New Imidazodiazepine Analogue, 5-(8-Bromo-6-(pyridin-2-yl)-4H-benzo[f]imidazo[1,5-a][1,4]diazepin-3-yl)oxazole, Provides a Simplified Synthetic Scheme, High Oral Plasma and Brain Exposures, and Produces Antiseizure Efficacy in Mice, and Antiepileptogenic Activity in Neural Networks in Brain Slices from a Patient with Mesial Temporal Lobe Epilepsy.

KRM-II-81 (1) is an imidazodiazepine GABAA receptor (GABAAR) potentiator with broad antiseizure efficacy and a low sedative burden. A brominated analogue, DS-II-73 (5), was synthesized and pharmacologically characterized as a potential backup compound as KRM-II-81 moves forward into development. The synthesis from 2-amino-5-bromophenyl)(pyridin-2yl)methanone (6) was processed in five steps with an overall yield of 38% and without the need for a palladium catalyst. GABAAR binding occurred with a Ki of 150 nM, and only 3 of 41 screened binding sites produced inhibition ≥50% at 10 µM, and the potency to induce cytotoxicity was ≥240 mM. DS-II-73 was selective for α2/3/5- over that of α1-containing GABAARs. Oral exposure of plasma and brain of rats was more than sufficient to functionally impact GABAARs. Tonic convulsions in mice and lethality induced by pentylenetetrazol were suppressed by DS-II-73 after oral administration and latencies to clonic and tonic seizures were prolonged. Cortical slice preparations from a patient with pharmacoresistant epilepsy (mesial temporal lobe) showed decreases in the frequency of local field potentials by DS-II-73. As with KRM-II-81, the motor-impairing effects of DS-II-73 were low compared to diazepam. Molecular docking studies of DS-II-73 with the α1ß3γ2L-configured GABAAR showed low interaction with α1His102 that is suggested as a potential molecular mechanism for its low sedative side effects. These findings support the viability of DS-II-73 as a backup molecule for its ethynyl analogue, KRM-II-81, with the human tissue data providing translational credibility.

KRM-II-81 suppresses epileptifom activity across the neural network of cortical tissue from a patient with pharmacoresistant epilepsy.

A clinical case of a 19-year-old male patient with pharmacoresistant seizures occurring following parieto-occipital tumor-resection at age 6 is described. Seizure surgery work-up included prolonged video EEG monitoring and head CT without contrast. Seizure focus was localized to the left temporal lobe, and we felt that the patient was an excellent candidate for seizure surgery. The patient underwent a left frontotemporal craniotomy for removal of the seizure focus with intraoperative electrocorticography (ECoG) conducted pre and post resection. ECoG recordings pre- and post-resection confirmed resolution of seizure generation. Imaging obtained immediately postoperatively showed complete resection of the residual tumor with no evidence of recurrence in follow-ups. A year after the surgery the patient is seizure-free but remains on seizure medication. With the patient's consent the excised epileptogenic tissue was used for ex-vivo research studies. The microelectrode recordings confirmed epileptiform activity in the excised tissue incubated in excitatory artificial cerebrospinal fluid. The epileptiform activity in the epileptogenic tissue was suppressed by addition of KRM-II-81, a novel α2/3 subtype preferring GABAA receptor (GABAAR) potentiator with previously demonstrated antiepileptic efficacy in multiple animal models of epilepsy and with reduced potential for CNS side-effects compared to classical benzodiazepine GABAAR potentiators. These findings support the proposition that KRM-II-81 might reduce seizure burden in pharmacoresistant patients.