Molecular and Cellular Mechanisms of the Lysosomal Storage Disease Cystinosis

溶酶体贮积病胱氨酸中毒的分子和细胞机制

• 批准号: 10801704
• 负责人: Sergio Daniel Catz
• 金额: $ 10.77万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-09 至 2025-05-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10801704
• 关键词: Affect; Animal Model; Apoptosis; Autophagocytosis; Brain; Cell Compartmentation; Cell Death; Cell model; Cell physiology; Cells; Cessation of life; Child; Collaborations; Complement; Complementary therapies; Creatinine; Cysteamine; Cystine; Cystinosis; Data; Defect; Deterioration; Development; Disease; Down-Regulation; Electrolytes; Eye; Failure; Fanconi Syndrome; Functional disorder; Glomerular Filtration Rate; Goals; Growth; Homeostasis; Human; Human Pathology; Immunofluorescence Immunologic; Impairment; Injury to Kidney; Kidney; Lead; Lesion; Link; Liver; Lysosomal Storage Diseases; Lysosomes; Mass Spectrum Analysis; Measures; Mediating; Membrane; Microscopy; Molecular; Molecular Chaperones; Mus; Mutation; Nature; Organ; Pathogenesis; Pathology; Pathway interactions; Patients; Pharmaceutical Chemistry; Pharmaceutical Preparations; Predisposition; Process; Proteins; Proteinuria; Recycling; Regulation; Renal function; Research; Resolution; Role; Sampling; Testing; Tissues; Vesicle; cell dedifferentiation; detection of nutrient; functional improvement; histological specimens; human disease; improved; in vivo; mutant; nephropathic cystinosis; nervous system disorder; novel; novel therapeutics; receptor; reconstitution; repaired; small molecule; tissue injury; trafficking; translational approach; urinary; young adult

SUMMARY Lysosomal function is crucial for cell homeostasis, autophagy, nutrient sensing, apoptosis and tissue remodeling. In lysosomal storage disorders (LSDs), characterized by genetic defects leading to anomalous accumulation of metabolites in lysosomes, cells are affected by lysosomal malfunction frequently leading to cell death. Cystinosis is a lysosomal storage disorder resulting from defects in the cystine transporter cystinosin (CTNS). Increased levels of intra-lysosomal cystine lead to cell malfunction and progressive tissue deterioration, which is especially manifested in kidneys. As with most LSDs, this leads to a slow but irreversible deterioration, organ dysfunction and early death. Patients with nephropathic cystinosis develop proximal tubule cell dedifferentiation, Fanconi syndrome and progressive renal injury, which are not corrected by the current therapy, cysteamine. Thus, cell malfunction and tissue failure occur despite cystine depletion, suggesting that cystine accumulation is not the only cause of all the defects observed in cystinosis. We recently revealed a defective mechanism of chaperone-mediated autophagy (CMA) in cystinosis. Defective CMA is directly linked to human disease, including kidney pathologies and neurological disorders. CMA defects in cystinosis are caused by mislocalization and downregulation of the only lysosomal CMA receptor, LAMP2A. Defective CMA activity correlates with high susceptibility to cell death in cystinosis. Importantly, the defect was not rescued by cystine depleting therapies supporting that it is independent of lysosomal overload. Our data highlight that CMA impairment is an important contributor to the pathogenesis of cystinosis and underline the need for new treatments to complement cystine depletion therapies. Our research plan aims to elucidate the molecular and cellular mechanisms leading to abnormal CMA activity in cystinosis. We also propose translational approaches that utilize small-molecule activators of CMA to improve cellular function in cystinosis. Our Specific Aims are: Aim 1: To understand the molecular basis of the regulation of LAMP2A function in cystinosis. To this end, we will study the interplay between the CTNS protein and the CMA receptor LAMP2A and elucidate the mechanisms that mediate LAMP2A trafficking and destabilization at the lysosomal membrane in cystinosis. Aim 2: To determine the molecular basis of the regulation of CMA activity and proximal tubule cell function in cystinosis. We will study the mechanisms mediated by CTNS to regulate CMA function and will test the hypothesis that the rescue of LAMP2A expression and CMA activity improves the function of proximal tubule cells from cystinotic patients. Aim 3: To utilize small-molecule CMA activators in vivo to improve renal function in cystinotic mice. We will correct cellular and renal function in cystinotic mice using CMA activators, alone, or in combination with cysteamine. Our research is highly significant because it aims to elucidate molecular mechanisms associated with a devastating human pathology and will help develop new therapies for the treatment of cystinosis and other human diseases.

摘要 溶酶体功能对细胞内稳态、自噬、营养感知、细胞凋亡和组织至关重要 改建。在溶酶体储存障碍(LSD)中，特征是遗传缺陷导致异常 代谢产物在溶酶体中积聚，细胞受溶酶体故障影响频繁导致细胞 死亡。胱氨酸病是一种溶酶体储存障碍，由胱氨酸转运蛋白的缺陷引起。 (CTNS)。溶酶体内胱氨酸水平升高导致细胞功能障碍和进行性组织 恶化，尤其表现在肾脏上。与大多数LSD一样，这将导致缓慢但不可逆转的 恶化、器官功能障碍和过早死亡。肾病患者出现近端小管 细胞去分化、Fanconi综合征和进行性肾损伤，这些都不能通过当前的 治疗，半胱胺。因此，尽管半胱氨酸耗尽，细胞功能障碍和组织衰竭仍会发生，这表明 胱氨酸积聚并不是观察到的胱氨酸病所有缺陷的唯一原因。我们最近透露了一种 伴侣介导的自噬(CMA)在胱氨酸病中的缺陷机制。有缺陷的CMA直接链接 人类疾病，包括肾脏病理和神经疾病。膀胱氨酸病的CMA缺陷有 由唯一的溶酶体CMA受体LAMP2A的错误定位和下调引起。CMA有缺陷 在胱氨酸病中，活性与细胞死亡的高度易感性相关。重要的是，缺陷不是由 胱氨酸消耗疗法支持它不受溶酶体过载的影响。我们的数据强调了CMA 损伤是胱氨酸病发病的一个重要因素，并强调了新的 补充半胱氨酸耗竭疗法的治疗。我们的研究计划旨在阐明分子和 导致胱氨酸病CMA活性异常的细胞机制。我们还提出了翻译方法 利用CMA的小分子激活剂改善胱氨酸病的细胞功能。我们的具体目标是： 目的1：了解LAMP2A功能调控在胱氨酸病中的分子基础。为此，我们 将研究CTNS蛋白和CMA受体LAMP2A之间的相互作用，并阐明 在胱氨酸病中介导LAMP2A运输和溶酶体膜失稳的机制。目标 2：确定CMA活性和近曲小管细胞功能调节的分子基础 胱氨酸病。我们将研究CTN介导的调节CMA功能的机制，并将测试 挽救LAMP2A表达和CMA活性可改善近曲小管功能的假说 来自膀胱虫病患者的细胞。目的3：体内利用小分子CMA激活剂改善肾功能 在胱氨酸病小鼠身上。我们将单独使用CMA激活剂纠正膀胱炎小鼠的细胞和肾功能，或 与半胱胺结合使用。我们的研究非常有意义，因为它旨在阐明分子 与毁灭性的人类病理相关的机制，并将有助于开发新的治疗方法 治疗胱氨酸病和其他人类疾病。