Development of GSK-3 Beta Inhibitors for the Treatment of Parkinson's Disease

开发用于治疗帕金森病的 GSK-3 Beta 抑制剂

• 批准号: 7596052
• 负责人: Jia Zhou
• 金额: $ 28.26万
• 依托单位: PSYCHOGENICS, INC.
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-30 至 2009-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7596052
• 关键词: 1-Methyl-4-phenylpyridinium; Adverse effects; Affect; Age; Aging; Agreement; Alzheimer' s Disease; Animals; Arizona; Arts; Autopsy; Benzofurans; Biochemistry; Bipolar Disorder; Brain; Brain region; Catechols; Cells; Central Nervous System Diseases; Chemistry; Chicago; Chronic; Collaborations; Comorbidity; Complex; Condition; Cytoskeleton; Data; Degenerative Disorder; Development; Disease; Dissociation; Dopamine Agonists; Dose; Ensure; Equipment; Family; Frontotemporal Dementia; Funding; Genes; Glycogen Synthase Kinase 3; Glycogen Synthase Kinases; Goals; Grant; Hand; Housing; Human; Illinois; In Vitro; Inclusion Bodies; Indazoles; Indoles; Injection of therapeutic agent; Investigation; Laboratory Research; Lead; Legal patent; Lesion; Levodopa; Lewy Bodies; Libraries; Licensing; Ligands; Link; Lithium; Maleimides; Modeling; Modification; Molecular Biology; Mus; Mutation; Nerve; Nerve Degeneration; Neurodegenerative Disorders; Neurofibrillary Tangles; Neurologic; Neurons; Neuroprotective Agents; Neurosciences; Numbers; Organic Chemistry; Parkinson Disease; Parkinson' s Dementia; Parkinsonian Disorders; Pathology; Patients; Pattern; Personal Satisfaction; Pharmaceutical Chemistry; Pharmaceutical Preparations; Pharmacotherapy; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Proteins; Public Health; Relative (related person); Research; Research Proposals; Role; Science; Screening procedure; Senior Scientist; Single Nucleotide Polymorphism; Site; Small Business Technology Transfer Research; Societies; Specific qualifier value; Staurosporine; Symptoms; Tauopathies; Technology; Testing; Therapeutic; Time; Toxin; Transferase; Treatment Efficacy; Triton X100; Universities; Work; age related; analog; base; benzofuran; chemical synthesis; cytotoxicity; day; design; drug development; experience; follow-up; hyperphosphorylated tau; in vivo; indole; inhibitor/antagonist; kinase inhibitor; neuroprotection; neurorestoration; novel; payment; presynaptic; prevent; response; sarkosyl; scale up; small molecule; synuclein; synucleinopathy; tau Proteins; tau aggregation; tau-protein kinase

DESCRIPTION (provided by applicant): Parkinson's disease (PD), a chronic and progressive neurological condition and one of aging and age- related diseases, affects approximately 1.5 million people in the US alone. PD not only places severe burden on the patients but also on their family and society. In 2003, approximately US $2.3 billion was spent on drug therapy worldwide to treat Parkinson's disease. Although great strides have been made in the development of agents to treat PD, the existing PD drugs such as dopamine (DA) agonists, levodopa and catechol-O-methyl transferase inhibitors (COMT) only treat the symptoms of the disease, and are also fraught with adverse effects and long-term complications. Consequently, there is a great need for developing disease-modifying and neuroprotective as well as neurorestorative drugs which can slow down or stop the disease from progressing. Recent research suggests that inhibition of the glycogen synthase kinase-3¿ (GSK-3¿) by small molecules may offer an important strategy in the treatment of a number of neurodegenerative diseases including Alzheimer's disease (AD), but its usefulness in PD has not been described. We have recently found in the cellular and animal MPTP model of parkinsonism, an induction in the hyperphosphorylated form of Tau, p-Tau, with hyperphosphorylation seen at many sites, including those found in neurofibrillary tangles of AD. The increase in p-Tau levels was strictly dependent upon the presence of a- Syn. This requirement for a-Syn was mandatory, since in a-Syn-/- mice, and in transfected cells not expressing any a-Syn, the toxin failed to induce p-Tau. MPTP also caused an increase in a-Syn protein levels. Hyperphosphorylation of Tau lead to its dissociation from the cytoskeleton, and aggregates of p-Tau were seen in sarkosyl-insoluble and Triton X-100-insoluble fractions. a-Syn was able to form stable heteromeric protein complexes with p-Tau, and p-Tau aggregates were seen in mature inclusion bodies of a-Syn. MPP+ caused the activation of several p-Tau-specific kinases, such as GSK-3¿ and p-ERK. Blockade of GSK-3¿ not only prevented, but also reduced, MPP+-induced p-Tau formation, a-Syn accumulation and cytotoxicity. Very new data obtained in human postmortem PD brains, in collaboration with Dr. Jeffrey Joyce, show a similar pattern of pathology: increased a-Syn accumulation, hyperphosphorylation of Tau at sites similar to the MPTP models [pSer262 and pSer396/404], lack of phosphorylation at sites not seen with MPTP [pSer202], and large increases in GSK-3¿. Interestingly, these pathological changes were further augmented in PD patients with dementia [PD + DEM]. Together, these findings in PD brains confirm the validity of our findings with the MPTP models. Importantly, prior to our findings, a possible role for GSK-3¿ in PD has not been previously described, although its role in AD is well studied. In addition to our findings, another study found strong linkage of two single nucleotide polymorphisms in the GSK-3¿ gene to sporadic PD. Thus, GSK-3¿ presents a novel target site in the development of novel therapies for PD. To date, we have identified some nM potency GSK-3¿ inhibitors that emerged from our SAR studies of staurosporine. A number of these designed staurosporine analogs have been screened against a family of 30 kinases. Among the compounds tested we found one, an indolyl-indazolylmaleimide, that was able to inhibit 98% of the kinase activity of GSK-3¿ when tested at a concentration of 10 ¿M. After further structural modifications described below, novel 3-(indol-3-yl)-4-(benzofuran-3-yl)maleimides having a Ki value as low as 2 nM selectively against GSK-3¿ relative to 30 additional kinases were identified. Moreover, we have been able to show that some of these ligands are able to exert a neuroprotective and neurorestorative action in vitro. The ultimate goal would be to identify one or two GSK-3¿ inhibitors that could be further developed for slowing or halting the progression of PD. Through funding from this STTR grant, we intend to follow-up on the exciting preliminary findings we have made in pursuit of novel GSK-3¿ inhibitors as potential therapeutics for the treatment of PD. To achieve this goal, the Specific Aims of this research proposal are as follows: 1. Compound selection and synthesis: Based upon the compound library in hand, 10 potent GSK- 3¿ inhibitors will be resynthesized for further in vitro pharmacological studies. Then 2 or 3 of the most promising ligands based upon the in vitro profile will be scaled up (about 5 grams each) for in vivo animal studies. 2. Neuroprotective In vitro studies in transfected cells and neurons: Investigation of time course and dose response of the compounds for inhibition of GSK-3¿, analyses of other kinases and phosphatases that are modulated by these compounds, and comparison of our findings with the effects using lithium. 3. Neurorestorative in vitro studies in transfected cells and neurons: After the initiation of cytotoxicity, we will examine the time and dose of GSK-3¿ inhibitors necessary for neurorestoration. 4. In vivo animal studies: Injection with MPTP for 5 days will be followed by simultaneous or delayed injections with increasing doses of the GSK-3¿ inhibitors for different time periods. GSK-3¿, a-Syn, Tau, kinases and phosphatases will be examined, as specified in Specific Aims 2 and 3. Key words: GSK-3¿ inhibitors, Kinases, Selectivity, Parkinson's disease, Therapeutics, Staurosporine analogs, Neuroprotective agents, Neurorestorative agents, Neuroprotection, a-Synuclein, p-Tau, Taupathies, Synucleopathies, MPTP model, Aging, Neurodegenerative, Age-related diseases, Alzheimer's disease. Project Des inson's disease (PD), the second most common neurodegenerative disease next to Alzheimer's disease (AD), is a progressive neurological condition associated with aging. Our proposal entails the chemical synthesis of novel staurosporine analogues as potent glycogen synthase kinase-3¿ (GSK-3¿) inhibitors, and in vitro studies in transfected cells and neurons as well as in vivo studies in animals of these compounds, which have the great potential to be developed as neuroprotective and neurorestorative therapies to slow down or stop Parkinson's disease from progressing. Public Health

描述（由申请人提供）：帕金森病（PD）是一种慢性进行性神经系统疾病，也是一种衰老和与年龄相关的疾病，仅在美国就影响着大约150万人，PD不仅给患者带来了沉重的负担。 2003 年，全世界用于治疗帕金森病的药物治疗花费了约 23 亿美元，尽管在治疗帕金森病的药物开发方面取得了巨大进展，但现有的帕金森病药物如多巴胺（DA）激动剂、左旋多巴和儿茶酚-O-甲基转移酶抑制剂（COMT）只能治疗该疾病的症状，而且也充满了不良反应和长期并发症，经测试，有很大的需要。修饰和神经保护以及神经恢复药物可以减缓或阻止疾病的进展。最近的研究表明，抑制糖原合酶激酶 3¿ (GSK-3¿) 的小分子可能为治疗包括阿尔茨海默氏病 (AD) 在内的许多神经退行性疾病提供重要策略，但我们最近在细胞和动物 MPTP 中尚未发现其在 PD 中的用途。帕金森病模型，Tau 的过度磷酸化形式（p-Tau）的诱导，在许多位点都可见过度磷酸化，包括在 AD 的神经原纤维缠结中发现的 p-Tau 水平增加。严格依赖于 a-Syn 的存在。对 a-Syn 的这种要求是强制性的，因为在 a-Syn-/- 小鼠中以及在不表达任何 a-Syn 的转染细胞中，该毒素无法诱导 p-Tau。 MPTP 还导致 Tau 的 a-Syn 蛋白水平增加，导致其从细胞骨架解离，并且在 sarkosyl-insolve 和 Triton 中观察到 p-Tau 的聚集。 X-100 不溶性组分能够与 p-Tau 形成稳定的异聚蛋白复合物，并且在 a-Syn 的成熟包涵体中观察到 p-Tau 聚集体，导致多个 p-Tau 特异性的激活。激酶，例如 GSK-3¿和 p-ERK 的封锁。与 Jeffrey Joyce 博士合作，在人类死后 PD 大脑中获得的非常新的数据显示了类似的病理模式：不仅阻止而且减少了 MPP+ 诱导的 p-Tau 形成、a-Syn 积累和细胞毒性。 -Syn 积累，Tau 在类似于 MPTP 模型 [pSer262 和 pSer396/404] 的位点过度磷酸化，在 MPTP 未见的位点缺乏磷酸化[pSer202]，并且 GSK-3 大幅增加¿暗示性的是，这些病理变化在患有痴呆的 PD 患者中进一步增强 [PD + DEM]，这些在 PD 大脑中的发现证实了我们在 MPTP 模型中的发现的有效性，重要的是，在我们的发现之前，GSK 可能发挥作用。 -3¿尽管 GSK-3 在 AD 中的作用已得到充分研究，但之前尚未对其在 PD 中的作用进行过描述。除了我们的发现之外，另一项研究还发现 GSK-3 中两个单核苷酸多态性之间存在很强的联系。因此，GSK-3¿迄今为止，我们已经确定了 GSK-3 的一些 nM 效力。我们对星形孢菌素的 SAR 研究中产生的抑制剂已针对 30 种激酶家族进行了筛选，我们发现一种吲哚基吲唑马来酰亚胺能够抑制 98% 的激酶。 GSK-3的活性¿当测试浓度为 10 ¿ M. 经过下述进一步的结构修饰，新型 3-(吲哚-3-基)-4-(苯并呋喃-3-基)马来酰亚胺具有低至 2 nM 的 Ki 值，选择性地对抗 GSK-3¿此外，我们还发现其中一些配体能够在体外发挥神经保护和神经恢复作用，最终目标是鉴定一种或两种 GSK-3。通过 STTR 拨款的资助，我们打算进一步开发新的 GSK-3 抑制剂，以减缓或阻止 PD 的进展。抑制剂作为治疗 PD 的潜在疗法 为了实现这一目标，本研究提案的具体目标如下： 1. 化合物选择和合成：基于现有的化合物库，10 种有效的 GSK-3¿抑制剂将被重新合成以用于进一步的体外药理学研究，然后根据体外特征将2或3个最有希望的配体放大（每个约5克）用于体内动物研究。细胞和神经元：研究化合物抑制 GSK-3 的时间过程和剂量反应¿ 、对这些化合物调节的其他激酶和磷酸酶的分析，以及我们的研究结果与使用锂的效果的比较。 3. 转染细胞和神经元的神经恢复体外研究：细胞毒性开始后，我们将检查时间和剂量。 GSK-3?? 4. 体内动物研究：注射 MPTP 5 天，然后同时或延迟注射增加剂量的 GSK-3¿不同时间段的 GSK-3 抑制剂。 、a-Syn、Tau、激酶和磷酸酶将按照具体目标 2 和 3 中的规定进行检查。关键词：GSK-3¿抑制剂、激酶、选择性、帕金森病、治疗学、十字孢菌素类似物、神经保护剂、神经恢复剂、神经保护、a-突触核蛋白、p-Tau、Taupathies、突触核蛋白病、MPTP 模型、衰老、神经退行性、年龄相关疾病、阿尔茨海默病项目。 Des inson 病 (PD)，是仅次于阿尔茨海默病的第二常见神经退行性疾病(AD) 是一种与衰老相关的进行性神经系统疾病，我们的建议需要化学合成新型十字孢菌素类似物作为有效的糖原合成酶激酶 3¿ (GSK-3¿) 抑制剂、转染细胞和神经元的体外研究以及这些化合物的动物体内研究，这些化合物具有开发为神经保护和神经恢复疗法以减缓或阻止帕金森病的巨大潜力。公共卫生进展