Novel Targets for Stroke Intervention - Gene Discovery for Modulators of Infarction

中风干预的新靶点——梗死调节基因的发现

• 批准号: 10055780
• 负责人: Scott R Floyd
• 金额: $ 63.83万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-12-01 至 2022-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10055780
• 关键词: Adult; Agonist; Alleles; Biochemical; Biological Assay; Blood Vessels; Brain; Brain Injuries; Brain region; Candidate Disease Gene; Cause of Death; Cerebral Infarction; Cessation of life; Chromosome 8; Chromosome Mapping; Clinical; Complex; Data; Development; Distal; Drug Targeting; Engineering; Event; Explosion; Genes; Genetic; Glucose; Goals; Human; Hybrids; Inbred Strain; Inbred Strains Mice; Individual; Infarction; Intervention; Ischemia; Ischemic Stroke; Laboratory mice; Location; Maps; Medical; Modeling; Molecular; Mouse Strains; Mus; Nature; Neurons; Nutrient; Operative Surgical Procedures; Organ; Oxygen; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phenotype; Physiological; Population; Proteins; Publishing; RNA Interference; Rapid screening; Reproducibility; Research; Resources; Risk Factors; Scientist; Slice; Stroke; Surveys; Symptoms; System; Testing; Therapeutic; Time; Transfection; Transgenic Mice; Translations; Variant; Work; blood vessel occlusion; brain tissue; clinically relevant; deprivation; design; disease phenotype; drug development; druggable target; experience; forward genetics; gain of function; gene discovery; genetic approach; genetic variant; genome wide association study; genome-wide; genomic locus; improved; in vivo; innovation; ischemic injury; lipid metabolism; loss of function; mature animal; middle cerebral artery; mouse genetics; mouse genome; neuroprotection; new therapeutic target; novel; novel strategies; novel therapeutics; screening; stroke clinical trials; stroke intervention; stroke outcome; stroke patient; stroke risk; stroke symptom; stroke therapy; trait; whole genome

Occlusion of the blood vessels supplying the brain leads to ischemic stroke and infarction—irreversible death of brain tissue. Risk factors causing stroke, especially those involving lipid metabolism, form the basis of current therapies to reduce stroke risk. However, despite decades of research on the molecular events occurring during infarction, the translation of these discoveries to “druggable” targets to treat stroke outcome (death of brain tissue) has been quite disappointing. Novel approaches will be required to identify new and more physiologically relevant targets. The scientific premise of our proposal is that naturally occurring allelic variation underlies the profound differences in seen in stroke outcomes and that these neuro-protective gene variants would provide a novel path towards new targets for stroke treatment. However, genetic mapping approaches for infarct size in the human (e.g., GWAS of infarct volume among ischemic stroke patients) are intrinsically problematic due to wide variation in the extent and location of the occluded vessel, and especially, variation in the time window between first recognized symptoms and medical intervention. To date, we can find no published GWAS for infarct volume in ischemic stroke. The Marchuk lab has taken an alternative, forward genetic approach to discover novel genes modulating infarction. We have surgically occluded the distal middle cerebral artery in over 35 inbred mouse strains and found that infarct volume differs more than 50-fold. These robust and highly reproducible differences in infarct size are at least 10-fold larger than that seen in any engineered mouse lines but, importantly, are caused by natural allelic variation in the mouse genome. We have mapped several of these genetic loci and the goals of Aims 1 and 2 are to identify these novel genes regulating in infarct size. However, this gene discovery approach has required in vivo surgical assays in thousands of adult animals. We need a more scalable yet physiologically relevant screening platform to transform this approach to full genome-wide scale. The Lo lab has pioneered the development of such a discovery platform for cerebral infarction, simulating stroke by Oxygen/Glucose Deprivation (OGD; a well-characterized model for ischemic injury) in ex vivo brain tissue explants. Unlike isolated neurons in culture, brain slice explants retain the complex multicellular nature of the intact organ, and thus retain and represent the complex intercellular interactions occurring in brain tissue during cerebral infarction. In Aims 1 and 2, this ex vivo OGD platform will be used to identify the causative genes in our previously mapped loci. Our experience gained in these aims will lead to Aim 3, where the ex vivo OGD assay will be used to directly map and identify novel cerebral infarction genes, using the genetic mapping resource population of the Collaborative Cross. Our study takes advantage of innovative approaches developed by the co-PIs to implement a novel strategy for identifying novel drug targets to treat ischemic stroke.

供应大脑的血管闭塞会导致缺血性中风 梗塞——脑组织不可逆转的死亡。导致中风的危险因素，尤其是那些涉及脂质的因素 代谢，构成当前降低中风风险疗法的基础。然而，尽管几十年来 对梗塞期间发生的分子事件的研究，将这些发现转化为 治疗中风结果（脑组织死亡）的“药物”靶点非常令人失望。小说 需要采取一些方法来确定新的、与生理学更相关的目标。科学的 我们建议的前提是自然发生的等位基因变异是 在中风结果中看到这些神经保护基因变异将提供一条新的途径 朝着中风治疗的新目标迈进。然而，针对梗死面积的基因图谱方法 人类（例如，缺血性中风患者梗塞体积的 GWAS）本质上是有问题的，因为 闭塞血管的范围和位置的广泛变化，特别是时间的变化 首次识别症状和医疗干预之间的窗口。迄今为止，我们还没有发现任何已发表的 GWAS 用于评估缺血性中风的梗塞体积。 Marchuk 实验室采取了另一种方法，即正向遗传 发现调节梗塞的新基因的方法。我们通过手术封堵了远端中段 研究人员对超过 35 个近交系小鼠的脑动脉进行了研究，发现梗塞体积相差 50 倍以上。 这些梗死面积的稳健且高度可重复的差异至少比观察到的大 10 倍 在任何工程小鼠品系中，但重要的是，是由小鼠的自然等位基因变异引起的 基因组。我们已经绘制了其中几个遗传位点的图谱，目标 1 和 2 的目标是确定 这些新基因调节梗塞面积。然而，这种基因发现方法需要在体内 对数千只成年动物进行手术测定。我们需要一个更具可扩展性但与生理相关的 筛选平台将这种方法转变为全基因组规模。 Lo 实验室开创了 开发这样一个脑梗塞发现平台，通过氧气/葡萄糖模拟中风 离体脑组织外植体的剥夺（OGD；一种充分表征的缺血性损伤模型）。不像 培养中分离的神经元，脑切片外植体保留了完整器官的复杂多细胞性质， 从而保留并代表脑组织中发生的复杂的细胞间相互作用 脑梗塞。在目标 1 和 2 中，该离体 OGD 平台将用于识别病因 我们之前绘制的基因座中的基因。我们在这些目标中获得的经验将导致目标 3，其中 离体 OGD 测定将用于直接绘制和鉴定新的脑梗塞基因，使用 协作交叉的遗传图谱资源群体。我们的研究利用 联合首席研究员开发的创新方法，用于实施识别新药的新策略 治疗缺血性中风的目标。