1. A Benzodiazepine Ligand with Improved GABA A Receptor α 5-Subunit Selectivity Driven by Interactions with Loop C.
- Author
-
Simeone X, Koniuszewski F, Müllegger M, Smetka A, Steudle F, Puthenkalam R, Ernst M, and Scholze P
- Subjects
- Animals, Benzodiazepines chemistry, Benzodiazepines pharmacology, Dose-Response Relationship, Drug, Female, Flunitrazepam chemistry, Flunitrazepam metabolism, Flunitrazepam pharmacology, GABA Modulators chemistry, GABA Modulators pharmacology, HEK293 Cells, Humans, Ligands, Molecular Docking Simulation methods, Protein Binding physiology, Protein Structure, Secondary, Rats, Receptors, GABA-A chemistry, Xenopus laevis, Benzodiazepines metabolism, GABA Modulators metabolism, Receptors, GABA-A metabolism
- Abstract
The family of GABA
A receptors is an important drug target group in the treatment of sleep disorders, anxiety, epileptic seizures, and many others. The most frequent GABAA receptor subtype is composed of two α -, two β -, and one γ 2-subunit, whereas the nature of the α -subunit critically determines the properties of the benzodiazepine binding site of those receptors. Nearly all of the clinically relevant drugs target all GABAA receptor subtypes equally. In the past years, however, drug development research has focused on studying α 5-containing GABAA receptors. Beyond the central nervous system, α 5-containing GABAA receptors in airway smooth muscles are considered as an emerging target for bronchial asthma. Here, we investigated a novel compound derived from the previously described imidazobenzodiazepine SH-053-2'F-R-CH3 (SH53d-ester). Although SH53d-ester is only moderately selective for α 5-subunit-containing GABAA receptors, the derivative SH53d-acid shows superior (>40-fold) affinity selectivity and is a positive modulator. Using two-electrode voltage clamp electrophysiology in Xenopus laevis oocytes and radioligand displacement assays with human embryonic kidney 293 cells, we demonstrated that an acid group as substituent on the imidazobenzodiazepine scaffold leads to large improvements of functional and binding selectivity for α 5 β 3 γ 2 over other α x β 3 γ 2 GABAA receptors. Atom level structural studies provide hypotheses for the improved affinity to this receptor subtype. Mutational analysis confirmed the hypotheses, indicating that loop C of the GABAA receptor α -subunit is the dominant molecular determinant of drug selectivity. Thus, we characterize a promising novel α 5-subunit-selective drug candidate. SIGNIFICANCE STATEMENT: In the current study we present the detailed pharmacological characterization of a novel compound derived from the previously described imidazobenzodiazepine SH-053-2'F-R-CH3. We describe its superior (>40-fold) affinity selectivity for α 5-containing GABAA receptors and show atom-level structure predictions to provide hypotheses for the improved affinity to this receptor subtype. Mutational analysis confirmed the hypotheses, indicating that loop C of the GABAA receptor α -subunit is the dominant molecular determinant of drug selectivity., (Copyright © 2020 by The Author(s).)- Published
- 2021
- Full Text
- View/download PDF