Trimetoquinol (TMQ) is a novel tetrahydroisoquinoline drug which interacts stereoselectively with beta-adrenergic (beta1-, beta2- and beta3-subtypes) and thromboxane A2 (TXA2) [alpha- and tau- subtypes] receptors. Our hypothesis is that TMQ binds in a three-point attachment to both of these receptor systems, and interactions with receptor domains include the catechol, amino and trimethoxybenzyl moieties. For the beta- adrenoceptor, we believe that TMQ binds with serines (Ser) 204 and 207 (catechol group), aspartic acid (Asp) 113 (amino group), and a third unknown region (trimethoxybenzyl group). The primary objective of this proposal is to synthesize and study highly selective photoaffinity and affinity labels of TMQ as probes of TXA2 and beta-adrenergic receptors, and to utilize mutant beta-adrenoceptor systems for defining specific receptor binding domains. Because of the major differences in optical isomers of TMQ analogs (TMQ and 8-fluoro TMQ) and high potency of the 3'-iodo and 3',5'-diiodo TMQ on TXA2 and beta-adrenergic receptors, we have designed optically active radiolabeled photoaffinity and affinity labels for probing these two distinct receptor systems. A second part of the proposal will be to synthesize a series of catechol mimetic analogs of TMQ that will investigate the importance of hydrogen bonding to receptors, and should provide more selective (beta2 versus beta1) and longer-acting (resistant to COMT metabolism) drugs than the parent molecule as beta-agonists (bronchial relaxants) and TXA2 antagonists (antiaggregatory and hypotensive activities). Radiolabeled affinity compounds will be used for biochemical characterization of molecular mass and isoelectric points of receptor proteins, and of peptide mapping and amino acid sequencing of relevant peptides. We will also examine ligand binding domains for TMQ analogs using mutant beta-adrenoceptors, and define relationships between receptor occupancy and signal transduction for TMQ analogs using cells expressing unmutated beta-adrenoceptors. Newly synthesized drugs will be evaluated for receptor specific interactions in beta-adrenergic (beta1-, beta2- and beta3-) and TXA2 (smooth muscle and endothelial cells, platelets) tissues. The main purpose is to characterize specific receptor site interactions of tetrahydroisoquinolines with beta-adrenergic and TXA2-receptor systems. Our long-term goal is to separate the desired bronchial dilation from the undesired beta2-adrenergic effects on skeletal muscle (tremors) and to differentiate between the platelet aggregatory and vasoconstrictive effects of TXA2.
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