Mitophagy-driven selection against heteroplasmic mitochondrial DNA mutations

线粒体自噬驱动的针对异质线粒体 DNA 突变的选择

• 批准号: 8323862
• 负责人: DAVID K. SIMON
• 金额: $ 21.75万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8323862
• 关键词: Age; Aging; Attenuated; Autophagocytosis; Behavioral; Blood - brain barrier anatomy; Brain; Cardiac; Cardiomyopathies; Cell Line; Cell physiology; Cells; Chronic; Clinical Treatment; Clinical Trials; Cognitive deficits; Complex; DNA; DNA Fingerprinting; DNA-Directed DNA Polymerase; Data; Defect; Disease; Dose; FDA approved; Family; Functional disorder; Gene Mutation; Genome; Hereditary Disease; Heredity; Human; Impairment; Inherited; Laboratories; Lead; Longevity; Metabolic; Mitochondria; Mitochondrial DNA; Mitochondrial Diseases; Mus; Mutation; Myopathy; Neurodegenerative Disorders; Neurologic; Non-Insulin-Dependent Diabetes Mellitus; Oral; Oxidative Stress; Parkinson Disease; Parkinsonian Disorders; Patients; Phenotype; Phosphotransferases; Premature aging syndrome; Process; Resistance; Resources; Role; Safety; Seizures; Sirolimus; Stroke; Testing; Therapeutic; Time; Tissues; Translations; Work; age related; cancer therapy; cell motility; clinical application; effective therapy; fly; gene therapy; high risk; human FRAP1 protein; kinase inhibitor; member; mitochondrial DNA mutation; mitochondrial genome; mutant; neurochemistry; novel; novel strategies; optic nerve disorder

DESCRIPTION (provided by applicant): Disorders caused by maternally inherited pathogenic mitochondrial DNA (mtDNA) mutations can lead to a wide array of neurological, cardiac, and other disorders. Unfortunately, clearly effective clinical treatments for these often devastating disorders are lacking. An ideal strategy would eliminate the mutant mtDNA and replace it with wild type (WT) DNA. However, classic "gene therapy" approaches are difficult to apply to mtDNA mutations. On the other hand, mitochondria undergo frequent turnover (every few days), even in postmitotic cells, with only a subset of copies of the mitochondrial genome being replicated during this process, providing an opportunity to influence which mtDNA molecules are replicated. We now propose to test a novel strategy to promote the selective elimination of deleterious mtDNA mutations that can be applied to heteroplasmic mtDNA mutations. Heteroplasmy is a common feature of pathogenic mtDNA mutations, and refers to a mix of WT and mutant mtDNA within the same cells or tissue. Our hypothesis takes advantage of a natural cellular process known as "mitophagy" (mitochondrial degradation by autophagy), which is a mechanism for selectively eliminating dysfunctional mitochondria. We hypothesize that some mitochondria within a cell will harbor greater levels of a heteroplasmic mtDNA mutation than others. Those with greater levels of a deleterious mutation will tend to have relatively greater impairment of mitochondrial function. Therefore, we propose to test the novel hypothesize that stimulating mitophagy by inhibiting mTOR kinase activity in cells harboring a heteroplasmic pathogenic mtDNA mutation will drive selection against the mutant mtDNA, over time leading to a substantial reduction in the mutational burden and hence an improvement in mitochondrial function. We have a unique resource available for testing this hypothesis: multiple SH-SY5Y cybrid cell lines harboring different levels of a heteroplasmic G11778A complex I (CI) gene mutation associated with Leber's Heredity Optic Neuropathy (LHON), all prepared at the same time from members of a single family. Our preliminary data with these cell lines support our hypothesis. A second important resource in our laboratory is the "mutator" mouse that expresses a proofreading deficient mtDNA polymerase 3 (Polg) leading to accumulation with age of heteroplasmic somatic mtDNA mutations in association with a premature aging phenotype. Our preliminary data demonstrate substantial metabolic, behavioral, and neurochemical deficits in these mice. We now hypothesize that enhancing mitophagy in the Polg mutator mice will attenuate the accumulation of somatic mtDNA mutations and ameliorate the deficits in these mice. Ultimately, clinical applications of this strategy have the potential to be of benefit to patients with classic mitochondrial disorders associated with heteroplasmic mtDNA mutations, to families harboring Polg mutations associated with familial parkinsonism and other disorders, and potentially for age-related neurodegenerative disorders.

描述（由申请人提供）：由母系遗传致病性线粒体DNA （mtDNA）突变引起的疾病可导致广泛的神经、心脏和其他疾病。不幸的是，对于这些通常具有毁灭性的疾病，缺乏明确有效的临床治疗方法。理想的策略是消除突变的mtDNA并用野生型（WT） DNA代替。然而，经典的“基因治疗”方法很难应用于mtDNA突变。另一方面，即使在有丝分裂后的细胞中，线粒体也经历频繁的更新（每隔几天），在这一过程中，只有一小部分线粒体基因组拷贝被复制，这为影响哪些线粒体dna分子被复制提供了机会。我们现在提议测试一种新的策略来促进选择性消除有害的mtDNA突变，这种策略可以应用于异质mtDNA突变。异质性是致病性mtDNA突变的共同特征，是指在同一细胞或组织内WT和突变mtDNA的混合。我们的假设利用了被称为“线粒体自噬”（线粒体自噬降解）的自然细胞过程，这是一种选择性消除功能失调线粒体的机制。我们假设细胞内的一些线粒体会比其他线粒体有更高水平的异质线粒体dna突变。那些有害突变水平较高的人往往线粒体功能受损相对较大。因此，我们提出验证一种新的假设，即通过抑制携带异质致病性mtDNA突变的细胞中的mTOR激酶活性来刺激线粒体自噬，将驱动对突变mtDNA的选择，随着时间的推移，导致突变负担的大幅减少，从而改善线粒体功能。我们有一个独特的资源可用于验证这一假设：多个SH-SY5Y杂交细胞系含有不同水平的与Leber遗传性视神经病变（LHON）相关的异质G11778A复合物I （CI）基因突变，所有这些突变都是在同一时间从一个家族成员中制备的。这些细胞系的初步数据支持我们的假设。我们实验室的第二个重要资源是“突变”小鼠，该小鼠表达校对缺陷mtDNA聚合酶3 (Polg)，导致与早衰表型相关的异质体细胞mtDNA突变随年龄增长而积累。我们的初步数据表明，这些小鼠存在大量的代谢、行为和神经化学缺陷。我们现在假设，增强Polg mutator小鼠的线粒体自噬将减少体细胞mtDNA突变的积累，并改善这些小鼠的缺陷。最终，该策略的临床应用有可能对与异质mtDNA突变相关的经典线粒体疾病患者、与家族性帕金森病和其他疾病相关的Polg突变家族以及与年龄相关的神经退行性疾病患者有益。