Characterization of specific drug receptors with fluorescent ligands

Abstract

Conjugates of fluorescent labels with specific, selective, and high affinity ligands for receptors have been synthesized and used to directly measure binding to receptors. In the examples, fluorescein conjugates of the high-affinity benzodiazepine receptor ligands Ro 15-1788 and Ro 7-1986 were synthesized. The binding of these fluorescent ligands (BD 621, BD 623 and BD 607) to benzodiazepine receptors was characterized by direct fluorescence measurement. Both the equilibrium dissociation constants (K.sub.D) of BD 621 and BD 607 and the maximum number of binding sites (B.sub.max) estimated by fluorescence monitoring were consistent with values obtained by using radioligand binding techniques. The binding of BD 621 and BD 607 assessed by fluorescence measurement was reversible, abolished by photoaffinity labeling with Ro 15-4513, and unaffected by a variety of substances that do not bind to benzodiazepine receptors. The potencies of chemically diverse benzodiazepine receptor compounds to inhibit fluorescent ligand binding were highly correlated, with potencies obtained from radioligand binding techniques. These findings demonstrate that direct fluorescence measurement techniques can be used to quantitate ligand-receptor interactions.