Ineffective mucociliary clearance (MCC) is a common pathophysiologic process contributing to chronic rhinosinusitis (CRS) - a highly prevalent disease with substantial morbidity. Evidence from our laboratory has demonstrated that brief exposures to lipopolysaccharide (LPS) from gram-negative bacteria lead to dysfunctional MCC by decreasing anion transport through CFTR apical Cl- channels in humans and other mammalian species. CFTR inhibition in this setting is caused by TLR4-mediated generation of reactive oxygen species (ROS), but independent of NFkB-derived inflammation. Furthermore, our group has confirmed the presence of acquired CFTR dysfunction in human sinuses, and has demonstrated that CFTR potentiators can stimulate Cl- secretion when partial CFTR dysfunction is present in multiple in vitro and preclinical models. Our central hypotheses are that LPS-induced acquired CFTR deficiency, 1) contributes substantially to the pathogenesis of CRS, and 2) can be treated with Ivacaftor, a CFTR potentiator developed for CF therapy.
Specific Aim 1 will investigate the mechanistic basis of LPS-mediated CFTR dysfunction in sinonasal epithelium by 1) examining the oxidant-dependent inhibition of CFTR via AMP-dependent kinase (an inhibitor of PKA-dependent phosphorylation of the CFTR regulatory domain), 2) assessing the impact of ROS (superoxide, hydrogen peroxide) on CFTR function (patch clamp analysis) and structure (mass spectrometry), and 3) measuring the effects of longer exposures to LPS on CFTR expression, maturational processing, and recycling.
Aim 2 will identify the efficiency of Ivacaftor in improving CFTR function in a pre-clinical rabbit model of acquired CFTR deficiency by 1) developing normative data for LPS-exposed rabbit maxillary sinus CFTR dysfunction, 2) assessing the effects of Ivacaftor on CFTR-related endpoints, and 3) evaluating Ivacaftor as therapy for Pseudomonas aeruginosa rabbit maxillary sinusitis.
Aim 3 will conduct a clinical study using Ivacaftor in CRS patients by 1) correlating the novel ?endoscopically-directed sinus potential difference? assay to validated measures of CRS disease severity and 2) performing a pilot clinical trial using Ivacaftor for CRS patients with refractory gram-negative bacterial CRS. The current proposal will help clarify mechanisms responsible for sinusitis pathogenesis, but also translates our laboratory findings to human subjects by providing a clinical trial using Ivacaftor for therapy of sinusitis. We believe our application will answer fundamental questions regarding pathomechanisms underlying CRS and establish the foundation for a new therapeutic approach to a serious and debilitating chronic disease.
Evidence from our laboratory has demonstrated that brief exposures to lipopolysaccharide ? a component of gram-negative bacterial cell walls - lead to dysfunctional mucus clearance by decreasing transport of chloride (Cl-) ions through the cystic fibrosis transmembrane conductance regulator (CFTR) present on the surface of airway cells. Furthermore, our group has confirmed the existence of acquired CFTR dysfunction in the sinuses of human subjects with chronic rhinosinusitis, and that the drug Ivacaftor (developed for therapy of cystic fibrosis) overcomes acquired Cl- secretory defects present in multiple in vitro and in vivo model systems. The current proposal will help clarify mechanisms responsible for sinusitis pathogenesis, and also translate our laboratory findings to human subjects by providing a clinical trial using Ivacaftor for therapy of non-CF sinusitis.
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