Airway inflammatory diseases such as asthma are characterized by inflammation, airway remodeling and hyperresponsiveness resulting in severe bronchoconstriction. Agonists of the beta adrenergic receptor (??AR) relax airway smooth muscle (ASM), bronchodilate, and hence are prominent asthma prophylaxis and rescue medicines. Recent studies have demonstrated multiple clinical problems associated with ??AR agonists including worsening of asthma symptoms, tachyphylaxis and asthma-associated mortality. Therefore, there is a clinical need to develop novel and improved ?2AR ligands. ?2ARs on ASM are coupled to Gs protein and relax ASM via a cAMP/protein kinase A mechanism. However, recent studies have demonstrated that ?2ARs bind to and activate additional signaling proteins (e.g. ?-arrestins) in target cells leading to untoward functional effects such as desensitization, proliferation, and altered gene expression. Molecular characterization of these regulatory events in airway cells is necessary for ?2AR. Receptor interaction with signaling proteins and thus activation is dependent on the receptor conformation which is based on the chemical structure of the ligand. Indeed, preliminary data suggest that agonist-induced conformational changes in ?2AR is distinct depending upon the structure of the ligand and the presence of interacting proteins such as Gs protein. Therefore, we hypothesize that the activation of multiple signaling pathways is dependent on the conformational heterogeneity of the ?2AR, which can be manipulated by ligands of differing chemical structures.
In Aim 1 we will employ advanced computational approaches including atomistic molecular dynamics simulations carried out at a superior spatial and temporal resolution in the presence of interacting proteins and ligands. Further, we will develop functional group affinity patterns, FragMaps, for different conformations of the receptor which will be used to screen for novel ligands. Select compounds will be tested for biological activity in Aim 2 using multiple experimental models including HEK293 cells expressing human ?2AR, human ASM cells, isolated airways and lung slices, and by determining intracellular signaling (cAMP generation, PKA activation, ERK phosphorylation) and functional effects (relaxation, proliferation). We will correlate the structural information obtained from Aim 1 with the signaling and functional findings from Aim 2 to identify novel ?2AR agonists. We anticipate that the ?2AR ligands will act in a biased fashion, differentially favoring certain G-protein dependent, or, ?-arrestin dependent processes, based on ligand structure and the receptor conformation established by that ligand. This information can be leveraged for the rational design of novel and improved bronchodilators for asthma.
Bronchodilator drugs targeting beta adrenergic receptors on airways have been used as rescue and preventive medications for asthma. However, this class of drugs has come under scrutiny recently due to their ineffectiveness in controlling asthma symptoms, and due to multiple side effects including asthma-associated death. Studies proposed in this application will identify and characterize new compounds that inhibit airway hyperresponsiveness in asthma paving the way for the development of a novel and improved class of bronchodilators.
| Pera, Tonio; Deshpande, Deepak A; Ippolito, Michael et al. (2018) Biased signaling of the proton-sensing receptor OGR1 by benzodiazepines. FASEB J 32:862-874 |
