New Inactivators of GABA Aminotransferase for Epilepsy and Neuropathic Pain

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Project Summary The two principal neurotransmitters involved in the regulation of brain neuronal activity are γ-aminobutyric acid (GABA), one of the most widely distributed inhibitory neurotransmitters, and L-glutamic acid, an excitatory neurotransmitter. The concentration of GABA is regulated by two pyridoxal 5'-phosphate (PLP)-dependent enzymes, L-glutamic acid decarboxylase (GAD), which catalyzes the conversion of L-glutamate to GABA, and GABA aminotransferase (GABA-AT), which degrades GABA to succinic semialdehyde and converts α- ketoglutarate to L-glutamic acid. When the concentration of GABA diminishes below a threshold level, convulsions result; raising GABA levels terminates the seizure. When epilepsy is defined broadly as any disease characterized by recurring convulsive seizures, then over 1% of the entire world population (including >3 million Americans) can be classified as having epilepsy. One approach to raise GABA levels is with a molecule that crosses the blood-brain barrier (BBB) and inhibits/inactivates GABA-AT. This effectively dampens excessive neural activity without affecting basal neuronal firing. Neuropathic pain, including chemotherapy-induced peripheral neuropathy (a problem for more than 60% of cancer patients treated with chemotherapy), affects 3- 17% of the world population. Inadequate current treatments of pain are exacerbated by adverse side effects, such as abuse liability, sedation, and altered mental status, which limit treatment utility. Two features of neuropathic pain that have been identified are reduced GABA levels and spinal GABAergic inhibitory function. The objective of this proposal is to design and evaluate new mechanism-based inactivator analogs of our previously successful GABA-AT inactivators to enhance potency and elucidate inactivation mechanisms using computer modeling and crystallography (Dr. Dali Liu does our crystal structures) as the driving force for design. New inactivators are being designed for selective GABA-AT inactivation. The effectiveness of our new molecules will be tested by my collaborator, Dr. Andrea Hohmann, for their effect on various neuropathic pains, including chemotherapy-induced peripheral neuropathy. Specific Aims 1-3 in grant R01 NS123057 describes a multitude of molecules that were proposed to synthesize for testing as inactivators of GABA-AT. Our 20-year old HPLC/mass spectrometer has been repaired multiple times and now has not functioned in over 8 months. This dramatically diminishes our productivity to synthesize new molecules, making the time to analyze and identify intermediates and products very long. Consequently, the following aim is proposed: Specific Aim 1. Purchase and installation of a new mass spectrometer coupled with high-performance liquid chromatography. A routine training/maintenance policy will be established, assisted by the vendor.