Krabbe disease: Combined therapies for the central and peripheral nervous systems

克拉伯病：中枢和周围神经系统的联合疗法

• 批准号: 7570708
• 负责人: DAVID A WENGER
• 金额: $ 30.9万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-09-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7570708
• 关键词: Affect; Amino Acids; Animal Model; Apoptosis; Apoptotic; Biochemical; Blood; Bone Marrow Cell Transplantation; Bone Marrow Cells; Brain; Breeding; CMV promoter; Canis familiaris; Catabolism; Cells; Cerebellum; Cessation of life; Clinical; Cognitive; Combined Modality Therapy; DNA; Demyelinations; Development; Disease; Dose; Drops; Elements; Embryo; Enzymes; Event; Galactolipids; Galactosylceramides; Genes; Globoid cell leukodystrophy; Gold; Health; Hematopoietic Stem Cell Transplantation; Hereditary Disease; Human; In Vitro; Indomethacin; Infant; Inflammation; Inflammatory; Injection of therapeutic agent; Intraventricular; Laboratories; Lead; Lentivirus Vector; Leukocytes; Life; Limb structure; Macaca mulatta; Maintenance; Major Histocompatibility Complex; Measures; Microglia; Minocycline; Modeling; Motor; Mus; Muscle; Myelin; Neonatal; Neurodegenerative Disorders; Normal Cell; Oligodendroglia; Organ; Outcome; Partner in relationship; Pathogenesis; Pathology; Patients; Peripheral Nervous System; Pharmaceutical Preparations; Play; Process; Psychosine; Regulatory Element; Residual state; Role; Schwann Cells; Source; Staging; Stem cell transplant; Testing; Tetanus Helper Peptide; Tetracycline Control; Tetracyclines; Time; Time Study; Transgenes; Transgenic Mice; Transgenic Organisms; Transplantation; Viral Vector; analog; base; cytokine; depressed; dosage; galactosylceramidase; gene therapy; improved; in vivo; macrophage; motor deficit; mouse model; myelination; nerve stem cell; neuroprotection; prevent; promoter; standard care; success; vector

DESCRIPTION (provided by applicant): This is a proposal to continue our studies on the treatment of the mouse models of Krabbe disease or globoid cell leukodystrophy (GLD), an autosomal recessive disorder caused by the deficiency of galactocerebrosidase (GALC) activity. GALC is responsible for the lysosomal catabolism of galactosylceramide and psychosine. In GLD, the accumulation of psychosine in the CNS and PNS results in the apoptotic death of myelin-forming cells. Other mechanisms, including activation of resident microglial cells, recruitment of blood macrophages and inflammation, also play important roles in the pathogenesis. The only treatment currently available for some patients is hematopoietic stem cell transplantation (HSCT). While this treatment helps to maintain cognitive abilities, motor function remains a problem. Therapy trials with viral vectors and HSCT in the dog and mouse models have not lead to complete clinical, biochemical and pathological correction. It is clear that combined therapy will be required for entirely successful treatment. Questions related to early pathological changes and timing and dosage of GALC activity remain to be answered. In Aim 1, we will generate transgenic mice that have GALC expression under control of a tetracycline (tet) responsive promoter. Initially, we will produce Tet-Off mice (trG) that will have high GALC expression in the absence of tet analogues. After detailed characterization of these mice, they will be mated with heterozygous GALC-deficient mice to produce mice with no GALC activity except that regulated by tet analogues. The trG mice will be used to answer important questions regarding the timing of treatment and dosage of GALC activity needed for correction, and as a source of stem cells for transplantation. In Aim 2, we will improve upon the outcome of BMT by preparing and testing viral vectors for in vivo and ex vivo therapy trials. We have prepared a high titer lentiviral vector both for raising GALC activity in bone marrow cells for transplantation and for direct intracerebral injection. In addition we will construct a lentiviral vector containing both the mouse GALC gene and a tet responsive element. This vector will be used to transduce mouse BM cells that will have GALC high activity, above the low levels present in untransduced cells from normal donor mice and, in addition, will have the expression of GALC under control of tet analogues. We will determine the ability of GALC-AAV2/1 injected into the hind limb muscles to attain high GALC expression in order to correct the residual motor deficits in mice receiving BMT. Also, intracerebral, intraventricular and intracerebellar injections of mGALC-AAV2/1 will be combined with BMT and approved drugs that could reduce inflammation and prevent apoptosis. Initially these drugs will include minocycline and Indomethacin. These drugs have been successful in reducing demyelination and in providing neuroprotection in other neurodegenerative disorders. Studies using the dog model of GLD have been dropped from this application. Krabbe disease a genetic disease affecting mostly infants. Treatment options are limited at this time, but the studies proposed in animal models could lead the way to improved therapies for human patients. A combination of therapies may be needed to both stop the pathological events and provide long-term correction.

描述（由申请人提供）：这是我们继续研究蟹黄病或球状细胞白质营养不良（GLD）小鼠模型的建议，GLD是一种由半乳糖脑苷酶（GALC）活性缺乏引起的常染色体隐性疾病。GALC负责半乳糖神经酰胺和精神素的溶酶体分解代谢。在GLD中，精神素在CNS和PNS中的积累导致髓磷脂形成细胞的凋亡性死亡。其他机制，包括常驻小胶质细胞的激活、血液巨噬细胞的募集和炎症，也在发病机制中发挥重要作用。目前对一些患者唯一可用的治疗方法是造血干细胞移植（HSCT）。虽然这种治疗有助于维持认知能力，但运动功能仍然是个问题。在狗和小鼠模型中使用病毒载体和HSCT的治疗试验尚未导致完全的临床、生化和病理纠正。很明显，联合治疗将需要完全成功的治疗。与早期病理改变以及GALC活性的时间和剂量有关的问题仍有待回答。在Aim 1中，我们将产生在四环素（tet）应答启动子控制下具有GALC表达的转基因小鼠。首先，我们将生产在缺乏tet类似物的情况下具有高GALC表达的tet - off小鼠（trG）。在详细描述了这些小鼠的特征后，将它们与GALC缺陷的杂合小鼠交配，产生除了受tet类似物调节外没有GALC活性的小鼠。trG小鼠将用于回答有关治疗时间和纠正所需GALC活性剂量的重要问题，并作为移植干细胞的来源。在目标2中，我们将通过准备和测试体内和体外治疗试验的病毒载体来改善BMT的结果。我们制备了一种高滴度的慢病毒载体，既可以提高移植用骨髓细胞GALC的活性，也可以直接用于脑内注射。此外，我们将构建包含小鼠GALC基因和tet应答元件的慢病毒载体。该载体将用于转导小鼠BM细胞，该细胞将具有GALC高活性，高于正常供体小鼠未转导细胞中存在的低水平，此外，GALC的表达将在tet类似物的控制下。我们将确定将GALC- aav2 /1注射到后肢肌肉中获得高GALC表达的能力，以纠正BMT小鼠的残留运动缺陷。此外，脑内、脑室和小脑内注射mGALC-AAV2/1将与BMT和已批准的减少炎症和防止细胞凋亡的药物联合使用。最初这些药物将包括米诺环素和吲哚美辛。这些药物已成功地减少脱髓鞘和提供神经保护在其他神经退行性疾病。使用GLD的狗模型的研究已从该应用程序中删除。克拉伯病是一种主要影响婴儿的遗传性疾病。目前的治疗选择有限，但在动物模型中提出的研究可能会为人类患者带来更好的治疗方法。可能需要联合治疗来阻止病理事件并提供长期纠正。