Development and characterization of bumetanide derivatives with selective effect on the neuronal cation-chloride cotransporter NKCC1 for studies on the role of NKCC1 in ictogenesis and epileptogenesis

• Applicant: Professor Dr. Wolfgang Löscher
• Subject Area: Molecular Biology and Physiology of Neurons and Glial Cells ; Cognitive, Systems and Behavioural Neurobiology ; Pharmacology ; Veterinary Medical Science
• Term: from 2015 to 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 279625255

Epilepsies are one of the most frequent neurological diseases. Despite the development of numerous new antiepileptic drugs in the last some 20 years, about 30% of the patients do not become seizure-free, i.e., are pharmacoresistant. For some types of epileptic seizures, e.g., neonatal seizures, the situation is even worse, because none of the available drugs exert any sufficient efficacy. Thus, novel strategies for development of more efficacious antiepileptic drugs are an urgent medical need. Furthermore, there is an urgent clinical need to develop drugs that prevent development of epilepsies after brain insults, i.e., exert a prophylactic (antiepileptogenic) effect. The diuretic drug bumetanide, a selective inhibitor of Na-K-Cl-cotransporters (NKCCs), has been discussed for years as a highly interesting antiepileptic and antiepileptogenic compound, because increased expression of neuronal NKCC1 and the resulting shift in GABA functionality seems to play an important role in epileptogenesis and as a mechanism of pharmacoresistance in neonatal and adult epilepsies. Experimental studies with bumetanide seemed to support this idea; however, first clinical trials in children with neonatal seizures were negative. This is an obvious consequence of the high ionization rate of bumetanide in plasma, resulting in poor penetration into the brain, which was first reported by the Löscher group in 2010. As a consequence, this group developed lipophilic prodrugs of bumetanide, which penetrate into the brain and release bumetanide. However, this strategy could not resolve some significant disadvantages of bumetanide, e.g., the lack of selectivity for renal NKCC2 vs. neuronal NKCC1. Aim of the planned studies described in this application is the development and characterization of novel brain-penetrating bumetanide derivatives with highly selective effect on neuronal NKCC1. Starting point for this aim are bumetanide derivatives, which have been synthesized by our cooperation partner Peter Feit during development of bumetanide, using derivatives that are significantly different from bumetanide in both structural and diuretic properties. Based on data with these derivatives, additional compounds will by synthesized by our cooperation partner Thomas Erker. The inhibitory potency of these compounds on NKCC1 vs. NKCC2 will be determined by an oocyte expression assay. In these experiments, we will also study the NKCC1-splice variant NKCC1b, which is predominantly expressed in the brain. The most interesting derivatives will be tested in in vivo models in mice and rats for (a) brain penetration, (b) diuretic potency, and (c) antiepileptic and antiepileptogenic effects. From these comprehensive studies, we expect not only an optimization of the bumetanide structure for treatment of brain diseases but also a better understanding of the role of NKCC1 in epileptogenesis and ictogenesis.

• DFG Programme: Research Grants