ic-87114 and Asthma

Bronchial epithelial cells are front sentinels eliciting innate and adaptive immunity to respiratory viral pathogens. Recognition of viral double-stranded RNA induces antiviral interferon (IFN) responses in bronchial epithelial cells. Co-inhibitory molecules programmed cell death 1 ligand 1 (PD-L1) and ligand 2 (PD-L2) were also induced on bronchial epithelial cells, which bind programmed cell death 1 on T cell and inhibit the function of virus-specific cytotoxic T lymphocyte. A previous study showed that antiviral type I IFN increased PD-L1 and PD-L2 expression in cultured melanoma cells. However, it remains unknown whether antiviral IFNs affect PD-L1 and PD-L2 expression in bronchial epithelial cells. In addition, we previously reported that inhibition of PI3Kδ signaling enhanced antiviral IFN responses in human primary bronchial epithelial cells (PBECs). Here we assessed the effect of exogenous IFNs or a selective PI3Kδ inhibitor IC87114 on PD-L1 and PD-L2 in PBECs stimulated with a synthetic double-stranded RNA poly I:C or human metapneumovirus. Treatment with IFNβ or IFNλ increased PD-L1 and PD-L2, and IFNβ or IFNλ treatment plus poly I:C further increased both expressions. Treatment with IC87114 or transfection with siRNA targeting PI3K p110δ enhanced poly I:C-induced gene and protein expression of PD-L2, whereas IC87114 suppressed poly I:C-induced PD-L1. IC87114 enhanced poly I:C-induced gene expression of IFNβ, IFNλ, and IFN-regulated genes

Topics: Adenine; Asthma; B7-H1 Antigen; Bronchi; Cells, Cultured; Epithelial Cells; Gene Expression Regulation; Humans; Interferon Regulatory Factor-3; Interferons; Metapneumovirus; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Poly I-C; Programmed Cell Death 1 Ligand 2 Protein; Protein Serine-Threonine Kinases; Quinazolines

Hyperactivation of phosphoinositol 3-kinase (PI3K) has been suggested to be a potential mechanism for endoplasmic reticulum (ER) stress-enhanced airway hyperresponsiveness, and PI3K inhibitors have been examined as asthma therapeutics. However, the regulatory mechanism linking PI3K to ER stress and related pathological signals in asthma have not been defined. To elucidate these pathogenic pathways, we investigated the influence of a selective PI3Kδ inhibitor, IC87114, on airway inflammation in an ovalbumin/lipopolysaccharide (OVA/LPS)-induced asthma model. In OVA/LPS-induced asthmatic mice, the activity of PI3K, downstream phosphorylation of AKT and activation of nuclear factor-κB (NF-κB) were all significantly elevated; these effects were reversed by IC87114. IC87114 treatment also reduced the OVA/LPS-induced ER stress response by enhancing the intra-ER oxidative folding status through suppression of protein disulfide isomerase activity, ER-associated reactive oxygen species (ROS) accumulation and NOX4 activity. Furthermore, inositol-requiring enzyme-1α (IRE1α)-dependent degradation (RIDD) of IRE1α was reduced by IC87114, resulting in a decreased release of proinflammatory cytokines from bronchial epithelial cells. These results suggest that PI3Kδ may induce severe airway inflammation and hyperresponsiveness by activating NF-κB signaling through ER-associated ROS and RIDD-RIG-I activation. The PI3Kδ inhibitor IC87114 is a potential therapeutic agent against neutrophil-dominant asthma.

Topics: Adenine; Animals; Asthma; Disease Models, Animal; Endoplasmic Reticulum Stress; Lipid Peroxidation; Lipopolysaccharides; Membrane Proteins; Mice; Nerve Tissue Proteins; NF-kappa B; Ovalbumin; Oxidation-Reduction; Oxidative Stress; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Quinazolines; Reactive Oxygen Species; Receptors, Cell Surface; Signal Transduction

Transforming growth factor (TGF) β1 increases pro-inflammatory cytokines and contractile protein expression by human airway smooth muscle (ASM) cells, which could augment airway inflammation and hyperresponsiveness. Phosphoinositide 3' kinase (PI3K) is one of the signaling pathways implicated in TGFβ1 stimulation, and may be altered in asthmatic airways. This study compared the expression of PI3K isoforms by ASM cells from donors with asthma (A), chronic obstructive pulmonary disease (COPD), or neither disease (NA), and investigated the role of PI3K isoforms in the production of TGFβ1 induced pro-inflammatory cytokine and contractile proteins in ASM cells. A cells expressed higher basal levels of p110δ mRNA compared to NA and COPD cells; however COPD cells produced more p110δ protein. TGFβ1 increased 110δ mRNA expression to the same extent in the three groups. Neither the p110δ inhibitor IC87114 (1, 10, 30 µM), the p110β inhibitor TGX221 (0.1, 1, 10 µM) nor the PI3K pan inhibitor LY294002 (3, 10 µM) had any effect on basal IL-6, calponin or smooth muscle α-actin (α-SMA) expression. However, TGFβ1 increased calponin and α-SMA expression was inhibited by IC87114 and LY294002 in all three groups. IC87114, TGX221, and LY294002 reduced TGFβ1 induced IL-6 release in a dose related manner in all groups of ASM cells. PI3K p110δ is important for TGFβ1 induced production of the contractile proteins calponin and α-SMA and the proinflammatory cytokine IL-6 in ASM cells, and may therefore be relevant as a potential therapeutic target to treat both inflammation and airway remodeling.

Topics: Adenine; Adult; Aged; Aged, 80 and over; Asthma; Benzamides; Cells, Cultured; Chromones; Class I Phosphatidylinositol 3-Kinases; Contractile Proteins; Dioxoles; Female; Gene Expression Regulation; Humans; Interleukin-6; Lung; Male; Middle Aged; Morpholines; Muscle, Smooth; Phosphatidylinositol 3-Kinases; Pulmonary Disease, Chronic Obstructive; Quinazolines; Tissue Donors; Transforming Growth Factor beta1

Patients with chronic obstructive pulmonary disease (COPD) show a poor response to corticosteroids, which has been linked to oxidative stress. Here we show that the long-acting β(2) -agonist formoterol (FM) reversed corticosteroid insensitivity under oxidative stress via inhibition of phosphoinositide-3-kinase (PI3K) signalling.. Responsiveness to corticosteroids dexamethasone (Dex), budesonide (Bud) and fluticasone propionate (FP) was determined, as IC(50) values on TNF-α-induced interleukin 8 release, in U937 monocytic cell line treated with hydrogen peroxide (H(2) O(2) ) or peripheral blood mononuclear cells (PBMCs) from patients with COPD or severe asthma.. PBMCs from severe asthma and COPD were less sensitive to Dex compared with those from healthy subjects. Both FM (10(-9)  M) and salmeterol (SM, 10(-8)  M) reversed Dex insensitivity in severe asthma, but only FM restored Dex sensitivity in COPD. Although H(2) O(2) exposure decreased steroid sensitivity in U937 cells, FM restored responsiveness to Bud and FP while the effects of SM were weaker. Additionally, FM, but not SM, partially inhibited H(2) O(2) -induced PI3Kδ-dependent (PKB) phosphorylation. H(2) O(2) decreased SM-induced cAMP production in U937 cells, but did not significantly affect the response to FM. The reduction of SM effects by H(2) O(2) was reversed by pretreatment with LY294002, a PI3K inhibitor, or IC87114, a PI3Kδ inhibitor.. FM reversed oxidative stress-induced corticosteroid insensitivity and decreased β(2) adrenoceptor-dependent cAMP production via inhibition of PI3Kδ signalling. FM will be more effective than SM, when combined with corticosteroids, for the treatment of respiratory diseases under conditions of high oxidative stress, such as in COPD.

Topics: 1-Phosphatidylinositol 4-Kinase; Adenine; Adrenal Cortex Hormones; Adrenergic beta-2 Receptor Agonists; Adult; Aged; Albuterol; Androstadienes; Asthma; Budesonide; Chromones; Dexamethasone; Drug Resistance; Ethanolamines; Female; Fluticasone; Formoterol Fumarate; Humans; Hydrogen Peroxide; Interleukin-8; Leukocytes, Mononuclear; Male; Middle Aged; Morpholines; Oxidative Stress; Pulmonary Disease, Chronic Obstructive; Quinazolines; Receptors, Adrenergic, beta-2; Salmeterol Xinafoate; U937 Cells; Young Adult

Trafficking and recruitment of eosinophils during allergic airway inflammation is mediated by the phosphatidylinositol 3-kinase (PI3K) family of signaling molecules. The role played by the p110δ subunit of PI3K (PI3K p110δ) in regulating eosinophil trafficking and recruitment was investigated using a selective pharmacological inhibitor (IC87114). Treatment with the PI3K p110δ inhibitor significantly reduced murine bone marrow-derived eosinophil (BM-Eos) adhesion to VCAM-1 as well as ICAM-1 and inhibited activation-induced changes in cell morphology associated with reduced Mac-1 expression and aberrant cell surface localization/distribution of Mac-1 and α4. Infused BM-Eos demonstrated significantly decreased rolling and adhesion in inflamed cremaster muscle microvessels of mice treated with IC87114 compared with vehicle-treated mice. Furthermore, inhibition of PI3K p110δ significantly attenuated eotaxin-1-induced BM-Eos migration and prevented eotaxin-1-induced changes in the cytoskeleton and cell morphology. Knockdown of PI3K p110δ with siRNA in BM-Eos resulted in reduced rolling, adhesion, and migration, as well as inhibition of activation-induced changes in cell morphology, validating its role in regulating trafficking and migration. Finally, in a mouse model of cockroach antigen-induced allergic airway inflammation, oral administration of the PI3K p110δ inhibitor significantly inhibited airway eosinophil recruitment, resulting in attenuation of airway hyperresponsiveness in response to methacholine, reduced mucus secretion, and expression of proinflammatory molecules (found in inflammatory zone-1 and intelectin-1). Overall, these findings indicate the important role played by PI3K p110δ in mediating BM-Eos trafficking and migration by regulating adhesion molecule expression and localization/distribution as well as promoting changes in cell morphology that favor recruitment during inflammation.

Topics: Adenine; Animals; Asthma; Bone Marrow Cells; Cell Adhesion; Cell Movement; Chemokine CCL11; Class I Phosphatidylinositol 3-Kinases; Eosinophilia; Eosinophils; Hypersensitivity; Intercellular Adhesion Molecule-1; Macrophage-1 Antigen; Methacholine Chloride; Mice; Mice, Inbred BALB C; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Quinazolines; Respiratory System; RNA Interference; RNA, Small Interfering; Vascular Cell Adhesion Molecule-1

Phosphoinositide 3-kinases (PI3Ks) contribute to the pathogenesis of asthma by regulating the activation of inflammatory mediators, inflammatory cell recruitment and immune cell function. Recent findings have indicated that PI3Ks also regulate the expression of interleukin (IL)-17, which has been recognised as an important cytokine involved in airway inflammation. In the present study, we investigated a role of PI3Kδ in the regulation of IL-17 expression in allergic airway disease using a murine model of asthma. After ovalbumin inhalation, administration of a selective p110δ inhibitor, IC87114, significantly attenuated airway infiltration of total cells, lymphocytes, neutrophils and eosinophils, as well as airway hyperresponsiveness, and attenuated the increase in IL-17 protein and mRNA expression. Moreover, IC87114 reduced levels of IL-4, -5 and -13, expression of keratinocyte chemoattractant protein and mRNA, and nuclear factor (NF)-κB activity. In addition, a NF-κB inhibitor, BAY 11-7085 substantially reduced the increase in IL-17 protein levels. Our results also showed that inhibition of IL-17 activity with an anti-IL-17 antibody remarkably reduced airway inflammation and hyperresponsiveness. These findings suggest that inhibition of the p110δ signalling pathway suppresses IL-17 expression through regulation of NF-κB activity and, thus, has therapeutic potential in asthma.

Topics: Adenine; Animals; Asthma; Bronchoalveolar Lavage Fluid; Chemotactic Factors; Eosinophils; Female; Interleukin-17; Interleukin-4; Interleukin-5; Lung; Lymphocytes; Mice; Mice, Inbred C57BL; Neutrophils; NF-kappa B; Nitriles; Phosphoinositide-3 Kinase Inhibitors; Quinazolines; Sulfones

Hypoxia-inducible factor-1α (HIF-1α) plays a critical role in immune and inflammatory responses. One of the HIF-1α target genes is vascular endothelial growth factor (VEGF), which is a potent stimulator of inflammation, airway remodeling, and physiologic dysregulation in allergic airway diseases. Using OVA-treated mice and murine tracheal epithelial cells, the signaling networks involved in HIF-1α activation and the role of HIF-1α in the pathogenesis of allergic airway disease were investigated. Transfection of airway epithelial cells with HIF-1α siRNA suppressed VEGF expression. In addition, the increased levels of HIF-1α and VEGF in lung tissues after OVA inhalation were substantially decreased by an HIF-1α inhibitor, 2-methoxyestradiol. Our data also show that the increased numbers of inflammatory cells, increased airway hyperresponsiveness, levels of IL-4, IL-5, IL-13, and vascular permeability in the lungs after OVA inhalation were significantly reduced by 2-methoxyestradiol or a VEGF inhibitor, CBO-P11. Moreover, we found that inhibition of the PI3K p110δ isoform (PI3K-δ) or HIF-1α reduced OVA-induced HIF-1α activation in airway epithelial cells. These findings indicate that HIF-1α inhibition may attenuate antigen-induced airway inflammation and hyperresponsiveness through the modulation of vascular leakage mediated by VEGF, and that PI3K-δ signaling may be involved in the allergen-induced HIF-1α activation.

Topics: 2-Methoxyestradiol; Adenine; Airway Remodeling; Animals; Asthma; Bronchoalveolar Lavage Fluid; Endothelial Growth Factors; Epithelial Cells; Estradiol; Female; Histocytochemistry; Hypoxia-Inducible Factor 1, alpha Subunit; Mice; Mice, Inbred C57BL; Ovalbumin; Peptides, Cyclic; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Quinazolines; Respiratory Function Tests; Reverse Transcriptase Polymerase Chain Reaction; RNA; RNA, Small Interfering; Signal Transduction; Specific Pathogen-Free Organisms; Vascular Endothelial Growth Factor A

P110delta phosphoinositide 3-kinase (PI3K) plays a pivotal role in the recruitment and activation of certain inflammatory cells. Recent findings revealed that the activity of p110delta also contributes to allergen-IgE-induced mast cell activation and vascular permeability. We investigated the role of p110delta in allergic airway inflammation and hyperresponsiveness using IC87114, a selective p110delta inhibitor, in a mouse asthma model. BALB/c mice were sensitized with OVA and, upon OVA aerosol challenge, developed airway eosinophilia, mucus hypersecretion, elevation in cytokine and chemokine levels, up-regulation of ICAM-1 and VCAM-1 expression, and airway hyperresponsiveness. Intratracheal administration of IC87114 significantly (P<0.05) attenuated OVA-induced influx into lungs of total leukocytes, eosinophils, neutrophils, and lymphocytes, as well as levels of IL-4, IL-5, IL-13, and RANTES in a dose-dependent manner. IC87114 also significantly (P<0.05) reduced the serum levels of total IgE and OVA-specific IgE and LTC(4) release into the airspace. Histological studies show that IC87114 inhibited OVA-induced lung tissue eosinophilia, airway mucus production, and inflammation score. In addition, IC87114 significantly (P<0.05) suppressed OVA-induced airway hyperresponsiveness to inhaled methacholine. Western blot analyses of whole lung tissue lysates shows that IC87114 markedly attenuated the OVA-induced increase in expression of IL-4, IL-5, IL-13, ICAM-1, VCAM-1, RANTES, and eotaxin. Furthermore, IC87114 treatment markedly attenuated OVA-induced serine phosphorylation of Akt, a downstream effector of PI3K signaling. Taken together, our findings implicate that inhibition of p110delta signaling pathway may have therapeutic potential for the treatment of allergic airway inflammation.

Topics: Adenine; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Adhesion Molecules; Chemokines; Chemotaxis, Leukocyte; Class I Phosphatidylinositol 3-Kinases; Cytokines; Eosinophilia; Female; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Models, Animal; Mucus; Ovalbumin; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Protein Processing, Post-Translational; Proto-Oncogene Proteins c-akt; Quinazolines

Bronchial asthma is characterized by inflammation of the airways, which is usually accompanied by increased vascular permeability, resulting in plasma exudation. Vascular endothelial growth factor (VEGF) has been implicated in contributing to asthmatic tissue edema through its effect on vascular permeability. Many cellular responses of VEGF are regulated by the lipid products of phosphoinositide 3-kinase (PI3K). However, the effect of PI3K catalytic subunit p110delta on VEGF-mediated signaling is unknown. Recently, an isoform-specific small molecule inhibitor, IC87114, which is selective for p110delta catalytic activity, has been identified.. We have sought to investigate the role of PI3K-delta, more specifically in the increase of vascular permeability.. Female BALB/c mice were sensitized and challenged with ovalbumin. We have investigated the effect of IC87114 on airway inflammation, T(H)2 cytokines expression, airway hyperresponsiveness, plasma extravasation, hypoxia-inducible factor 1alpha expression, and VEGF expression in a murine model of asthma.. Our current study has revealed that IC87114 reduces antigen-induced airway infiltration of inflammatory cells, secretion of T(H)2 cytokines in lungs, airway hyperresponsiveness, and vascular permeability. Moreover, we have found that inhibition of p110delta reduces ovalbumin-induced upregulation of VEGF level.. These results suggest that PI3K-delta inhibitor attenuates antigen-induced airway inflammation and hyperresponsiveness by preventing vascular leakage in mice.. These findings provide a crucial molecular mechanism for the potential role of PI3K-delta in asthma and other airway inflammatory disorders.

Topics: Adenine; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capillary Permeability; Cinnamates; Class I Phosphatidylinositol 3-Kinases; Cytokines; Disease Models, Animal; Enzyme Inhibitors; Female; Hypoxia-Inducible Factor 1, alpha Subunit; Indoles; Leukocyte Count; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Phosphoinositide-3 Kinase Inhibitors; Pneumonia; Quinazolines; Receptors, Vascular Endothelial Growth Factor; Vascular Endothelial Growth Factor A