R. M. Mroz

Efficacy and Safety of Aclidinium Bromide Compared with Placebo and Tiotropium in Patients with Moderate-to-Severe Chronic Obstructive Pulmonary Disease: Results from a 6-week, Randomized, Controlled Phase Iiib Study

Abstract Background: This randomized, double-blind, Phase IIIb study evaluated the 24-hour bronchodilatory efficacy of aclidinium bromide versus placebo and tiotropium in patients with moderate-to-severe chronic obstructive pulmonary disease (COPD). Methods: Patients received aclidinium 400 μg twice daily (morning and evening), tiotropium 18 μg once daily (morning), or placebo for 6 weeks. The primary endpoint was change from baseline in forced expiratory volume in 1 second area under the curve for the 24-hour period post-morning dose (FEV1 AUC0–24) at week 6. Secondary and additional endpoints included FEV1 AUC12–24, COPD symptoms (EXAcerbations of chronic pulmonary disease Tool-Respiratory Symptoms [E-RS] total score and additional symptoms questionnaire), and safety. Results: Overall, 414 patients were randomized and treated (FEV1 1.63 L [55.8% predicted]). Compared with placebo, FEV1 AUC0–24 and FEV1 AUC12–24 were significantly increased from baseline with aclidinium (∆ = 150 mL and 160 mL, respectively; p < 0.0001) and tiotropium (∆ = 140 mL and 123 mL, respectively; p < 0.0001) at week 6. Significant improvements in E-RS total scores over 6 weeks were numerically greater with aclidinium (p < 0.0001) than tiotropium (p < 0.05) versus placebo. Only aclidinium significantly reduced the severity of early-morning cough, wheeze, shortness of breath, and phlegm, and of nighttime symptoms versus placebo (p < 0.05). Adverse-event (AE) incidence (28%) was similar between treatments. Few anticholinergic AEs (<1.5%) or serious AEs (<3%) occurred in any group. Conclusions: Aclidinium provided significant 24-hour bronchodilation versus placebo from day 1 with comparable efficacy to tiotropium after 6 weeks. Improvements in COPD symptoms were consistently numerically greater with aclidinium versus tiotropium. Aclidinium was generally well tolerated.

Increased levels of Treg cells in bronchoalveolar lavage fluid and induced sputum of patients with active pulmonary sarcoidosis

ObjectiveIt has recently been described that circulatory and BAL regulatory T-cells (Tregs), defined as CD4+CD25highCD127low are increased in patients with active sarcoidosis compared with other interstitial lung diseases.Materials and methodsWe studied prospectively 17 patients (10 women, 7 men) of median age 39 years (range 27-65) with active granulomatous lung diseases (GLD) (10 patients with sarcoidosis (BBS), and 7 with hypersensitivity pneumonitis (HP), and 9 healthy controls. Bronchoalveolar lavage fluid (BAL) and induced sputum Treg counts, CD4+, CD8+, CD25+ cells were quantified by flow cytometry. Disease activity was measured by ACE serum level. Pulmonary function tests were performed using an Elite DL Medgraphics body box.ResultsWe found Treg cells count significantly elevated in induced sputum from active GLD (38.3% vs. 7.1% and 5.3% in BBS, HP, and control, respectively). A significantly higher percentage of Treg cells characterized BAL cells from HP patients (2.27%; 9.5%; 2.1%, in BBS, HP and control, respectively). There was a strong correlation with ACE serum level and Treg cell count in BAL fluid of BBS patients, with no such correlation within HP patient group, nor Treg cell count and pulmonary function tests.ConclusionsOur data suggest a potential role of CD4+CD25 high CD127 low induced sputum and BAL lymphocytes from patients with active granulomatous lung diseases and hypersensitivity pneumonitis. An increased number of Treg cells in active GLD may be involved in immune regulation in active granulomatous lung diseases. The results indicate that analysis of these cells could be useful as markers of disease activity in granulomatous lung diseases.

Tiotropium Increases PPARγ and Decreases CREB in Cells Isolated from Induced Sputum of COPD Patients

Chronic obstructive pulmonary disease (COPD) is characterized by progressive airflow limitation and chronic inflammation of airways and lung parenchyma. Our aim was to assess two important elements of intracellular signaling involved in regulation of inflammation in COPD in patients subjected to long-acting beta2-agonist or long-acting beta2-agonist plus long-acting antimuscarinic: peroxisome proliferator-activated receptor gamma (PPARγ) protein, which has antiinflammatory and immunomodulatory properties and cAMP response element binding protein (CREB) and activated (CREB-P) protein which has histone acetyltransferase activity and increases histone acetylation and transcriptional activation of chromatin. Twenty one stable COPD patients (18 males and 3 females, mean age 65 years) receiving 12 μg B.I.D formoterol were assayed before and after 3 month add-on therapy, consisting of 18 μg Q.D. tiotropium. In all patients, sputum induction, spirometry, lung volumes, and DLCO were performed before and after therapy. Sputum cells were isolated and processed to isolate cytosolic and nuclear fractions. PPARγ, CREB, or CREB-P proteins were quantified in subcellular fractions using Western blot. Tiotropium add-on therapy improved respiratory parameters: FEV1 and lung volumes. After therapy mean expression of PPARγ in cell nuclei was significantly increased by about 180%, while CREB and phosphorylated CREB levels in cytosol and nuclei were decreased by about 30%. Our data show that the mechanism whereby tiotropium reduces exacerbations may be associated not only with persistent increase in airway functions and reduced hyperinflation mediated by muscarinic receptors, but also with possible anti-inflammatory effects of the drug, involving increased PPARγ and decreased CREB signaling.

Ttiotropium increases cytosolic muscarinic M3 receptors and acetylated H3 histone proteins in induced sputum cells of COPD patients

ObjectiveChronic obstructive pulmonary disease (COPD) is characterized by irreversible progressive airflow limitation related to tobacco smoking. This limitation is caused by chronic inflammation of the airways and lung parenchyma and is associated with increased activity of parasympathetic system. The most effective bronchodilators in COPD are muscarinic receptor antagonists (MRA), which reverse, at least in part, compromised respiratory function. MRA also contribute to control inflammatory processes via interactions with inflammatory signaling molecules. The use of the long-acting cholinolytic bronchodilatator - tiotropium, with high affinity to M3 receptors, is suggested as a first line maintenance treatment in COPD patients.Materials and methodsIn this study we assessed M3 receptor protein expression in induced sputum of 27 stable COPD patients before and after therapy consisting of 18 μg once daily tiotropium for 12 weeks. Lung function tests including spirometry, lung volumes, and DLCO were performed before and after therapy in all COPD patients. The patients were subjected to the sputum induction procedures before and after therapy. Sputum cells were isolated, sample-specific cell profiles were characterized, and the cells were processed to isolate pure cytosolic fractions. Cytosolic M3 protein and HDAC2 levels and nuclear acetylated histone H3 (AcH3) expression was quantified using specific antibodies against human proteins and Western blot with enhanced luminescence detection.ResultsTherapy significantly increased the mean forced expiratory volume in one second (FEV1) and forced vital capacity (FVC) volume (P < 0.05). The mean expression of M3 protein was higher by 37% after therapy (P < 0.05), HDAC2 expression was not altered, while AcH3 level was increased by about 90% (P < 0.01), compared with the corresponding data before therapy. HDAC2 expression before therapy was positively correlated with AcH3 expression (r = 0.74), while after therapy no correlation was detected. FEV1, FCV, and cytosolic M3 protein expression did not correlate with other biochemical parameters tested.ConclusionsTwelve weeks of tiotropium therapy in COPD patients improves clinical indices of lung function and involves alterations in sputum cell chromatin acetylation and also increased cholinergic M3 receptor internalization.

Nuclear HSP90 and HSP70 in COPD patients treated with formoterol or formoterol and corticosteroids

ObjectiveHeat shock proteins assist cellular protein folding and are required for the normal activity of steroid receptors. In this study we assessed nuclear HSP90 and HSP70 proteins and mRNA levels in cells isolated from induced sputum of chronic obstructive pulmonary disease patients treated for 4 weeks with formoterol (F) or formoterol+budesonide (F/ICS).MethodsNuclear heat shock protein levels were assessed by Western blot and specific mRNAs were quantified in cell lysates using qRT-PCR.ResultsBoth HSP90 and HSP70 protein levels were higher in the F/ICS-treated patients in comparison with the F-treated group (by 31%, P < 0.05 and 28%, P < 0.05, respectively), while specific mRNAs were lowered. HSP86/HSP89 and D6S182/HSP90-BETA were repressed by about 40% (P < 0.05)whileHSP70-1/HSP70-1A, HSP70-1B/HSP70-2, and HSP70-HSC54/HSC70 were repressed by 47% (P < 0.01), 57% (P < 0.01) and 65% (P < 0.01), respectively.ConclusionsIt is possible that increased nuclear heat shock proteins may play a role in the attenuation of the response to glucocorticoids in COPD patients.

Altered histone deacetylase activity and iNOS expression in cells isolated from induced sputum of COPD patients treated with tiotropium.

Chronic obstructive pulmonary disease (COPD) is the only major disease with increasing death rate. In COPD, progressive reduction in quality of life is closely related to the increasing limitation of airflow due to chronic bronchitis, cell hyperplasia, fibrosis, and irreversible lung damage. Signaling pathways involved in inflammatory processes in COPD and inflammatory response to therapy are unknown. Our aim was to isolate cells from induced sputum of COPD patients treated with formoterol or formoterol + tiotropium and assess enzymatic activity of histone deacetylases (HDACs) acetylated histone 4 (AcH4) and expression of inducible nitric oxide synthase (iNOS). HDACs are important in signal transduction and inflammation. iNOS is generating nitric oxide (NO) relevant to blood pressure regulation, inflammation and infections. Thirty stable COPD patients (21 males and 9 females, mean age 67 years) receiving 12 μg b.i.d. formoterol were assayed before and after 3 months add-on therapy consisting of 18 μg q.i.d. tiotropium. In all patients, spirometry, lung volumes, and DLCO were performed before and after tiotropium therapy and all patients were subjected to sputum induction. Sputum cells were isolated and processed to obtain cytosolic and nuclear fractions. HDAC activity was measured in nuclear fraction using colorimetric assay. Expression AcH4 and iNOS was quantified using Western blot. In patients receiving both drugs, FEV1 and lung volumes significantly improved compared with formoterol-only treated patients. Mean HDAC activity was slightly decreased (P < 0.05), while AcH4 levels and iNOS expression were significantly elevated in tiotropium-treated patients (increase by about 65 %; P < 0.01 and 77 %; P < 0.01 respectively). Our data show that beneficial effects of tiotropium in add-on therapy to formoterol may be related to altered histone signaling and increased iNOS expression.

Tregs and HLA-DR Expression in Sputum Cells of COPD Patients Treated with Tiotropium and Formoterol

Immune cells expressing the activation markers HLA-DR and regulatory T cells (Tregs) may be involved in the regulation of chronic inflammation in chronic obstructive pulmonary disease (COPD). In this study we analyzed native and activated cell profiles in sputum of 22 stable COPD patients receiving formoterol (F) or formoterol + tiotropium (F + T) for 3 months. Cells were isolated from induced sputum and were examined on Coulter flow cytometer using fluorescent antibodies specific for CD3, CD4, CD8, CD14, CD19, CD25, CD127, and HLA-DR antigens. Cell profiles and cell activation were assessed by analysis of HLA-DR, CD25, and CD127 co-expression in double-stained samples. Tregs were defined as CD4⁺CD25(high) CD127(low) cells. We found that the combined therapy significantly decreased the CD8⁺ cell number (p < 0.01). At baseline, HLA-DR was expressed in about 10 % of sputum T or B cells and a higher expression was found on monocytes. The HLA-DR expression on lymphocytes, but not monocytes, was significantly lower (p < 0.01) in patients treated with F + T. Fractions of activated [CD4⁺ CD25⁺] cells were also significantly lower in the combined therapy group, except for the subpopulation of CD4⁺CD25(high) CD127(low) cells which was not altered. We conclude that tiotropium in add-on therapy to formoterol affects Treg cell profiles and decreases HLA-DR expression in airway lymphocytes.

Costs of Treatment of Chronic Obstructive Pulmonary Disease

The aim of this study was to analyze direct costs of COPD therapy in relation with clinical course and stage of the disease. Sixty patients with moderate to severe COPD were included into the study. The average cost was taken from institutional data file and was also assessed from a social perspective. Results were presented as average costs per patient per year. Forty two percent of patients was classified as GOLD D category, while categories A, B, and C accounted for 8 %, 27 %, and 23 %, respectively. Approximately 65 % of patients had 2-3 degrees of dyspnea according to the Modified Medical Research Council Dyspnea Scale. About 60 % of patients underwent two or three exacerbations per year and those patients had one or two co-morbidities diagnosed. Treatment costs almost doubled with disease progression, mainly due to exacerbations. In patients in Group C and Group D with exacerbations the direct costs were several times higher than in group A or B and the difference increased with progression of the disease. In Groups A and B, the costs of treatment of stable disease or with exacerbation were comparable. We conclude that costs of treatment of COPD patients were highest in advanced disease and were strongly related to COPD exacerbations.

Improvement in 24-hour bronchodilation and symptom control with aclidinium bromide versus tiotropium and placebo in symptomatic patients with COPD: post hoc analysis of a Phase IIIb study

Background A previous Phase IIIb study (NCT01462929) in patients with moderate to severe COPD demonstrated that 6 weeks of treatment with aclidinium led to improvements in 24-hour bronchodilation comparable to those with tiotropium, and improvement of symptoms versus placebo. This post hoc analysis was performed to assess the effect of treatment in the symptomatic patient group participating in the study. Methods Symptomatic patients (defined as those with Evaluating Respiratory Symptoms [E-RS™] in COPD baseline score ≥10 units) received aclidinium bromide 400 μg twice daily (BID), tiotropium 18 μg once daily (QD), or placebo, for 6 weeks. Lung function, COPD respiratory symptoms, and incidence of adverse events (AEs) were assessed. Results In all, 277 symptomatic patients were included in this post hoc analysis. Aclidinium and tiotropium treatment improved forced expiratory volume in 1 second (FEV1) from baseline to week 6 at all time points over 24 hours versus placebo. In addition, improvements in FEV1 from baseline during the nighttime period were observed for aclidinium versus tiotropium on day 1 (aclidinium 157 mL, tiotropium 67 mL; P<0.001) and week 6 (aclidinium 153 mL, tiotropium 90 mL; P<0.05). Aclidinium improved trough FEV1 from baseline versus placebo and tiotropium at day 1 (aclidinium 136 mL, tiotropium 68 mL; P<0.05) and week 6 (aclidinium 137 mL, tiotropium 71 mL; P<0.05). Aclidinium also improved early-morning and nighttime symptom severity, limitation of early-morning activities, and E-RS Total and domain scores versus tiotropium (except E-RS Chest Symptoms) and placebo over 6 weeks. Tolerability showed similar incidence of AEs in each arm. Conclusion In this post hoc analysis of symptomatic patients with moderate to severe COPD, aclidinium 400 μg BID provided additional improvements compared with tiotropium 18 μg QD in: 1) bronchodilation, particularly during the nighttime, 2) daily COPD symptoms (E-RS), 3) early-morning and nighttime symptoms, and 4) early-morning limitation of activity.

Systematic biobanking, novel imaging techniques, and advanced molecular analysis for precise tumor diagnosis and therapy: The Polish MOBIT project

Personalized and precision medicine is gaining recognition due to the limitations by standard diagnosis and treatment; many areas of medicine, from cancer to psychiatry, are moving towards tailored and individualized treatment for patients based on their clinical characteristics and genetic signatures as well as novel imaging techniques. Advances in whole genome sequencing have led to identification of genes involved in a variety of diseases. Moreover, biomarkers indicating severity of disease or susceptibility to treatment are increasingly being characterized. The continued identification of new genes and biomarkers specific to disease subtypes and individual patients is essential and inevitable for translation into personalized medicine, in estimating both, disease risk and response to therapy. Taking into consideration the mostly unsolved necessity of tailored therapy in oncology the innovative project MOBIT (molecular biomarkers for individualized therapy) was designed. The aims of the project are: (i) establishing integrative management of precise tumor diagnosis and therapy including systematic biobanking, novel imaging techniques, and advanced molecular analysis by collecting comprehensive tumor tissues, liquid biopsies (whole blood, serum, plasma), and urine specimens (supernatant; sediment) as well as (ii) developing personalized lung cancer diagnostics based on tumor heterogeneity and integrated genomics, transcriptomics, metabolomics, and radiomics PET/MRI analysis. It will consist of 5 work packages. In this paper the rationale of the Polish MOBIT project as well as its design is presented. (iii) The project is to draw interest in and to invite national and international, private and public, preclinical and clinical initiatives to establish individualized and precise procedures for integrating novel targeted therapies and advanced imaging techniques.