New Inactivators of GABA Aminotransferase for Addiction

描述（由申请人提供）：本研究的长期目标是设计新的化合物来抑制GABA氨基转移酶（GABA-AT），该酶分解代谢抑制性神经递质GABA，用于治疗化学成瘾和癫痫。抑制GABA-AT，提高GABA水平，已被证明可以有效地抑制过度的神经活动，而不影响基础神经元放电。增加GABA水平可以阻断大鼠的可卡因、尼古丁、甲基苯丙胺、酒精和海洛因成瘾以及人类的可卡因成瘾，而不会影响对食物的渴望。此外，当GABA的浓度降低到大脑中的阈值水平以下时，会导致惊厥;提高大脑GABA水平会终止癫痫发作。对滥用药物的神经化学反应是脑桥核中多巴胺水平的急剧增加，这激活了负责快乐和奖励反应的神经元。多巴胺和相关行为的增加可以通过GABA浓度的增加来拮抗。氨己烯酸（商品名SabrilTM）是一种不可逆的GABA-AT抑制剂，目前作为儿科患者的单药治疗和成人难治性癫痫发作的预防治疗上市。正电子发射断层扫描（PET）显示，在灵长类动物中，氨己烯酸抑制可卡因诱导的多巴胺增加。然而，氨己烯酸用于治疗癫痫和兴奋剂成瘾两者的接受受到了对在长期施用大量氨己烯酸后25-50%的患者中的视野缺陷（VFD）的担忧的阻碍;典型剂量是每天1-3克。导致VFD的机制尚不清楚;尽管如此，如果普遍认为VFD是由长期暴露于大剂量药物引起的是正确的，并且如果可以使用低得多的药物剂量，则可能没有不良后果。发现一种新的GABA-AT灭活剂，（1 S，3S）-3-氨基-4-二氟亚甲基-1-环戊酸（2），在调节成瘾物质诱导的多巴胺增加和逆转大鼠可卡因成瘾方面的效力比氨己烯酸强300倍。由于2的效力，可以使用比氨己烯酸低得多的剂量，这可以预防VFD。本研究的一个重要目的是阐明2的失活机制，这将对未来的抑制剂设计非常有益。将进行涉及2和磷酸吡哆醛（PLP）辅酶的同位素标记的研究。另一个目的是合成新的GABA-AT灭活剂并研究其灭活机制;所提出的化合物之一应通过避免可能产生VFD的氨己烯酸潜在毒性副产物的途径进行。其他新化合物与2的结构相关，以进一步增强效力。第三个目的是确定酶上的灭活剂附着位点。第四个目标将涉及合作者Stephen Dewey博士使用PET研究这些新化合物对大鼠大脑中多巴胺释放的影响及其对大鼠成瘾的影响。最后一个目的是确定这些灭活剂对GABA-AT相对于其他PLP依赖性酶的选择性。 公共卫生相关性：这项研究的目的是设计新的化合物来阻断GABA氨基转移酶，这种酶催化抑制性神经递质GABA的破坏，从而增加大脑中GABA的水平。增加GABA水平可以阻断大鼠的可卡因、尼古丁、甲基苯丙胺、酒精和海洛因成瘾以及人类的可卡因成瘾，而不会影响对食物的渴望。此外，当GABA的浓度降低到脑中的阈值水平以下时，导致惊厥;提高脑GABA水平，例如通过抑制GABA转氨酶，终止癫痫发作。

DESCRIPTION (provided by applicant): The long-term goal of this research is to design new compounds to inactivate GABA aminotransferase (GABA- AT), the enzyme that catabolizes the inhibitory neurotransmitter GABA, for the treatment of chemical addiction and epilepsy. Inhibition of GABA-AT, which raises GABA levels, has been shown to effectively dampen excessive neural activity without affecting basal neuronal firing. Increasing GABA levels blocks cocaine, nicotine, methamphetamine, alcohol, and heroin addiction in rats and cocaine addiction in humans without affecting the craving for food. Also, when the concentration of GABA diminishes below a threshold level in the brain, convulsions result; raising the brain GABA levels terminates the seizure. The neurochemical response to drugs of abuse is a sharp increase in dopamine levels in the nucleus accumbens, which activates the neurons responsible for pleasure and reward responses. The rise in dopamine and associated behaviors can be antagonized by an increase in the concentration of GABA. Vigabatrin (trade name SabrilTM), an irreversible inhibitor of GABA-AT, is currently marketed as a monotherapy for pediatric patients and as an adjunctive therapy for adults with refractory seizures. It was shown by positron emission tomography (PET) in primates that vigabatrin inhibits these cocaine-induced dopamine increases. The acceptance of vigabatrin for the treatment of both epilepsy and stimulant addiction, however, has been hampered by concerns about visual field defects (VFDs) in 25-50% of patients following chronic administration of large amounts of vigabatrin; the typical dose is 1-3 grams a day. The mechanism leading to the VFDs is not known; nonetheless, if the prevailing belief that VFDs arise from prolonged exposure to large doses of drug is correct, and if much lower doses of a drug can be used, there may be no untoward consequences. A new GABA-AT inactivator, (1S,3S)- 3-amino-4-difluoromethylenyl-1-cyclopentanoic acid (2), was found to be 300 times more potent than vigabatrin in modulation of the dopamine increase induced by addictive substances and in reversal of cocaine addiction in rats. Because of the potency of 2, much lower doses can be used than those with vigabatrin, which may prevent the VFDs. An important aim of this proposal is to elucidate the inactivation mechanism of 2, which will be very beneficial to future inhibitor design. Studies involving isotopic labeling of 2 and of the pyridoxal phosphate (PLP) coenzyme will be carried out. Another aim is to synthesize new inactivators of GABA-AT and study their inactivation mechanisms; one of the proposed compounds should proceed by a pathway that avoids a potential toxic by-product of vigabatrin that might produce the VFDs. Other new compounds are related to the structure of 2 to enhance potency further. A third aim is to determine the site of inactivator attachment on the enzyme. A fourth aim will involve studies by collaborator Dr. Stephen Dewey on the effect of these new compounds on dopamine release in rat brains using PET and their effect on addiction in rats. The last aim is to determine the selectivity of these inactivators for GABA-AT relative to other PLP-dependent enzymes. PUBLIC HEALTH RELEVANCE: The aim of this research is to design new compounds to block GABA aminotransferase, the enzyme that catalyzes the destruction of the inhibitory neurotransmitter GABA, thereby increasing the brain levels of GABA. Increasing GABA levels blocks cocaine, nicotine, methamphetamine, alcohol, and heroin addiction in rats and cocaine addiction in humans without affecting the craving for food. Also, when the concentration of GABA diminishes below a threshold level in the brain, convulsions result; raising the brain GABA levels, for example by inhibiting GABA aminotransferase, terminates the seizure.