Development of predictive in vitro model of drug absorption across the human BBB

描述（由申请人提供）：血脑屏障（BBB）解释了为什么在血液中循环的某些物质能够进入大脑，而其他物质则不能。血脑屏障是由脑毛细血管形成的，看起来像简单的内皮细胞管。实验和临床证据表明，血脑屏障维持神经元功能的化学环境，并保护大脑免受有害物质的侵害。每年，制药公司花费数百万美元开发和测试体外血脑屏障模型的药物，其中许多最终在体内不起作用。在包括啮齿动物在内的小动物身上进行的研究不能直接外推到人体组织中。本实验室和其他实验室的初步结果令人信服地表明，使用啮齿动物脑内皮细胞和一般非人类来源的内皮细胞系作为临床药理学模型有时是不完美的。事实上，虽然许多耐药分子在啮齿动物的大脑中表达，但临床和体外观察表明，人类大脑似乎要复杂得多。相反，大多数体外有效的有希望的CNS药物由于疾病相关的多药耐药现象的出现而在临床试验中失败。我们建议开发和验证一个人性化的、实用的血脑屏障体外模型，以更准确地反映血脑屏障的临床现实。特别是，我们希望1)比较临床相关抗癫痫药物在药物反应患者中的血脑屏障通透性与由相同受试者或其他正常、非癫痫、非耐药人脑细胞组成的体外模型中测量的值。这将通过术中或体外测量血清和脑抗癫痫药物水平来实现；2)比较多重耐药患者中相同药物的血脑屏障通透性值与由同一受试者的细胞组成的体外模型中测量的值。这将通过测量药物在多重耐药人脑中的渗透程度，以及在手术期间从人脑中分离出的细胞间的渗透程度，以缓解难治性癫痫发作。PAR-07-049希望“加速药物发现的过程”和“开发允许化合物初步筛选的细胞分析”。我们坚信，提高体外血脑屏障模型的预测价值是朝着这个方向迈出的关键一步。然而，本文提出的实验将对高通量筛选假定的中枢神经系统药物的成功产生深远影响。公共卫生相关性：三分之一的癫痫患者患有耐药性癫痫，这与死亡风险增加和使人衰弱的社会心理后果有关。人类癫痫耐药的基础尚不清楚。与癌症耐药的相似之处表明，耐药蛋白可能发挥了作用。我们认为，多重耐药癫痫患者的血管中布满了从大脑中挤出药物的分子，这对成功缓解癫痫发作造成了障碍。如果我们的假设是正确的，我们可能会获得治疗儿童和成人癫痫的新工具。

DESCRIPTION (provided by applicant): The blood-brain barrier (BBB) explains why certain substances circulating in the blood are able to get into the brain and others do not. The BBB is formed by brain capillaries that look like simple tubes of endothelial cells. Experimental and clinical evidence suggests that the BBB maintains the chemical environment for neuronal function and protects the brain from harmful substances. Every year millions of dollars are spent by pharmaceutical companies to develop and test drugs using in vitro models of the BBB, many of which end up not working in vivo. Studies performed in small animals including rodents cannot be directly extrapolated to human tissue. Preliminary results from this and other laboratories have convincingly demonstrated that use of rodent brain endothelial cells and in general endothelial cell lines from non human sources as models of clinical pharmacology are sometimes imperfect. In fact, while a number of drug resistance molecules are expressed in rodent brain, the human brain appears to be much more complicated as demonstrated by both clinical and in vitro observations. Conversely, most of the promising CNS drugs that are effective in vitro have failed clinical trials due to disease- related appearance of the multidrug resistance phenomenon. We propose to develop and validate a humanized, practical in vitro model of the BBB that more accurately reflects the clinical reality of the BBB. In particular, we wish to 1) compare the BBB permeability of clinically relevant antiepileptic drugs in pharmacorespondent patients to values measured in an in vitro model comprised of cells from the same subjects or other sources of normal, non-epileptic, non drug resistant human brain. This will be achieved by measuring serum and brain antiepileptic drug levels intraoperatively or in vitro; and 2) to compare BBB permeability values of the same drugs measured in multiple drug resistant patients to values measured in an in vitro model comprised of cells from the same subjects. This will be performed by measuring the penetration of drugs into multiple drug resistant human brain and across cells isolated from human brain during surgeries to relieve refractory seizures. PAR-07-049 wishes to "accelerate the process of drug discovery" and to "Develop cellular assays that permit the preliminary screening of compounds". We strongly believe that improving the predictive value of in vitro models of the blood- brain barrier is a crucial step in this direction. The experiments proposed herein will however have a profound impact on the success of high-throughput screening of putative CNS drugs. PUBLIC HEALTH RELEVANCE: One third of patients with epilepsy have drug-resistant epilepsy, which is associated with an increased risk of death and debilitating psychosocial consequences. The basis of drug resistance in human epilepsy is not understood. Parallels with resistance in cancer suggest that drug resistance proteins may have a role. We believe that blood vessels of multiple drug resistant epileptics are lined with molecules that extrude drugs from the brain, causing an impediment to successful relief of seizures. If our hypothesis is correct we may acquire new tools to treat epilepsy in children and adults.