580. Prevention of Airway Inflammation by Simultaneous Inhibition of NFkB and STST6 Using Chimeric Decoy Oligonucleotides in a Mouse Model of Asthma

Abstract

Allergic asthma is a common inflammatory disease of the airways characterized by reversible airflow obstruction and airway hyperresponsiveness (AHR) to bronchoconstrictor stimuli. The pathophysiology in asthma mainly occurs as a consequence of overexpression of the TH2 cytokines. These inflammatory factors are tightly regulated by a transcriptional network system. Nuclear factor kappaB (NFkB) and STAT6 are well known to regulate a set of gene associated with inflammatory and immune responses, and is thought to play an important role in the induction of allergic asthma. Therefore, we focused on the simultaneous inhibition of these important transcription factors using a decoy strategy to develop a novel therapeutic approach for treating asthma. For increased efficacy of decoy oligodeoxynucleotides (ODN) in vivo, we employed chimeric decoy ODN containing consensus sequences of both NFkB and STAT6 binding sites. In addition, two strands of decoy ODN were combined by the chemical spacer to increase its resistance to endonuclease for intratracheal administration. The therapeutic effect of chimeric decoy ODN was investigated using OVA-induced experimental asthma in mice. Mice were sensitized by intraperitoneal injections of 20 ug of ovalbumin (OVA) and 2 mg aluminum hydroxide constituted in 0.1 ml of saline on days 0 and 14. On days 21, 22 and 23, mice were challenged with 1% OVA aerosol for 20 min. At 24 hours after the last challenge, AHR was measured by methacholine-induced airflow obstruction. Intratracheal administration of decoy ODN was performed in OVA-sensitized mice at 3 days before the first challenge with aerosolized OVA. FITC-labeled chimeric decoy ODN could be detected in macrophages and monocytes migrating into the lung and airway, and NFkB and STAT6 activity were simultaneously inhibited by chimeric decoy ODN. Twenty-four hours after the last OVA challenge, treatment with chimeric or single transfection of NFkB decoy ODN was protected from methacholine-induced AHR, while mice treated with scrambled decoy ODN or saline developed a significantly increase in airway reactivity to methacholine. Importantly, this inhibitory effect of chimeric decoy ODN on airway hyperresponsiveness was significantly greater than that of NFkB decoy ODN. Treatment with chimeric decoy ODN markedly suppressed airway inflammation after OVA sensitization and challenge as compared with control and scrambled decoy ODN treatment. Inflammatory infiltrate, such as macrophage, was significantly inhibited by chimeric decoy ODN through suppression of ICAM-1 and eotaxin expression. In addition, secretion of Th2 cytokines including IL-4, IL-5 and IL-13 in BALF, and histamine in the whole lung were reduced by chimeric decoy ODN. Furthermore, a significant reduction of mucin secretion was observed by chimeric decoy ODN treatment accompanied by a suppression of MUC5AC gene expression. However, intratracheal administration of chimeric decoy ODN did not affected IgE synthesis. The present study provides a novel strategy for treating bronchial asthma by the simultaneous inhibition of both NFkB and STAT6 using chimeric decoy ODN. Further modification of chimeric decoy ODN would be useful to treat asthma as a decoy-based therapy.