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Theophylline is a natural compound present in tea. Because of its property to relax smooth muscle it is used in pharmacology for the treatment of airway diseases (ie, chronic obstructive pulmonary disease, asthma). However, this effect on smooth muscle is dose dependent and it is related to the development of side effects. Recently, an increasing body of evidence suggests that theophylline, at low concentrations, also has anti-inflammatory effects related to the activation of histone deacetylases. In this study, we evaluated the effects of theophylline alone and in combination with corticosteroids on human bronchial epithelial cells under inflammatory stimuli. Theophylline administrated alone was not able to reduce growth-stimulating signaling via extracellular signal-regulated kinases activation and matrix metalloproteases release, whereas it strongly counteracts this biochemical behavior when administered in the presence of corticosteroids. These data provide scientific evidence for supporting the rationale for the pharmacological use of theophylline and corticosteroid combined drug. © 2017 Gallelli et al. Theophylline is a natural compound present in tea. Because of its property to relax smooth muscle it is used in pharmacology for the treatment of airway diseases (ie, chronic obstructive pulmonary disease, asthma). However, this effect on smooth muscle is dose dependent and it is related to the development of side effects. Recently, an increasing body of evidence suggests that theophylline, at low concentrations, also has anti-inflammatory effects related to the activation of histone deacetylases. In this study, we evaluated the effects of theophylline alone and in combination with corticosteroids on human bronchial epithelial cells under inflammatory stimuli. Theophylline administrated alone was not able to reduce growth-stimulating signaling via extracellular signal-regulated kinases activation and matrix metalloproteases release, whereas it strongly counteracts this biochemical behavior when administered in the presence of corticosteroids. These data provide scientific evidence for supporting the rationale for the pharmacological use of theophylline and corticosteroid combined drug.

Within the context of the heterogeneous phenotypic stratification of asthmatic population, many patients are characterized by moderate-to-severe eosinophilic asthma, not adequately controlled by relatively high dosages of inhaled and even oral corticosteroids. Therefore, these subjects can obtain significant therapeutic benefits by additional biologic treatments targeting interleukin-5 (IL-5), given the key pathogenic role played by this cytokine in maturation, activation, proliferation, and survival of eosinophils. In particular, reslizumab is a humanized anti-IL-5 monoclonal antibody that has been found to be an effective and safe add-on therapy, capable of decreasing asthma exacerbations and significantly improving disease control and lung function in patients experiencing persistent allergic or nonallergic eosinophilic asthma, despite the regular use of moderate-to-high doses of inhaled corticosteroids. These important therapeutic effects of reslizumab, demonstrated by several controlled clinical trials, have led to the recent approval by US Food and Drug Administration of its use, together with other antiasthma medications, for the maintenance treatment of patients suffering from severe uncontrolled asthma.

The humanized monoclonal anti-IgE antibody omalizumab is currently the only biologic drug approved for asthma treatment. Omalizumab inhibits allergic responses by binding to serum immunoglobulins E (IgE), thus preventing their interactions with cellular IgE receptors. Omalizumab is also capable of down-regulating the expression of high affinity IgE receptors on inflammatory cells, as well as the numbers of eosinophils in both peripheral blood and induced sputum. The clinical effects of omalizumab include relevant improvements in respiratory symptoms and quality of life, paralleled by a marked reduction of asthma exacerbations, emergency room visits, and use of systemic corticosteroids and rescue bronchodilators. Moreover, some recent studies suggest potential benefits of omalizumab also in non allergic phenotypes of severe asthma. Very interesting are also further recent reports referring to the potential inhibitory effect of omalizumab with regard to bronchial structural changes, especially occurring in severe asthma and globally defined as airway remodeling. Omalizumab is relatively well tolerated, and only very rarely induces anaphylactic reactions. Therefore, this drug represents a valid option as add-on therapy for patients with severe persistent asthma, inadequately controlled by high doses of standard inhaled treatments.

The propensity to develop an exaggerated antibody response to inhaled antigens operated by immunoglobulin E (IgE), which is defined as atopy, plays a central role in the pathobiology of allergic asthma [1]. The pleiotropic effects of IgE are mediated by activation of specific IgE receptors expressed by both immune-inflammatory and airway structural cells [2–4]. IgE were identified in 1967 by Ishizaka and Ishizaka, and since then these antibodies have been considered as suitable molecular targets for the development of anti-allergy therapies [5, 6]. However, it took almost 40 years to translate this key immunological discovery into the approval of the anti-IgE antibody omalizumab for the treatment of severe allergic asthma [7]. Indeed, omalizumab has been the first and for a long time the only biologic drug available in clinical practice for add-on therapy of uncontrolled asthma [8].

Tumour necrosis factor-α (TNF-α) plays a relevant role in many features of asthma pathobiology by exerting its effects on both airway inflammatory and structural cells (Fig. 7.1) [1]. This pleiotropic cytokine is produced by several cell types including Th1 lymphocytes, macrophages and mast cells and induces the recruitment of neutrophils and eosinophils into the airways via up-regulation of epithelial and endothelial adhesion molecules [2, 3]. Moreover, by enhancing TGF-β expression, TNF-α stimulates airway smooth muscle cell proliferation, as well as fibroblast growth and maturation into myofibroblasts [4–6]. Therefore, TNF-α can contribute to the development of severe asthma not only by inducing airway inflammation and hyperresponsiveness but also by promoting bronchial remodelling [7]. Indeed, TNF-α is overexpressed in the airways of patients with severe asthma and also directly stimulates airway smooth muscle contraction through changes in intracellular calcium fluxes [1].

Interleukin-25 (IL-25), interleukin-33 (IL-33) and thymic stromal lymphopoietin (TSLP) are innate cytokines produced by the airway epithelium, which play a key role in asthma pathobiology by contributing to the development of Th2-mediated adaptive immune responses [1]. In addition to stimulating Th2 cell-dependent allergic bronchial inflammation, IL-25, IL-33 and TSLP are also crucially involved in non-allergic eosinophilic asthma because of their ability to induce type 2 innate lymphoid cells (ILC2s) to produce T2-type cytokines such as IL-5, IL-9 and IL-13 [2]. Therefore, IL-25, IL-33 and TSLP are suitable targets for antiasthma biological therapies [3].

Potentially suitable targets for antiasthma treatments also include other pro-inflammatory cytokines such as interleukin-9 (IL-9), granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as interleukin-17 (IL-17) and interleukin-23 (IL-23) [1–4].

Even if severe asthma (SA) accounts for 5–10% of all cases of the disease, it is currently a crucial unmet need, owing its difficult clinical management and its high social costs. For this reason several networks, focused on SA have been organized in some countries, in order to select these patients, to recognize their clinical features, to evaluate their adherence, to classify their biological/clinical phenotypes, to identify their eligibility to the new biologic therapies and to quantify the costs of the disease. Aim of the present paper is to describe the ongoing Italian Severe Asthma Network (SANI). Up today 49 centres have been selected, widespread on the national territory. Sharing the same diagnostic protocol, data regarding patients with SA will be collected and processed in a web platform. After their recruitment, SA patients will be followed in the long term in order to investigate the natural history of the disease. Besides clinical data, the cost/benefit evaluation of the new biologics will be verified as well as the search of peculiar biomarker(s) of the disease.

Dupilumab is a fully human IgG4 monoclonal antibody directed against the α subunit of the interleukin (IL)-4 receptor (IL-4Rα). Since the activation of IL-4Rα is utilized by both IL-4 and IL-13 to mediate their pathophysiological effects, dupilumab behaves as a dual antagonist of these two sister cytokines, which blocks IL-4/IL-13-dependent signal transduction. Areas covered: Herein, the authors review the cellular and molecular pathways activated by IL-4 and IL-13, which are relevant to asthma pathobiology. They also review: the mechanism of action of dupilumab, the phase I, II and III studies evaluating the pharmacokinetics as well as the safety, tolerability and clinical efficacy of dupilumab in asthma therapy. Expert opinion: Supported by a strategic mechanism of action, as well as by convincing preliminary clinical results, dupilumab currently appears to be a very promising biological drug for the treatment of severe uncontrolled asthma. It also may have benefits to comorbidities of asthma including atopic dermatitis, chronic sinusitis and nasal polyposis.

Continuous spreading of HIV infection may be due to lack of knowledge, especially among young people. It is important to monitor level of knowledge and risk behaviours especially in young people to set up appropriate preventative and informative campaigns.We assessed knowledge of HIV infection and risk factors in a sample of students at the "Magna Graecia" University of Catanzaro, using an anonymous multiple-choice questionnaire.Two-hundred and sixteen medical students attending the first year at "Magna Graecia" University (age: 18-20 years) were prescribed an anonymous multiple-choice questionnaire. Knowledge of HIV infection was scarce in a significant percentage of students. They were often practicing risk behaviours at risk for acquisition of HIV infection and other sexually transmitted infections.This study shows that preventative and informative campaigns are urgently needed in earlier stage of adolescence to avoid acquisition of HIV infection and other sexually transmitted diseases.

Because of the central role played by interleukin-5 (IL-5) in maturation, activation, proliferation and survival of eosinophils, this cytokine is a key target for treatment of eosinophilic asthma [1–4]. In this regard it is noteworthy that, among the pleiotropic effects of corticosteroids, inhibition of IL-5 synthesis is one of the most important mechanisms underlying the very effective antiasthma action of these drugs [5]. Corticosteroids are indeed powerful inducers of eosinophil apoptosis [6, 7]; nevertheless, despite a regular or almost continuous use of inhaled and even systemic corticosteroids, some subgroups of asthmatic subjects display persistent bronchial and/or blood eosinophilia, associated with an inadequate control of asthma [8]. Therefore, these patients can potentially benefit from additional therapies based on the use of biologic drugs targeting IL-5.

Airway inflammation, sustained by a multitude of pro-inflammatory mediators such as cytokines and chemokines produced by both immune-inflammatory and airway structural cells, is a hallmark of asthma [1, 2]. Several inflammatory phenotypes of asthma have been characterized, which include eosinophilic, neutrophilic, mixed and paucigranulocytic patterns [3, 4]. Eosinophils are the inflammatory cells most frequently infiltrating the airways of asthmatic patients; indeed, their maturation, activation, survival and recruitment within the bronchial wall and airway lumen are crucially implicated in the development of both allergic and non-allergic asthma [5, 6]. Eosinophilic asthma originates from the activation of immunopathologic and pro-inflammatory pathways, mainly coordinated by T-helper 2 (Th2) lymphocytes, which release interleukin-5, interleukin-4 and interleukin-13 (IL-5, IL-4 and IL-13) (Fig. 2.1). In addition to being driven by adaptive immune responses, airway eosinophilia can also arise from innate immune mechanisms, which are mediated by intercellular communications involving dendritic cells, bronchial epithelial cells and innate lymphoid cells [7, 8]. Whilst bronchial eosinophilic infiltration is mostly responsible for mild-to-moderate asthma, more severe disease is often characterized by mixed patterns of inflammation including both eosinophils and neutrophils, with the latter which can represent the predominant inflammatory cells detectable in the induced sputum obtained from patients experiencing uncontrolled asthmatic symptoms and exacerbations. Neutrophilic airway inflammation, associated with severe asthma, is triggered by Th1 and especially Th17 lymphocytes (Fig. 2.1) [9, 10]. Within this pathobiologic context, a pivotal role is played by dendritic cells, which direct the commitment to the various Th lineages. In particular, polarization towards the different Th subsets is closely linked to the particular airway milieu consisting of specific co-stimulatory molecules and cytokines, which drive the various Th cell programmes [11].

In addition to chronic inflammation, both allergic and non-allergic phenotypes of asthma are also frequently characterized by dynamic structural changes which involve all layers of the bronchial wall, including large and small airways [1, 2]. Such structural changes, collectively referred as airway remodelling, likely originate from repetitive cycles of tissue injury, causing subsequent cellular responses leading to abnormal repair [3]. Although the concomitance in asthmatic airways of structural modifications and inflammation had previously suggested that the latter could causally determine the development and progression of bronchial remodelling, these two phenomena are currently believed to be quite independent [4, 5]. Indeed, airway remodelling can start during the very initial stages of the natural history of asthma, even at early childhood well before a long-term firm establishment of chronic inflammation [6]. However, close interactions tightly connect the cellular and molecular mechanisms underpinning airway inflammation and remodelling (Fig. 3.1), and both pathologic processes certainly contribute together to induce bronchial hyperresponsiveness as well as asthma persistence and progression, especially in most severe disease phenotypes [7–9].

During the last years, asthma treatment has entered a new fascinating era of trustworthy realities and promising perspectives [1]. These significant advances have been made possible by the recent relevant progress achieved towards a better understanding of the cellular and molecular mechanisms underlying asthma pathobiology [2, 3]. Undoubtedly, the most important new therapeutic tools available for the treatment of asthma, especially with regard to the more severe disease phenotypes, are biologic drugs [4]. Indeed, omalizumab and mepolizumab are already included in the step 5 of GINA guidelines for asthma management [5], and reslizumab has been recently approved by FDA [6]. Moreover, the current intense development of molecular-targeted therapies, many of which are undergoing advanced stages of clinical investigation, will probably lead in the next future to the introduction in real life of several other biologics indicated for asthma treatment [7–11].

Current advances in the knowledge of asthma pathobiology suggest that anticytokine therapies can be potentially useful for the treatment of this complex and heterogeneous airway disease. Recent evidence is accumulating in support of the efficacy of anti-IL-4, anti-IL-5, and anti-IL-13 drugs. Therefore, these new developments are now changing the global scenario of antiasthma therapies, especially with regard to more severe disease. Current findings referring to variability of individual therapeutic responses highlight that the different asthma subtypes need to be well characterized, in order to implement phenotype-targeted treatments which in the near future will hopefully be mainly based on cytokine-directed biologics.

Inhaled glucocorticoids are the mainstay of asthma treatment. Indeed, such therapeutic agents effectively interfere with many pathogenic circuits underpinning asthma. Among these drugs, during the last decades budesonide has been probably the most used molecule in both experimental studies and clinical practice. Therefore, a large body of evidence clearly shows that budesonide, either alone or in combination with long-acting bronchodilators, provides a successful control of asthma in many patients ranging throughout the overall spectrum of disease severity. These excellent therapeutic properties of budesonide basically depend on its molecular mechanisms of action, capable of inhibiting within the airways the activity of multiple immune-inflammatory and structural cells involved in asthma pathobiology.

Girolamo Pelaia,1 Alessandro Vatrella,2 Maria Teresa Busceti,1 Luca Gallelli,3 Cecilia Calabrese,4 Rosa Terracciano,3 Nicola Lombardo,1 Rosario Maselli11Department of Medical and Surgical Sciences, Magna Græcia University of Catanzaro, Catanzaro, 2Departmentof Medicine and Surgery, University of Salerno, Salerno, 3Department of Health Science, Magna Græcia University of Catanzaro, Catanzaro, 4Department of Cardio-Thoracic and Respiratory Sciences, Second University of Naples, Naples, ItalyAbstract: Bronchodilators are the most important drugs used for the treatment of chronic obstructive pulmonary disease (COPD). In particular, these therapeutic agents are mostly long-acting compounds utilized via inhalation, and include LAMA (long-acting muscarinic receptor antagonists) and LABA (long-acting β2-adrenoceptor agonists). Because LAMA and LABA induce bronchodilation by distinct mechanisms of action, LABA/LAMA combinations provide a reciprocal potentiation of the pharmacological effects caused by each component. Hence, many COPD patients who do not achieve a satisfactory control of their symptoms using a single, either LAMA or LABA bronchodilator, can experience relevant benefits with the use of LAMA/LABA fixed combinations. Many different LAMA/LABA combinations have been recently developed and evaluated in randomized clinical trials. In this context, our review focuses on the pharmacological mechanisms underpinning the bronchodilation elicited by the LAMA tiotropium bromide and the LABA olodaterol. We also discuss the results of the most important clinical studies carried out in COPD patients to assess the efficacy and safety of tiotropium/olodaterol combinations.Keywords: LAMA, LABA, tiotropium, olodaterol, dual bronchodilation, tiotropium/olodaterol combinations

Induced sputum is recognized as being of increasing importance for the diagnosis and monitoring of chronic inflammatory lung diseases. The main purpose of this study is to provide a valid approach to better fractionate and characterize the still under-estimated low-molecular weight proteome of induced sputum by using mesoporous silica beads (MSBs) SPE coupled to MALDI-TOF MS. Sputum peptides were captured from both derivatized and non-derivatized MSBs and then profiled by MALDI-TOF MS. Depending on the chemical groups present on the mesoporous surface, complex peptide mixtures were extracted from induced sputum and converted into reproducible MALDI profiles. The number of peaks detected as a function of S/N was evaluated for each mesoporous surface. More than 400 peaks with an S/N>5 were obtained in comparison to 200 peaks detected without MSBs. Additionally, as a proof-of-principle, we investigated the ability of this platform to discriminate between the "sputome" of patients with asthma and chronic obstructive pulmonary disease, and between these groups and those of healthy control subjects. Six m/z peaks emerged as potential diagnostic peptidic patterns able to differentiate these inflammatory airway diseases in the sputome range. Human α-defensins (human neutrophil peptide (HNP)1, HNP2, HNP3) and three C-terminal amidated peptides, one of which is phosphorylated on serine, were identified by MALDI-TOF/TOF MS. These findings may contribute to defining a high-throughput screening MS-based platform for monitoring key peptidic-biomarkers for inflammatory and chronic respiratory diseases in induced sputum samples.

Omalizumab is a humanized murine monoclonal antibody directed toward a portion of the IgE indicated in Europe for the treatment of severe persistent allergic asthma, inadequately controlled despite high-dose of ICS (mean BDP equivalent dose of inhaled corticosteroid 2224.68microg/die) in association with long-acting beta(2) agonists. Our aim was to describe the experience, efficacy and safety in a cohort of Italian patients treated with omalizumab in a real-life clinical setting. One hundred and forty two patients from 13 Italian Centers were observed and analysed. The dosage of omalizumab was established according to the labelling indication, with a median dose of IgE of 297.38IU/ml or kU/l. During the previous year, all patients experienced frequent exacerbations (mean=4.87), emergency visits (mean=4.45) and hospitalisation (mean=1.53). Following treatment with omalizumab, the annual rate of exacerbations, emergency visits and hospitalisation decreased by 79%, 88% and 95%, respectively. The proportion of patients without exacerbation, not needing emergency visits and hospitalization increased by 610%, 154% and 28%, respectively. The response to omalizumab measured with the GETE (global evaluation of treatment effectiveness) scale rated as good to excellent in 77% of patients. Overall, 9.6% (n=9) of the patients experienced one single adverse effect. Only one patient reported a serious adverse event (local reaction at the site of injection) leading to interruption of treatment. The observed reduction of asthma-related events in particularly poorly controlled patients in this Italian real-life setting is consistent with the results of other observational studies.

Bronchial hyperresponsiveness and airway infiltration with eosinophils and T lymphocytes are key features of asthma. In particular, CD4+ T cells are currently believed to play a pivotal role as initiators and coordinators of the asthmatic inflammatory response and, therefore, they represent a crucial target of corticosteroid treatment. The aim of the present investigation is thus to evaluate, in patients with mild asthma, the effects of inhaled corticosteroid therapy on the following parameters: (i) functional state of CD4+ T cells; (ii) airway eosinophilia; (iii) bronchial hyperresponsiveness to methacholine. The study was completed by twenty asthmatic, atopic subjects, subdivided into two groups of ten and treated for 12 weeks with either inhaled budesonide (200 μg twice daily) or terbutaline alone (500 μg twice daily), respectively. Expression of CD4+ T cell activation markers was measured in induced sputum at baseline and after 1, 4, 8 and 12 weeks of treatment by flow cytometry, which showed a down-regulation of HLA-DR and CD25 surface proteins in the budesonide group, compared with the control group; these differences resulted as being statistically significant through weeks 4–12. Budesonide also induced a quick, sharp reduction in the percentage of eosinophils detectable in induced sputum, as well as a more gradual progressive improvement in airway hyperresponsiveness to methacholine. Therefore, in addition to assessing various indices of bronchial inflammation, flow cytometry can be reliably applied to induced sputum in order to monitor, even in mildly symptomatic patients, the effects of anti-asthma treatments on T cell activation.

Chronic obstructive pulmonary disease (COPD) is a complex respiratory disorder, whose medical and socioeconomic burden as one of the main causes of morbidity and mortality is increasing worldwide. COPD pathophysiology includes chronic airway/lung inflammation and progressive airflow limitation. Therefore, anti-inflammatory and bronchodilator agents are key players in COPD treatment.This review article discusses the complementary molecular interactions connecting the respective mechanisms of action of inhaled corticosteroids (ICS) and long-acting β2-adrenergic agonists (LABAs). Moreover, attention is also focused on clinical trials, which have shown that ICS/LABA combinations are very effective in improving COPD symptoms and lung function, being also able to significantly reduce disease exacerbations.In selected subgroups of COPD patients, ICS/LABA combinations represent a very useful therapeutic approach for this widespread chronic respiratory disease. In addition to the well-known fixed-dose drug associations such as fluticasone propionate/salmeterol xinafoate and budesonide/formoterol fumarate, other newly developed ICS/LABA combinations are currently emerging as very interesting pharmacological options for COPD treatment.

COPD is a widespread inflammatory respiratory disorder characterized by a progressive, poorly reversible airflow limitation. Currently available therapies are mostly based on those used to treat asthma. However, such compounds are not able to effectively reduce the gradual functional deterioration, as well as the ongoing airway and lung inflammation occurring in COPD patients. Therefore, there is an urgent need to improve the efficacy of the existing drug classes and to develop new treatments, targeting the main cellular and molecular mechanisms underlying disease pathogenesis. These therapeutic strategies will be highlighted in the present review.

Lung cancer is characterized by a high mortality rate probably attributable to early metastasis. Oxidative stress is involved in development and progression of lung cancer, through cellular and molecular mechanisms which at least in part overlap with proinflammatory pathways. Simvastatin is a statin with pleiotropic effects that can also act as an anti-oxidant agent, and these pharmacologic properties may contribute to its potential anti-cancer activity. Therefore, the aim of this study was to evaluate, in the human lung adenocarcinoma cell line GLC-82, the effects of a 24-hour treatment with simvastatin on hydrogen peroxide (H2O2)-induced changes in cell viability, ERK phosphorylation, matrix metalloproteinase (MMP) expression, innate immunity signaling, NF-κB activation and IL-8 secretion. Cell counting was performed after trypan blue staining, cell proliferation was assessed using MTT assay, and apoptosis was evaluated through caspase-3 activation and Tunel assay. Western blotting was used to analyze protein extracts, and IL-8 release into cell culture supernatants was assessed by ELISA. Our results show that simvastatin (30 μM) significantly (P < 0.01) inhibited the proliferative effect of H2O2 (0.5 mM) and its stimulatory actions on ERK1/2 phosphorylation, NF-κB activation and IL-8 production. Furthermore, simvastatin decreased H2O2-mediated induction of the cellular expression of MMP-2 and MMP-9, as well as of several components of the signaling complex activated by innate immune responses, including MyD88, TRAF2, TRAF6 and TRADD. In conclusion, these findings suggest that simvastatin could play a role in prevention and treatment of lung cancer via modulation of important proinflammatory and tumorigenic events promoted by oxidative stress.. © 2014 Gallelli et al.

Genetic factors play a key role in determining the widely heterogeneous response to pharmacological treatment detectable among asthmatics. In particular, polymorphisms of the genes encoding relevant anti-asthma drug targets contribute significantly to such a variability. Therefore, it is very important to characterize asthmatic patient's genotypes and the related phenotypic patterns, in order to predict the individual therapeutic outcome. This pharmacogenetic approach will eventually help clinicians to optimize and personalize anti-asthma treatment, and will also provide useful information with regard to pre- and post-marketing evaluation of both effectiveness and side effects of newly introduced drugs.

The airway epithelium is continuously exposed to inhaled oxidants, including airborne pollutants and cigarette smoke, which can exert harmful proinflammatory and cytotoxic effects. Therefore, the aim of our study was to investigate, in primary cultures of human bronchial epithelial cells (HBEC), the signal transduction pathways activated by increasing concentrations (0.25, 0.5, and 1 mM) of hydrogen peroxide (H(2)O(2)), as well as their effects on IL-8 production and cell viability. The reported results show that H(2)O(2) elicited, in a concentration-dependent fashion, a remarkable increase in phosphorylation-dependent activation of mitogen-activated protein kinases (MAPKs), associated with a significant induction of IL-8 synthesis and a dramatically enhanced cell death. Pre-treatment of HBEC with MAPK inhibitors was able to significantly inhibit the effects of H(2)O(2) on IL-8 secretion, and to effectively prevent cell death. Therefore, these findings suggest that MAPKs play a key role as molecular transducers of the airway epithelial injury triggered by oxidative stress, as well as potential pharmacologic targets for indirect antioxidant intervention.

Non-small cell lung cancer (NSCLC) is the leading cause of cancer death in the world. Traditional chemotherapy for advanced NSCLC is often considered excessively toxic. Recent clinical trials documented that gemcitabine may represent a good therapeutical option in patients with NSCLC. Aim of our research was to retrospectively evaluate the adverse effects induced by gemcitabine in patients with NSCLC from 1 January 1997 to 31 December 2002, in clinical records of Oncology Divisions of "S. Giovanni di Dio" Hospital of Crotone, "Ospedali Riuniti" Hospital of Reggio Calabria, Hospital of Paola, and in Pneumological Oncology Division of "Mariano Santo" Hospital of Cosenza, Italy. Clinical records of patients treated with gemcitabine (1000mgm(-2) on days 1 and 8) were reviewed and following data were obtained: sex and age of the patients, histologic diagnosis and disease stage, World Health Organisation (WHO) performance status and toxic effects induced by gemcitabine. We reported that 71.6% of NSCLC patients (age range 48-77 years; 135 males, 27 females; performance status 0=53, 1=109) were eligible for our study. Side effect of gemcitabine involved gastrointestinal system (nausea, vomiting and diarrhoea) and only in the last cycles (VIII-XI) emopoiethic system (leukopenia, neutropenia, thrombocytopenia and anemia). Grade IV vomiting occurred in three patients, thrombocytopenia in two. Grade III leukopenia was observed in three patients. Other toxicities were mild. None of the patients died during chemotherapy. In conclusion, these data showed that gemcitabine present a very good tolerability in patients with NSCLC. Therefore, it could be considered as a new therapeutic agents to use as first line therapy for this disease.

We retrospectively analysed the adverse drug reactions (ADRs) associated with bronchodilator therapy and reported over a 7-year period, from January 1995 to December 2001, in clinical notes of two Pulmonary division of "Mater Domini" University Hospital and "Pugliese-Ciaccio" Hospital, both located in Catanzaro, Italy. Bronchodilators were responsible for 45 (18.5%) out of 243 episodes of ADRs. Theophylline was the drug most involved in ADRs (53.4%), and skin was the body system most susceptible to ADRs induced by all bronchodilators (47.7%). We determined that the drug-ADR relationship was certain in 73% of the reports; withdrawal of the suspected drug led to recovery in 86% of cases. In conclusion, this retrospective evaluation demonstrated that bronchodilators are a common cause of ADRs in hospitalised patients and, therefore, drug surveillance can successfully identify adverse events related with drug administration in hospitalised patients.

Bronchodilators are the cornerstone of the treatment of chronic obstructive pulmonary disease (COPD). In particular, the most commonly used drugs are inhaled long-acting agents, including long-acting F2-adrenergic agonists (LABAs) and long-acting muscarinic receptor antagonists (LAMAs). The combination of a LABA with a LAMA, i.e. of molecules characterized by different mechanisms of action, results in a synergistic enhancement of their clinical and functional effects. Therefore, this combined treatment can be implemented in a number of cases in which disease control is not adequately achieved by a single active agent such as a LABA or a LAMA. Several LABA/LAMA fixed-dose combinations, mainly made up of newly developed compounds, are currently in advanced phases of experimental evaluation. Within such a context, the aim of this review is to outline the pharmacological basis of dual bronchodilation as well as to discuss the results of the main trials carried out using the drug combination consisting of indacaterol and glycopyrronium, a LABA and a LAMA recently introduced in the treatment of COPD. i 2014 S. Karger AG, Basel

Simultaneously with the steady progress towards a better knowledge of the pathobiology of asthma, the potential usefulness of anticytokine therapies is emerging as one of the key concepts in the newly developing treatments of this widespread airway disease. In particular, given the key role played by interleukin (IL)-4 and IL-13 in the pathophysiology of the most typical aspects of asthma, such as chronic airway inflammation, tissue remodeling, and bronchial hyperresponsiveness, these pleiotropic cytokines are now considered as suitable therapeutic targets. Among the recently developed antiasthma biologic drugs, the monoclonal antibody dupilumab is very promising because of its ability to inhibit the biological effects of both IL-4 and IL-13. Indeed, dupilumab prevents IL-4/13 interactions with the α-subunit of the IL-4 receptor complex. A recent trial showed that in patients with difficult-to-control asthma, dupilumab can markedly decrease asthma exacerbations and improve respiratory symptoms and lung function; these effects were paralleled by significant reductions in T-helper 2-associated inflammatory biomarkers. However, further larger and longer trials are required to extend and validate these preliminary results, and also to carefully study the safety and tolerability profile of dupilumab.

The most recent guidelines define COPD in a multidimensional way, nevertheless the diagnosis is still linked to the limitation of airflow, usually measured by the reduction in the FEV1/FVC ratio below 70%. However, the severity of obstruction is not directly correlated to symptoms or to invalidity determined by COPD. Thus, besides respiratory function, COPD should be evaluated based on symptoms, frequency and severity of exacerbations, patient’s functional status and health related quality of life (HRQoL). Therapy is mainly aimed at increasing exercise tolerance and reducing dyspnea, with improvement of daily activities and HRQoL. This can be accomplished by a drug-induced reduction of pulmonary hyperinflation and exacerbations frequency and severity. All guidelines recommend bronchodilators as baseline therapy for all stages of COPD, and long-acting inhaled bronchodilators, both beta-2 agonist (LABA) and antimuscarinic (LAMA) drugs, are the most effective in regular treatment in the clinically stable phase. The effectiveness of bronchodilators should be evaluated in terms of functional (relief of bronchial obstruction and pulmonary hyperinflation), symptomatic (exercise tolerance and HRQoL), and clinical improvement (reduction in number or severity of exacerbations), while the absence of a spirometric response is not a reason for interrupting treatment, if there is subjective improvement in symptoms. Because LABA and LAMA act via different mechanisms of action, when administered in combination they can exert additional effects, thus optimizing (i.e. maximizing) sustained bronchodilation in COPD patients with severe airflow limitation, who cannot benefit (or can get only partial benefit) by therapy with a single bronchodilator. Recently, a fixed combination of ultra LABA/LAMA (indacaterol/glycopyrronium) has shown that it is possible to get a stable and persistent bronchodilation, which can help in avoiding undesirable fluctuations of bronchial calibre.

Mitogen-activated protein kinases (MAPK) play a central role in signal transduction by regulating many nuclear transcription factors involved in inflammatory, immune, and proliferative responses. The aim of this study was to investigate, in human pulmonary endothelial cells, the effects of synthetic glucocorticosteroids on activation of c-jun N-terminal kinases, extracellular signal-regulated kinases, and p38 subgroups of the MAPK family. Human microvascular endothelial cells from lung were stimulated for 2 h with either H 2O2 (2 mM), IL-1 (10 ng/mL), or tumour necrosis factor- (10 ng/mL). Under these conditions, a remarkable increase in the phosphorylation pattern of c-jun N-terminal kinases, extracellular signal-regulated kinases 1/2, and p38 was detected. Pretreatment for 12 h with dexamethasone (100 nM) was able to prevent phosphorylation-dependent MAPK activation in stimulated cells, without substantially affecting the expression levels of these enzymes. Our results suggest that inhibition of MAPK signaling pathways in human pulmonary endothelial cells may significantly contribute, by interfering with activation of several different transcription factors, to the antiinflammatory and immunosuppressive effects of glucocorticosteroids. © 2001 Elsevier Science Inc. All rights reserved.

The role of polymorphonuclear leukocytes (PMNs) in the immune defence against intracellular bacteria has long been neglected. Only recently have studies begun to address this issue. In this study the behavior of peripheral PMNs in Bacillus Calmette-Guerin (BCG) vaccinated subjects was investigated. Twenty healthy and purified protein derivative-negative adults were studied before, and two and four months after, BCG administration. Luminol-amplified chemiluminescence (CL) emission was evaluated in whole blood phagocytes using a soluble stimulus, such as phorbol mirystate acetate, or particulates such as zymosan opsonized with homologous (OZH) or autologous (OZA) serum. Specific IgG, IgA and IgM against antigen -60 by ELISA, total immunoglobulin, C3 and C4 components of complement, were assessed by immunochemical tests. The results revealed a late heightened production of reactive oxygen intermediates in vaccinated subjects in presence of OZA and OZH. Our findings confirm that the role of PMNs and their mediators in immunoregulation of intracellular diseases needs to be re-evaluated. © 2000 John Wiley & Sons, Ltd.

Combined pulmonary fibrosis and emphysema (CPFE) is a computed tomography (CT)-defined syndrome of combined pulmonary fibrosis and emphysema, characterized by subnormal spirometry, impairment of gas exchange, and high prevalence of pulmonary hypertension. Although endothelin-1 (ET-1) plays an important role in the development of lung fibrosis as well as in pulmonary hypertension, no ET-1-targeted therapy is currently recommended. Here we report a case of CPFE successfully treated with ambrisentan, an endothelin-A receptor antagonist, and also discuss the biologic mechanisms underlying the observed therapeutic effects. A 79-year-old man with chronic obstructive pulmonary disease (COPD) was referred to our respiratory unit as an outpatient for dyspnea. Clinical, radiologic, and laboratory findings suggested a diagnosis of chronic hypoxemic, type 1 respiratory failure, due to combined pulmonary fibrosis and emphysema, complicated by severe, precapillary pulmonary hypertension. Pharmacologic treatment with ambrisentan induced an initial improvement in clinical symptoms that proved to be very relevant 9 months later. In order to investigate the biologic mechanisms underlying the clinical effects of ambrisentan, we performed an "in vitro" study on primary cultures of fibrotic human lung fibroblasts, as well as on human umbilical vein endothelial cells, incubated for 24 and 48 h with ET-1, in the absence or presence of an overnight treatment with ambrisentan. ET-1 significantly increased cell proliferation and mitogen-activated protein kinase activation (P < 0.01). These effects were significantly (P < 0.01) inhibited by ambrisentan in both cell cultures. In conclusion, we hypothesize that the clinical benefits induced by ambrisentan in this patient with CPFE can be attributed to its vasodilator and anti-proliferative actions, exerted on pulmonary the vascular bed and lung fibroblasts. © 2013 Springer International Publishing Switzerland.

IgE antibodies are crucially involved in mediating, maintaining and amplifying the allergic cascade. The humanized monoclonal anti-IgE antibody omalizumab is currently the only biologic drug approved for asthma treatment. Anti-IgE inhibits allergic responses by binding to serum IgE, thus preventing their interactions with cellular IgE receptors. Omalizumab is also capable of down-regulating the expression of high affinity IgE receptors on inflammatory cells, as well as the numbers of eosinophils in both peripheral blood and induced sputum. Randomized clinical trials showed relevant clinical effects of omalizumab including improvements of respiratory symptoms and quality of life. Moreover, a marked reduction of asthma exacerbations, emergency room visits, and use of systemic corticosteroids and rescue bronchodilators was also observed. Omalizumab is relatively well tolerated, and only rarely induces anaphylactic reactions. Therefore, this drug represents a valid option as add-on therapy for most severe patients with persistent allergic asthma, inadequately controlled by high doses of standard treatments.

Chronic obstructive pulmonary disease (COPD) is a complex disorder involving both airways and lung parenchyma, usually associated with progressive and poorly reversible airflow limitation. In order to better characterize the phenotypic heterogeneity and the prognosis of patients with COPD, there is currently an urgent need for discovery and validation of reliable disease biomarkers. Within this context, proteomic and peptidomic techniques are emerging as very valuable tools that can be applied to both systemic and pulmonary samples, including peripheral blood, induced sputum, exhaled breath condensate, bronchoalveolar lavage fluid, and lung tissues. Identification of COPD biomarkers by means of proteomic and peptidomic approaches can thus also lead to discovery of new molecular targets potentially useful to improve and personalize the therapeutic management of this widespread respiratory disease.

Objective 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors (statins) can affect post-translational processes, thus being responsible for decreased farnesylation and geranylgeranylation of intracellular small G proteins such as Ras, Rho and Rac, essential for cell survival and proliferation. In this regard, recent in vitro and in vivo studies suggest a possible role for both statins and farnesyl transferase inhibitors in the treatment of malignancies. Within such a context, the aim of our study was to investigate effects of either simvastatin (at concentrations of 1, 15, and 30 μm) or the farnesyl transferase inhibitor R115777 (at concentrations of 0.1, 1, and 10 μm), on two cultures of human non-small lung cancer cells, adenocarcinoma (GLC-82) and squamous (CALU-1) cell lines. In particular, we evaluated actions of these two drugs on phosphorylation of the ERK1/2 group of mitogen-activated protein kinases and on apoptosis, plus on cell numbers and morphology. Materials and Methods Western blotting was used to detect ERK phosphorylation, and to assess apoptosis by evaluating caspase-3 activation; apoptosis was also further assessed by terminal deoxynucleotidyl-mediated dUTP nick end labelling (TUNEL) assay. Cell counting was performed after trypan blue staining. Results and conclusion In both GLC-82 and CALU-1 cell lines, simvastatin and R115777 significantly reduced ERK phosphorylation; this effect, which reached the greatest intensity after 36 h treatment, was paralleled by a concomitant induction of apoptosis, documented by significant increase in both caspase-3 activation and TUNEL-positive cells, associated with a reduction in cell numbers. Our results thus suggest that simvastatin and R115777 may exert, in susceptible lung cancer cell phenotypes, a pro-apoptotic and anti-proliferative activity, which appears to be mediated by inhibition of the Ras/Raf/MEK/ERK signalling cascade.

Here, the authors discuss how cytokines could be used as therapeutic targets for individuals with asthma that is inadequately controlled using current therapies. They also highlight the need for phenotyping asthma subtypes in order to achieve the best possible patient-focused treatment. Recent advances in the knowledge of asthma pathobiology suggest that biological therapies that target cytokines can be potentially useful for the treatment of this complex and heterogeneous airway disease. The use of biologics in asthma has been established with the approval of the humanized monoclonal immunoglobulin E-targeted antibody omalizumab (Xolair; Genentech/Novartis) as an add-on treatment for inadequately controlled disease. Furthermore, evidence is accumulating in support of the efficacy of other biologics, such as interleukin-5 (IL-5)- and IL-13-specific drugs. Therefore, these new developments are changing the scenario of asthma therapies, especially with regard to more severe disease. The variability among patients' individual therapeutic responses highlights that it will be necessary to characterize the different asthma subtypes so that phenotype-targeted treatments based on the use of biologics can be implemented.

Non-typeable Haemophilus influenzae (NTHi) is one of the most frequently involved pathogens in bacterial exacerbations of chronic obstructive pulmonary disease (COPD). In the airways, the main tissue target of NTHi is bronchial epithelium, where this pathogen can further amplify the inflammatory and structural changes induced by proinflammatory cytokines such as tumour necrosis factor-alpha (TNF-alpha). Therefore, the aim of this study is to investigate, in primary cultures of human bronchial epithelial cells, the effects of NTHi on signal transduction pathways, apoptotic events and chemokine production activated by TNF-alpha. Moreover, we also evaluated the effects exerted on such cellular and molecular phenomena by a corticosteroid drug. p38 mitogen-activated protein kinase (MAPK) phosphorylation was analyzed by Western blotting, using an anti-phospho-p38 MAPK monoclonal antibody. Apoptosis was assayed by active caspase-3 expression. Interleukin-8 (IL-8/CXCL8) was detected in cell-free culture supernatants by ELISA. TNF-alpha induced a significant increase in p38 MAPK phosphorylation. NTHi was able to potentiate the stimulatory actions of TNF-alpha on caspase-3 expression and, to a lesser extent, on IL-8 secretion. These effects were significantly (P less than 0.01) inhibited by a pharmacological pre-treatment with budesonide. These results suggest that TNF-alpha is able to stimulate, via activation of p38 MAPK signalling pathway, IL-8 release and airway epithelial cell apoptosis; the latter effect can be markedly potentiated by NTHi. Furthermore, budesonide can be very effective in preventing, through inhibition of p38 MAPK phosphorylation, both structural and proinflammatory changes elicited in bronchial epithelium by TNF-alpha and NTHi.

SummaryAirway smooth muscle (ASM) plays a key role in bronchomotor tone, as well as in structural remodeling of the bronchial wall. Therefore, ASM contraction and proliferation significantly participate in the development and progression of asthma. Many contractile agonists also behave as mitogenic stimuli, thus contributing to frame a hyperresponsive and hyperplastic ASM phenotype. In this review, the molecular mechanisms and signaling pathways involved in excitation–contraction coupling and ASM cell growth will be outlined. Indeed, the recent advances in understanding the basic aspects of ASM biology are disclosing important cellular targets, currently explored for the implementation of new, more effective anti-asthma therapies.

OBJECTIVES: Both interleukin-6 (IL-6) and transforming growth factor-beta (TGF-beta) are crucially involved in fibrotic events that characterize interstitial lung diseases (ILD). Therefore, the aim of this study was to investigate in primary cultures of normal and fibrotic human lung fibroblasts (HLF), exposed to either IL-6 or TGF-beta1, the effects on phosphorylation of mitogen-activated protein kinases (MAPK) and cell growth of IL-6 signalling inhibition, performed by the IL-6 receptor superantagonist Sant7. MATERIALS AND METHODS: MAPK phosphorylation was detected by Western blotting, HLF viability and proliferation were evaluated using the trypan blue staining and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, respectively. RESULTS: Sant7, at a concentration of 1 microg/mL, was capable of significantly inhibiting HLF proliferation and MAPK phosphorylation induced by cell exposure to IL-6 (100 ng/mL) or TGF-beta1 (10 ng/mL), whose actions were more evident in fibrotic cells. CONCLUSIONS: These findings suggest that, in HLFs derived from patients with ILDs, the proliferative mechanisms activated by TGF-beta1 are at least in part mediated by an increased release of IL-6, leading to phosphorylation-dependent MAPK activation. Such preliminary findings may thus open new therapeutic perspectives for fibrogenic ILDs, based on inhibition of signal transduction pathways stimulated by the IL-6 receptor.

Inflammation, oxidative stress and apoptosis, which are involved in chronic obstructive pulmonary disease (COPD) pathogenesis, may activate the p38 subgroup of mitogen-activated protein kinases (MAPKs). Therefore, the aim of the present study was to evaluate the expression of the phosphorylated, active form of p38 MAPK (phospho-p38) in the lungs of COPD patients. Surgical specimens were obtained from 18 smokers with COPD at different stages of disease severity, plus nine smoking and eight nonsmoking subjects with normal lung function. Phospho-p38+ cells were quantified by immunohistochemistry in both alveolar spaces and alveolar walls. Moreover, a Western blot analysis of phospho-p38 and total p38alpha isoform expressed by alveolar macrophages was performed. Phospho-p38+ alveolar macrophages and phospho-p38+ cells in alveolar walls were increased in patients with severe and mild/moderate COPD, compared with smoking and nonsmoking controls. Moreover, they were inversely correlated to values of forced expiratory volume in one second (FEV(1)) and FEV(1)/forced vital capacity. Western blot analysis showed that phosphorylated p38, but not the total p38alpha isoform, was specifically increased in alveolar macrophages from COPD patients. Activation of the p38 mitogen-activated protein kinase pathway appears to be involved in the pathogenesis of chronic obstructive pulmonary disease. The present findings suggest that this protein may be a suitable pharmacological target for therapeutic intervention.

1 Department of Experimental and Clinical Medicine, Chair of Pharmacology, Clinical Pharmacology and Pharmacovigilance Unit, University ‘Magna Graecia’ of Catanzaro, Regional Pharmacovigilance Center, ‘Mater Domini’ University Hospital, Catanzaro, Italy 2 Department of Experimental and Clinical Medicine, Chair of Respiratory Diseases, University ‘Magna Graecia’ of Catanzaro, Regional Pharmacovigilance Center, ‘Mater Domini’ University Hospital, Catanzaro, Italy