes life-threatening gastrointestinal infections. It is a high-risk pathogen due to a lack of effective treatments, antimicrobial resistance, and a poorly conserved genomic core. Herein, we report 30 X-ray structures from a structure genomics pipeline spanning 13 years, representing 10.2% of the X-ray structures for this important pathogen.

Longitudinal Follow-Up of Participants With Tobacco Exposure and Preserved Spirometry.

Importance: People who smoked cigarettes may experience respiratory symptoms without spirometric airflow obstruction. These individuals are typically excluded from chronic obstructive pulmonary disease (COPD) trials and lack evidence-based therapies. Objective: To define the natural history of persons with tobacco exposure and preserved spirometry (TEPS) and symptoms (symptomatic TEPS). Design, Setting, and Participants: SPIROMICS II was an extension of SPIROMICS I, a multicenter study of persons aged 40 to 80 years who smoked cigarettes (>20 pack-years) with or without COPD and controls without tobacco exposure or airflow obstruction. Participants were enrolled in SPIROMICS I and II from November 10, 2010, through July 31, 2015, and followed up through July 31, 2021. Exposures: Participants in SPIROMICS I underwent spirometry, 6-minute walk distance testing, assessment of respiratory symptoms, and computed tomography of the chest at yearly visits for 3 to 4 years. Participants in SPIROMICS II had 1 additional in-person visit 5 to 7 years after enrollment in SPIROMICS I. Respiratory symptoms were assessed with the COPD Assessment Test (range, 0 to 40; higher scores indicate more severe symptoms). Participants with symptomatic TEPS had normal spirometry (postbronchodilator ratio of forced expiratory volume in the first second [FEV1] to forced vital capacity >0.70) and COPD Assessment Test scores of 10 or greater. Participants with asymptomatic TEPS had normal spirometry and COPD Assessment Test scores of less than 10. Patient-reported respiratory symptoms and exacerbations were assessed every 4 months via phone calls. Main Outcomes and Measures: The primary outcome was assessment for accelerated decline in lung function (FEV1) in participants with symptomatic TEPS vs asymptomatic TEPS. Secondary outcomes included development of COPD defined by spirometry, respiratory symptoms, rates of respiratory exacerbations, and progression of computed tomographic-defined airway wall thickening or emphysema. Results: Of 1397 study participants, 226 had symptomatic TEPS (mean age, 60.1 [SD, 9.8] years; 134 were women [59%]) and 269 had asymptomatic TEPS (mean age, 63.1 [SD, 9.1] years; 134 were women [50%]). At a median follow-up of 5.76 years, the decline in FEV1 was -31.3 mL/y for participants with symptomatic TEPS vs -38.8 mL/y for those with asymptomatic TEPS (between-group difference, -7.5 mL/y [95% CI, -16.6 to 1.6 mL/y]). The cumulative incidence of COPD was 33.0% among participants with symptomatic TEPS vs 31.6% among those with asymptomatic TEPS (hazard ratio, 1.05 [95% CI, 0.76 to 1.46]). Participants with symptomatic TEPS had significantly more respiratory exacerbations than those with asymptomatic TEPS (0.23 vs 0.08 exacerbations per person-year, respectively; rate ratio, 2.38 [95% CI, 1.71 to 3.31], P < .001). Conclusions and Relevance: Participants with symptomatic TEPS did not have accelerated rates of decline in FEV1 or increased incidence of COPD vs those with asymptomatic TEPS, but participants with symptomatic TEPS did experience significantly more respiratory exacerbations over a median follow-up of 5.8 years.

A blood and bronchoalveolar lavage protein signature of rapid FEV1 decline in smoking-associated COPD.

Accelerated progression of chronic obstructive pulmonary disease (COPD) is associated with increased risks of hospitalization and death. Prognostic insights into mechanisms and markers of progression could facilitate development of disease-modifying therapies. Although individual biomarkers exhibit some predictive value, performance is modest and their univariate nature limits network-level insights. To overcome these limitations and gain insights into early pathways associated with rapid progression, we measured 1305 peripheral blood and 48 bronchoalveolar lavage proteins in individuals with COPD [n = 45, mean initial forced expiratory volume in one second (FEV1) 75.6 ± 17.4% predicted]. We applied a data-driven analysis pipeline, which enabled identification of protein signatures that predicted individuals at-risk for accelerated lung function decline (FEV1 decline ≥ 70 mL/year) ~ 6 years later, with high accuracy. Progression signatures suggested that early dysregulation in elements of the complement cascade is associated with accelerated decline. Our results propose potential biomarkers and early aberrant signaling mechanisms driving rapid progression in COPD.

Spatial attention is not affected by alpha or beta transcranial alternating current stimulation: A registered report.

Using Electroencephalography (EEG) an event-related change in alpha activity has been observed over primary sensory cortices during the allocation of spatial attention. This is most prominent during top-down, or endogenous, attention, and nearly absent in bottom-up, or exogenous orienting. These changes are highly lateralised, such that an increase in alpha power is seen ipsilateral to the attended region of space and a decrease is seen contralaterally. Whether these changes in alpha oscillatory activity are causally related to attentional resources, or to perceptual processes, or are simply epiphenomenal, is unknown. If alpha oscillations are indicative of a causal mechanism whereby attention is allocated to a region of space, it remains an open question as to whether this is driven by ipsilateral increases or contralateral decreases in alpha power. This preregistered report set out to test these questions. To do so, we used transcranial Alternating Current Stimulation (tACS) to modulate alpha activity in the somatosensory cortex whilst measuring performance on established tactile attention paradigms. All participants completed an endogenous and exogenous tactile attention task in three stimulation conditions; alpha, sham and beta. Sham and beta stimulation operated as controls so that any observed effects could be attributed to alpha stimulation specifically. We replicated previous behavioural findings in all stimulation conditions showing a facilitation of cued trials in the endogenous task, and inhibition of return in the exogenous task. However, these were not affected by stimulation manipulations. Using Bayes-factor analysis we show strong support for the null hypotheses - that the manipulation of Alpha by tACS does not cause changes in tactile spatial attention. This well-powered study, conducted over three separate days, is an important contribution to the current debate regarding the efficiency of brain stimulation.

A Structural Systems Biology Approach to High-Risk CG23 Klebsiella pneumoniae.

Klebsiella pneumoniae is a leading cause of antibiotic-resistant-associated deaths in the world. Here, we report the deposition of 14 structures of enzymes from both the core and accessory genomes of sequence type 23 (ST23) K1 hypervirulent K. pneumoniae.

The contribution of endocytosis to sensitization of nociceptors and synaptic transmission in nociceptive circuits.

ABSTRACT: Chronic pain involves sensitization of nociceptors and synaptic transmission of painful signals in nociceptive circuits in the dorsal horn of the spinal cord. We investigated the contribution of clathrin-dependent endocytosis to sensitization of nociceptors by G protein-coupled receptors (GPCRs) and to synaptic transmission in spinal nociceptive circuits. We determined whether therapeutic targeting of endocytosis could ameliorate pain. mRNA encoding dynamin (Dnm) 1 to 3 and adaptor-associated protein kinase 1 (AAK1), which mediate clathrin-dependent endocytosis, were localized to primary sensory neurons of dorsal root ganglia of mouse and human and to spinal neurons in the dorsal horn of the mouse spinal cord by RNAScope. When injected intrathecally to mice, Dnm and AAK1 siRNA or shRNA knocked down Dnm and AAK1 mRNA in dorsal root ganglia neurons, reversed mechanical and thermal allodynia and hyperalgesia, and normalized nonevoked behavior in preclinical models of inflammatory and neuropathic pain. Intrathecally administered inhibitors of clathrin, Dnm, and AAK1 also reversed allodynia and hyperalgesia. Disruption of clathrin, Dnm, and AAK1 did not affect normal motor functions of behaviors. Patch clamp recordings of dorsal horn neurons revealed that Dnm1 and AAK1 disruption inhibited synaptic transmission between primary sensory neurons and neurons in lamina I/II of the spinal cord dorsal horn by suppressing release of synaptic vesicles from presynaptic primary afferent neurons. Patch clamp recordings from dorsal root ganglion nociceptors indicated that Dnm siRNA prevented sustained GPCR-mediated sensitization of nociceptors. By disrupting synaptic transmission in the spinal cord and blunting sensitization of nociceptors, endocytosis inhibitors offer a therapeutic approach for pain treatment.

Bronchodilator Responsiveness in Tobacco-Exposed People With or Without COPD.

BACKGROUND: Bronchodilator responsiveness (BDR) in obstructive lung disease varies over time and may be associated with distinct clinical features. RESEARCH QUESTION: Is consistent BDR over time (always present) differentially associated with obstructive lung disease features relative to inconsistent (sometimes present) or never (never present) BDR in tobacco-exposed people with or without COPD? STUDY DESIGN AND METHODS: We retrospectively analyzed data from 2,269 tobacco-exposed participants in the Subpopulations and Intermediate Outcome Measures in COPD Study with or without COPD. We used various BDR definitions: change of ≥ 200 mL and ≥ 12% in FEV1 (FEV1-BDR), change in FVC (FVC-BDR), and change in in FEV1, FVC or both (ATS-BDR). Using generalized linear models adjusted for demographics, smoking history, FEV1 % predicted after bronchodilator administration, and number of visits that the participant completed, we assessed the association of BDR group: (1) consistent BDR, (2) inconsistent BDR, and (3) never BDR with asthma, CT scan features, blood eosinophil levels, and FEV1 decline in participants without COPD (Global Initiative for Chronic Obstructive Lung Disease [GOLD] stage 0) and the entire cohort (participants with or without COPD). RESULTS: Both consistent and inconsistent ATS-BDR were associated with asthma history and greater small airways disease (%parametric response mapping functional small airways disease) relative to never ATS-BDR in participants with GOLD stage 0 disease and the entire cohort. We observed similar findings using FEV1-BDR and FVC-BDR definitions. Eosinophils did not vary consistently among BDR groups. Consistent BDR was associated with FEV1 decline over time relative to never BDR in the entire cohort. In participants with GOLD stage 0 disease, both the inconsistent ATS-BDR group (OR, 3.20; 95% CI, 2.21-4.66; P < .001) and consistent ATS-BDR group (OR, 9.48; 95% CI, 3.77-29.12; P < .001) were associated with progression to COPD relative to the never ATS-BDR group. INTERPRETATION: Demonstration of BDR, even once, describes an obstructive lung disease phenotype with a history of asthma and greater small airways disease. Consistent demonstration of BDR indicated a high risk of lung function decline over time in the entire cohort and was associated with higher risk of progression to COPD in patients with GOLD stage 0 disease.

Clinical Implications of Low Absolute Blood Eosinophil Count in the SPIROMICS COPD Cohort.

BACKGROUND: The Global Initiative for Chronic Obstructive Lung Disease (GOLD) considers blood eosinophil counts < 100 cells/µL (BEC≤100) in people with COPD to predict poor inhaled corticosteroid (ICS) responsiveness. However, the BEC≤100 phenotype is inadequately characterized, especially in advanced COPD. RESEARCH QUESTION: Are there differences between GOLD group D patients with high BEC and those with low BEC regarding baseline characteristics and longitudinal outcomes? STUDY DESIGN AND METHODS: We used multivariable mixed models and logistic regression to contrast clinical characteristics and outcomes of BEC≤100 vs BEC > 100 (BEC100+) in all subjects with COPD (n = 1,414) and GOLD group D subjects (n = 185) not receiving ICS. RESULTS: We identified n = 485 with BEC≤100 (n = 61 GOLD group D) and n = 929 people with BEC100+ (n = 124 GOLD group D). BEC≤100 status was stable at 6 weeks and approximately 52 weeks (intraclass correlations of 0.78 and 0.71, respectively). Compared with BEC100+, BEC≤100 comprised more women, with greater current smoking, and less frequent childhood asthma. Among all analyzed participants, the two BEC-defined subsets showed similar rates of lung function decline (mean slope, BEC≤100 vs BEC100+, -50 vs -39 mL/y; P = .140), exacerbations (0.40 vs 0.36/y; P = .098), subsequent ICS initiation (2.5% vs 4.4%; P = .071), and mortality (7.8% vs 8.4%; P = .715). However, in GOLD group D, people with BEC≤100 showed higher exacerbation rates within 365 days of enrollment (0.62 vs 0.33/y; P = .002) and total follow-up (1.16 vs 0.83/y; P = .014). They also had greater lung function decline (mean slope of -68 mL/y vs -23 mL/y; P = .036) and had greater emphysema at baseline (voxels < 950 Hounsfield units at total lung capacity of 7.46% vs 4.61%; P = .029). INTERPRETATION: In non-ICS-treated GOLD group D COPD, people with BEC≤100 had more baseline emphysema, prospective exacerbations, and lung function decline. Our analysis has identified a particularly vulnerable subpopulation of people with COPD, suggesting the need for studies focused specifically on their therapeutic treatment. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov; No.: NCT01969344; URL: www. CLINICALTRIALS: gov.

Structure of the Monkeypox virus profilin-like protein A42R reveals potential functional differences from cellular profilins.

The infectious disease human monkeypox is spreading rapidly in 2022, causing a global health crisis. The genomics of Monkeypox virus (MPXV) have been extensively analyzed and reported, although little is known about the virus-encoded proteome. In particular, there are no reported experimental MPXV protein structures other than computational models. Here, a 1.52 Šresolution X-ray structure of the MPXV protein A42R, the first MPXV-encoded protein with a known structure, is reported. A42R shows structural similarity to profilins, which are cellular proteins that are known to function in the regulation of actin cytoskeletal assembly. However, structural comparison of A42R with known members of the profilin family reveals critical differences that support prior biochemical findings that A42R only weakly binds actin and does not bind poly(L-proline). In addition, the analysis suggests that A42R may make distinct interactions with phosphatidylinositol lipids. Overall, the data suggest that the role of A42R in the replication of orthopoxviruses may not be readily determined by comparison to cellular profilins. Furthermore, these findings support the need for increased efforts to determine high-resolution structures of other MPXV proteins to inform physiological studies of the poxvirus infection cycle and to reveal potential new strategies to combat human monkeypox should this emerging infectious disease with pandemic potential become more common in the future.

To bind, or not to bind, that is the question .

The unexpected findings that are described by Czub et al. [IUCrJ (2022), 9, 551-561] provide a very interesting study demonstrating how small differences in structure can result in significant changes in the relative affinities of the seemingly promiscuous binding sites that are seen with serum albumins.

Sensitive and Robust LC-MS/MS Analysis of Salivary Cortisol in Negative Mode.

Cortisol is one of the most important glucocorticoids involved in the regulation of human metabolism and physiological stress. Monitoring of levels of cortisol is of immense clinical benefit. In particular, salivary cortisol levels have been shown to correlate well with diurnal changes in cortisol levels in serum and have been used widely for monitoring of cortisol levels for diagnosis and prognosis purposes. We present a sensitive, fast, and robust quantitative liquid chromatography and tandem mass spectrometry (LC-MS/MS) assay for salivary cortisol in negative mode. This assay employs protein precipitation followed by reversed-phase liquid chromatographic separation, negative-mode electrospray ionization (ESI), and MS/MS detection. This assay has a total run time of 5.8 minutes and a limit of quantification of 0.5 ng/mL with a linear range up to 100 ng/mL. No carryover was observed at 10 µg/mL. This assay also incorporates the routine monitoring of prednisolone, a potential interferent to salivary cortisol.

Listeria monocytogenes two component system PieRS regulates secretion chaperones PrsA1 and PrsA2 and enhances bacterial translocation across the intestine.

Listeria monocytogenes (Lm) is a widespread environmental Gram-positive bacterium that can transition into a pathogen following ingestion by a susceptible host. To cross host barriers and establish infection, Lm is dependent upon the regulated secretion and activity of many proteins including PrsA2, a peptidyl-prolyl cis-trans isomerase with foldase activity. PrsA2 contributes to the stability and activity of a number of secreted virulence factors that are required for Lm invasion, replication, and cell-to-cell spread within the infected host. In contrast, a second related secretion chaperone, PrsA1, has thus far no identified contributions to Lm pathogenesis. Here we describe the characterization of a two-component signal transduction system PieRS that regulates the expression of a regulon that includes the secretion chaperones PrsA1 and PrsA2. PieRS regulated gene products are required for bacterial resistance to ethanol exposure and are important for bacterial survival during transit through the gastrointestinal tract. PrsA1 was also found to make a unique contribution to Lm survival in the GI tract, revealing for the first time a non-overlapping requirement for both secretion chaperones PrsA1 and PrsA2 during the process of intra-gastric infection.

Significance of FEV3/FEV6 in Recognition of Early Airway Disease in Smokers at Risk of Development of COPD: Analysis of the SPIROMICS Cohort.

BACKGROUND: Small airways are known to be affected early in the course of COPD; however, traditional spirometric indices may not accurately identify small airways disease. RESEARCH QUESTION: Can forced expiratory volume in 3 s/forced expiratory volume in 6 s (FEV3/FEV6) identify early airflow abnormalities and predict future clinically important respiratory-related outcomes, including development of COPD? STUDY DESIGN AND METHODS: The study included 832 current and former smokers with post-bronchodilator FEV1/FVC ≥ 0.7 from the Subpopulations and Intermediate Outcome Measures in COPD Study (SPIROMICS) cohort. Participants were classified as having a reduced pre-bronchodilator FEV3/FEV6 based on lower limit of normal (LLN) values. Repeatability analysis was performed for FEV3 and FEV6. Regression modeling was used to evaluate the relationship between baseline FEV3/FEV6 and outcome measures, including functional small airways disease, on thoracic imaging and respiratory exacerbations. Interval-censored analysis was used to assess progression to COPD. RESULTS: FEV3/FEV6 less than the LLN at baseline, defined as reduced compared with FEV3/FEV6 at or above the LLN, was associated with lower FEV1, poorer health status (St. George's Respiratory Questionnaire score), more emphysema, and more functional small airways disease on quantitative imaging. FEV3 and FEV6 showed excellent agreement between repeat measurements. A reduced FEV3/FEV6 was associated with increased odds of a severe respiratory exacerbation within the first year of follow-up and decreased time to first exacerbation. A low FEV3/FEV6 was also associated with development of COPD according to spirometry results (post-bronchodilator FEV1/FVC < 0.7) during study follow-up. INTERPRETATION: FEV3/FEV6 is a routinely available and repeatable spirometric index that can be useful in the evaluation of early airflow obstruction in current and former smokers without COPD. A reduced FEV3/FEV6 can identify those at risk for future development of COPD and respiratory exacerbations. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov; No.: NCT01969344; URL: www. CLINICALTRIALS: gov: ClinicalTrials.gov.

Identification of Sputum Biomarkers Predictive of Pulmonary Exacerbations in COPD.

BACKGROUND: Improved understanding of the pathways associated with airway pathophysiologic features in COPD will identify new predictive biomarkers and novel therapeutic targets. RESEARCH QUESTION: Which physiologic pathways are altered in the airways of patients with COPD and will predict exacerbations? STUDY DESIGN AND METHODS: We applied a mass spectrometric panel of metabolomic biomarkers related to mucus hydration and inflammation to sputa from the multicenter Subpopulations and Intermediate Outcome Measures in COPD Study. Biomarkers elevated in sputa from patients with COPD were evaluated for relationships to measures of COPD disease severity and their ability to predict future exacerbations. RESULTS: Sputum supernatants from 980 patients were analyzed: 77 healthy nonsmokers, 341 smokers with preserved spirometry, and 562 patients with COPD (178 with Global Initiative on Chronic Obstructive Lung Disease [GOLD] stage 1 disease, 303 with GOLD stage 2 disease, and 81 with GOLD stage 3 disease) were analyzed. Biomarkers from multiple pathways were elevated in COPD and correlated with sputum neutrophil counts. Among the most significant analytes (false discovery rate, 0.1) were sialic acid, hypoxanthine, xanthine, methylthioadenosine, adenine, and glutathione. Sialic acid and hypoxanthine were associated strongly with measures of disease severity, and elevation of these biomarkers was associated with shorter time to exacerbation and improved prediction models of future exacerbations. INTERPRETATION: Biomarker evaluation implicated pathways involved in mucus hydration, adenosine metabolism, methionine salvage, and oxidative stress in COPD airway pathophysiologic characteristics. Therapies that target these pathways may be of benefit in COPD, and a simple model adding sputum-soluble phase biomarkers improves prediction of pulmonary exacerbations. TRIAL REGISTRY: ClinicalTrials.gov; No.: NCT01969344; URL: www. CLINICALTRIALS: gov.