Discovery of H-10 as a potent and safe GABAA receptor positive allosteric modulator: A novel benzothiazine-triazole lead compound with broad-spectrum anticonvulsant activity.

Abstract

Epilepsy is a severe chronic neurological disorder predominantly caused by aberrant neuronal firing within the central nervous system. Herein, we report the design, synthesis, and biological evaluation of a series of 6-alkoxy-2-(4H-1,2,4-triazol-4-yl)-4H-benzo[d][1,3]thiazine derivatives based on a novel benzothiazine-triazole scaffold. Whole-cell patch-clamp recordings were performed on HEK-293 cells stably expressing human α1β3γ2 GABAA receptors (GABAARs). Lead compound H-10 was identified as a micromolar-potent GABAAR positive allosteric modulator (PAM), producing a 16.24-fold enhancement of GABA-evoked currents at 10 μM, with apparent functional preference for α1-containing receptor isoforms. Microscale thermophoresis verified the micromolar-affinity interaction between H-10 and the GABRA1 subunit. In preclinical seizure models, H-10 exerted robust antiseizure protection in pentylenetetrazol-challenged mice with an ED50 of 8.86 mg/kg and a favorable protective index of 10.6. Mechanistic studies revealed that H-10 restores cerebral GABA homeostasis and modulates hippocampal GABAergic-related protein expression. Pharmacokinetic profiling demonstrated high oral bioavailability (93.9%) and prominent brain penetration (brain-to-plasma AUC ratio = 3.51). H-10 elicited dose-dependent seizure suppression and alleviated multi-organ pathological damage in both acute and chronic epileptic mouse models. Collectively, H-10 is an orally bioavailable, efficacious GABAAR-targeted PAM with broad-spectrum antiseizure activity, deserving further optimization for anticonvulsant drug discovery.