This proposal aims to provide preclinical data in support of a novel therapy for epilepsy in Tuberous sclerosis complex (TSC), a disorder that includes early life epilepsy, mental retardation and autism. We propose that bumetanide, an inhibitor of the chloride cotransporter NKCC1, will be effective for the prevention of seizures in TSC alone or in combination with a canonical GABA agonist phenobarbital or the recently approved anticonvulsant vigabatrin. Our recent publication shows that NKCC1 is overexpressed in human TSC tissue, as well as in another cause of refractory epilepsy, focal cortical dysplasia Type IIb (FCD IIb). Increased NKCC1 causes enhanced intracellular chloride, and this impairs inhibitory actions of GABA receptor activation by causing it to mediate depolarization and excitation. We have recently generated a novel Tsc1cc Nestin-rtTA+ tet-OP-cre+ mutant mouse model that is epileptic and shows NKCC1 overexpression and depolarizing GABA responses.
Aim 1 will use this mouse model to further text the efficacy of bumetanide alone or in combination with phenobarbital or vigabatrin in cortical slices in vitro and by treatment in vivo using EEG recordings.
Aim 2 will examine the effects of bumetanide alone and in combination with phenobarbital or vigabatrin on GABA responses in brain slices acutely prepared from human TSC and FCD IIb surgical biopsy tissue. The proposed work represents a new approach in studying the mechanisms of epileptogenesis in TSC and has the potential to generate an adjuvant novel mechanism-based anticonvulsant strategy for seizure control in TSC patients. It will also provide proof-of-principle for this mechanism, as well as preclinical efficacy in a TSC mouse model with clinically relevant outcomes. Finally our proposal includes target validation as well as mechanism in human tissue. These are critical elements of a path to first-in-human trials for this compound in TSC, and may also extend to FCD IIb.
Epilepsy is a common disorder affecting over 1% of the population, and over 30% of patients have treatment resistant epilepsy. Some genetic conditions have even higher incidences of epilepsy: Tuberous Sclerosis Complex (TSC) is a genetic condition with and incidence of over 90% of patients with epilepsy. In addition, TSC epilepsy can be resistant to conventional anti-seizure drugs, and seizures in children with TSC can be associated with high rates of later intellectual disability. In some cases, the only treatment is surgical removal of the TSC tubers that are thought to be the foci of seizures. Here we show that a novel target, the NKCC1 protein, is expressed at higher than normal levels in human TSC brain as well as in a mouse model of TSC. The NKCC1 protein may cause seizures as well as be responsible for the resistance of TSC seizures from conventional drugs. We propose to use a blocker of NKCC1 activity, bumetanide, as add-on therapy in combination with an approved antiepileptic drugs phenobarbital and vigabatrin, to improve seizure control in a mouse model of TSC but also directly in brain slices from human TSC as well as surgical samples that are removed during surgery. We will also study tissue from patients with Focal Cortical Dysplasia Type IIb, as we have recently shown similar dysregulation of NKCC1 in these cases. As bumetanide is already FDA approved as a diuretic, if successful, the present proposal would provide important preclinical evidence and human target validation to support an early Phase I/II trial of bumetanide as an add-on therapy with conventional GABA agonist anticonvulsants like phenobarbital or other newer agents like vigabatrin for seizure control in TSC patients. Importantly, as a number of features of TSC patients are also seen in non-TSC patients with severe refractory epilepsy, such as Focal Cortical Dysplasia Type IIb, and these results may have wider application to more general cases of severe epilepsy. This grant application addresses several components of the Epilepsy Research Benchmarks (www.ninds.nih.gov/research/epilepsyweb/2007_benchmarks.htm) Area I. Prevent epilepsy and its progression D. Identify new ways to prevent epilepsy or stop it once it begins E. Develop new animal models to study epileptogenesis. Area II. Develop new treatment strategies and improve current approaches in order to cure epilepsy C. Improve current treatments and develop new technologies
|Cleary, Ryan T; Sun, Hongyu; Huynh, Thanhthao et al. (2013) Bumetanide enhances phenobarbital efficacy in a rat model of hypoxic neonatal seizures. PLoS One 8:e57148|