Synthesis and characterization of a radioiodinated photoaffinity probe for the alpha 1-adrenergic receptor.

Abstract

A radioiodinated aryl azide analog, 2-[4-(4-azido-3- iodobenzoyl ) piperazin -1-yl]-4-amino-6, 7- dimethoxyquinazoline [(125I] CP65 ,526), of the highly selective alpha 1-adrenergic antagonist prazosin was synthesized and characterized using rat hepatic plasma membranes. Prior to photolysis, this ligand bound with high affinity (Kd 0.3 nM), stereoselectively and in a saturable manner to sites with an alpha 1-adrenergic specificity. When membranes pretreated with [125I] CP65 ,526 were irradiated with ultraviolet light, the ligand incorporated irreversibly into the receptor-binding sites, also with typical alpha 1-adrenergic specificity. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of such labeled membranes followed by radioautography revealed major bands at Mr = 77,000, 68,000, and 59,000 daltons. Labeling of each of these bands was inhibitable by a variety of adrenergic ligands, stereoselectively and with a specificity typical of the alpha 1-adrenergic receptor. Smaller peptides with molecular weights of 42,000 and 31,000 daltons also displayed prazosin-inhibitable [125] CP65 ,526-binding. However, as the labeling of these protein species was not inhibitable by other adrenergic agonists or antagonists, they are unlikely to represent subunits of the receptor. Further evidence that [125I] CP65 ,526 incorporates covalently upon photolysis was the ability to specifically label immunoglobulin heavy and light chains of an antiserum that recognized both this ligand and the parent compound, prazosin. This new, radioiodinated, high-affinity probe should thus be uniquely valuable for the molecular characterization of the alpha 1-adrenergic receptor.

Alpha-1 Blockers. A New Generation of Antihypertensive Agents

“…Much work has gone into modifying the structure of the basic prazosin molecule to retain selective a-1 blockade but obtain a longer, more gradual action. Seidman and colleagues (84) found a photoaffinity a-1 selective probe to characterize the a-1 adrenoceptor, which demonstrated that the group in the N-4 position of the piperazine moiety was unnecessary for activity; similarly, Timmermans e t al. (85), reported that a piperidine derivative had potent a-1 inhibitory activity.…”

“…Although doxazosin is also found to have a longer half-life than prazosin, the benzodioxan moiety may not be as water-soluble as the tetrahydrofuran group in terazosin. There is a benzodioxan double-ring in the simple a-blocker piperoxan, but the exact identity of the substitute seems unimportant (84)(85)(86)(87).…”

Advances in the design and synthesis of prazosin derivatives over the last ten years

“…Iodoazidoaryl prazosin (IAAP), a photoactive analog of prazosin, has been previously reported to act as a multidrug resistance reversal agent by binding and inhibiting Pgp1 function [90]. As the synthesis of IAAP following the earlier route [91] proved to be dangerously unreliable, a convergent route involving direct addition of an acylated piperazine to 2-chloroquinazoline intermediate was established [92]. Erlotinib can also inhibit multidrug resistance by reversing ABC subfamily B member 1 (ABCB1; P-glycoprotein) and ABC subfamily G member 2 (ABCG2; breast cancer resistance protein/mitoxantrone resistance protein) functions in cancer cells through direct inhibition of the drug efflux function of ABCB1 and ABCG2 [93].…”