Neuroinflammation, Protein Aggregates, ApoE4 Drug Targeting, and Autophagy Rescue

神经炎症、蛋白质聚集体、ApoE4 药物靶向和自噬拯救

• 批准号: 10768318
• 负责人: Sue Tilton Griffin
• 金额: $ 47.03万
• 依托单位: UNIV OF ARKANSAS FOR MED SCIS
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2028-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10768318
• 关键词: Acute-Phase Proteins; Alzheimer' s Disease; Alzheimer' s neuropathogenesis; Amyloid beta-Protein; Apolipoprotein E; Astrocytes; Autophagocytosis; Binding; Brain; Brain Diseases; Brain region; Chemical Agents; Chronic; DNA; DNA Sequence; Development; Drug Targeting; Event; Exhibits; Feedback; Functional disorder; Genes; Genetic Transcription; Genotype; Glial Fibrillary Acidic Protein; Histologic; Individual; Inherited; Interleukin-1 beta; Lead; Microglia; Modification; Molecular; Motion; Neurofibrillary Tangles; Neurons; Outcome; Parents; Pathogenesis; Patients; Pharmaceutical Preparations; Phosphotransferases; Play; Prevention strategy; Production; Protein Inhibition; Proteins; Proteomics; Reporting; Repression; Role; Stress; Testing; Work; apolipoprotein E-4; brain tissue; chemical binding; cytokine; drug candidate; gain of function; hyperphosphorylated tau; neuroinflammation; novel; protein aggregation; protein protein interaction; small molecule; transcription factor

We established the pluripotent interleukin-1β (IL-1 β) cytokine as a significant player in the pathogenesis of Alzheimer’s disease as it sets in motion a self-amplifying positive-feedback cycle in which neuronal stress induces synthesis of the neuron's acute phase protein β APP and release of its fragments sAPPa and Aβ. Both these proteins activate microglia and a progressive elevation of IL-1 β, which drives chronically enhanced formation of the hallmark aggregates of AD: A β plaques and, via IL-1 β -induced synthesis and activation of specific kinases, hyperphosphorylation of tau in neurofibrillary tangles in neurons and glial fibrillary acidic protein (GFAP) in astrocytes. Interestingly, both events are dramatically enhanced in AD patients who inherit the Alzheimer gene from both parents (genotype ApoE4,4). Although these IL-1 β and ApoE genotype driven events favor negative outcomes, our progress in drug-development initiatives shows that they are amenable to treatment. Specifically, GFAP-binding chemical agents were shown to inhibit protein aggregation. Further, we discovered a novel function of the APOE£4 gene, which is a toxic gain-of- function exhibited by its protein product, ApoE4. We have demonstrated that ApoE4 competes with Transcriptional Factor EB (TFEB) for binding to the CLEAR DNA motifs, thus, hindering the transcription of three proteins crucial for lysosomal autophagy. We show this to be the case in brain tissues from AD 4,4, but not AD 3,3 patients. Now, importantly, we have identified a lead compound that binds to ApoE4 protein, obviating its interactions with CLEAR DNA and restoring the expression of three autophagy genes, that encode for production of p62, LC3B, and LAMP2 proteins. Now, we are prepared to further elucidate the role of IL-1 β in cellular pathophysiology, establishing its effects on kinases and kinase targets that manifest the modifications which drive predominant aggregate nucleation or propagation events. Our advanced molecular and histological approaches will be applied to confirm predictions of protein-protein interactions derived from proteomics approaches, particularly those involving GFAP. Moreover, these interactions can now be evaluated across brain regions and disease states to test their concordance with known AD parameters. Finally, we will elucidate the mechanisms of action of our identified novel small-molecule drugs targeted to ApoE4 in inhibiting all its pathognomonic interactions with other ApoE4 targeted DNA sequences. Successful completion of our proposed work promises a preventive strategy for foiling the known dramatic role that ApoE4 plays in Alzheimer neuropathogenesis. RELEVANCE: Successful completion of this proposed work will obviate the many pathognomonic aspects of inheritance of the Alzheimer gene (APOEE4) in the 1 in 4 individuals in the US, i.e., 80 million, who inherit one or both copies of this gene. Through the action of our specific drug candidates to inhibit the ApoE4 protein we will, therefore, restore the lysosomal autophagy necessary for efficient clearance of large aggregates, which, as we reported, is repressed in the brains of those who inherit one or both APOE4 genes.

我们确定多能白介素 1β (IL-1 β) 细胞因子是阿尔茨海默病发病机制中的重要参与者，因为它启动了自我放大的正反馈循环，其中神经元应激诱导神经元急性期蛋白 β APP 的合成并释放其片段 sAPPa 和 Aβ。这两种蛋白都会激活小胶质细胞和 IL-1 β 的逐渐升高，从而驱动 AD 标志性聚集体的长期增强形成：A β 斑块，并且通过 IL-1 β 诱导的特定激酶的合成和激活，神经元中的神经原纤维缠结中的 tau 蛋白和神经元中的神经胶质纤维酸性蛋白 (GFAP) 过度磷酸化。 星形胶质细胞。有趣的是，这两种事件在从父母那里继承阿尔茨海默基因（基因型 ApoE4,4）的 AD 患者中显着增强。尽管这些 IL-1 β 和 ApoE 基因型驱动的事件有利于负面结果，但我们在药物开发计划方面的进展表明它们适合治疗。具体来说，GFAP 结合化学试剂被证明可以抑制蛋白质聚集。此外，我们还发现了 APOE£4 基因的一个新功能，即其蛋白质产物 ApoE4 所表现出的有毒功能获得性。我们已经证明，ApoE4 与转录因子 EB (TFEB) 竞争与 CLEAR DNA 基序的结合，从而阻碍对溶酶体自噬至关重要的三种蛋白质的转录。我们在 AD 4,4 患者的脑组织中发现了这种情况，但在 AD 3,3 患者的脑组织中则不然。现在，重要的是，我们已经鉴定了一种与 ApoE4 蛋白结合的先导化合物，消除了它与 CLEAR DNA 的相互作用，并恢复了三个自噬基因的表达，这三个自噬基因编码 p62、LC3B 和 LAMP2 蛋白的产生。现在，我们准备进一步阐明 IL-1 β 在细胞病理生理学中的作用，确定其对激酶和激酶靶标的影响，这些激酶和激酶靶标表现出驱动主要聚集成核或增殖事件的修饰。我们先进的分子和组织学方法将用于确认源自蛋白质组学方法的蛋白质-蛋白质相互作用的预测，特别是涉及 GFAP 的方法。此外，现在可以跨大脑区域和疾病状态评估这些相互作用，以测试它们与已知 AD 参数的一致性。最后，我们将阐明我们确定的针对 ApoE4 的新型小分子药物在抑制其与其他 ApoE4 靶向 DNA 序列的所有特异性相互作用方面的作用机制。我们提出的工作的成功完成有望提供一种预防策略，以挫败 ApoE4 在阿尔茨海默病神经发病机制中所发挥的已知的戏剧性作用。 相关性：成功完成这项拟议的工作将消除美国四分之一的个体（即 8000 万人）中阿尔茨海默基因 (APOEE4) 遗传的许多特征方面，这些人继承了该基因的一个或两个拷贝。因此，通过我们的特定候选药物抑制 ApoE4 蛋白的作用，我们将恢复有效清除大聚集物所必需的溶酶体自噬，正如我们报道的那样，大聚集物在继承一种或两种 APOE4 基因的人的大脑中受到抑制。