Pathogenesis/Treatment-Inherited Cholesterol Deficiency

遗传性胆固醇缺乏症的发病机制/治疗

基本信息

批准号：
8644814
负责人：
CEDRIC HOWARD SHACKLETON
金额：
$ 33.15万
依托单位：
CHILDREN'S HOSPITAL & RES CTR AT OAKLAND
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-04-15 至 2016-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8644814
关键词：
7-dehydrocholesterol reductase Adverse effects Aftercare Animals Behavioral Biochemical Biochemical Markers Biochemistry Birth Blood Blood - brain barrier anatomy Brain CMV promoter Cell Count Cells Cerebrospinal Fluid Cholesterol Cholesterol Homeostasis Complex DNA Dehydrocholesterols Detection Development Diagnosis Diagnostic Procedure Disease Dose Dysmorphology Effectiveness Ensure Enzymes Family Fetal Development Fluorescent Antibody Technique Frequencies Gene Delivery Gene Expression Gene Transfer Gene Transfer Techniques Genes Genetic Goals Growth Healthcare Systems Hepatocyte Hereditary Disease Human Inherited Learning Liver Longevity Mass Spectrum Analysis Measures Memory Mental Retardation Messenger RNA Metabolism Methods Microscopic Monitor Mus Mutant Strains Mice Mutation Neuraxis Newborn Infant North America Oxidoreductase Pathogenesis Patients Performance Phenotype Photosensitivity Physiological Plant Roots Preventive Quality of life Reaction Reverse Transcriptase Polymerase Chain Reaction Risk Role Safety Severities Siblings Smith-Lemli-Opitz Syndrome Staining method Stains Sterols Survival Rate Symptoms Technology Testing Therapeutic Time Tissues Vertebral column Viral Vector Weight Gain base behavior test cellular transduction disease characteristic fetal gene therapy gene transfer vector grasp improved in utero killings morris water maze mouse model mutant neuromuscular novel diagnostics pre-clinical prenatal promoter public health relevance research study retinal rods vector

项目摘要

DESCRIPTION (provided by applicant): Smith-Lemli-Opitz syndrome (SLOS) is a dysmorphology and mental retardation disorder lacking adequate therapy options. It is caused by cholesterol deficiency due to inactivity of 7-dehydrocholesterol reductase (DHCR7). Biochemically, it is characterized by low concentrations of cholesterol (C) in blood and tissues and high concentrations of dehydrocholesterol (DHC). Although mutations in the DHCR7 gene have been well described, the pathogenic development of the disease characteristics is not well understood. Among inherited errors of metabolism, SLOS is relatively frequent, and new diagnostic methods aid its prenatal detection. The long-term objective of this proposal is to develop a basis for the treatment of SLOS. For experiments, gene transfer techniques and mouse models that mimic human SLOS will be used. Three Specific Aims are proposed: (Aim 1) Define the limitations of post-natal, systemic gene therapy. The ratio of DHC/C is an indicator of SLOS severity. So far we have improved, but not completely normalized, this ratio by gene therapy. We now seek to enhance the effectiveness of this gene therapy. Our hypothesis is that the number of liver cells able to express the introduced DHCR7 gene is the limiting factor. We shall test this by varying the dose and type of vector carrying a tagged DHCR7 gene, following the extent of transduction by microscopic identification of cells expressing a tagged enzyme (FLAG-DHCR7), and comparing with DHC/C. (Aim 2) Normalize cholesterol in the CNS by treating newborns. Cholesterol metabolism in the CNS must be treated separately because it is isolated from systemic cholesterol by the blood brain barrier. Gene transfer vector will be injected intrathecally into the cerebrospinal fluid of newborn SLOS mice. At various times brains will be analyzed for sterols, DHCR7 DNA and mRNA, and the distribution of cells expressing FLAG-DHCR7. Sustained normalization of DHC/C in brain will prompt behavioral tests to see if learning, memory and neuromuscular performance are improved. (Aim 3) Correct cholesterol levels during fetal development. Here the emphasis is on preventive treatment and establishing when during development irreversible damage occurs. Biochemical markers, dysmorphic features, and physiological indicators (e.g., survival rates, weight gain and photosensitivity) will all be monitored. If there is significant improvement we shall again explore neuromuscular and behavioral performance. For the three types of treatment, systemic, CNS and fetal, long- term consequences, safety and potential adverse effects will also be monitored. Due to the multiple essential roles of cholesterol, SLOS is a complicated and devastating disorder in spite of its relatively simple genetic origin. The frequency of SLOS in North America is estimated at about 1 in 60,000, although many cases are thought to be never diagnosed because of prenatal lethality or mild phenotype. Through the proposed experiments using gene transfer and SLOS mice, we hope to learn more about the timing and biochemistry of disease development and to establish a basis for eventual treatment in humans.

描述（由申请人提供）：Smith-Lemli-Opitz 综合征 (SLOS) 是一种缺乏足够治疗选择的形态畸形和精神发育迟滞障碍。它是由于 7-脱氢胆固醇还原酶 (DHCR7) 失活而导致胆固醇缺乏所致。从生化角度看，其特点是血液和组织中胆固醇（C）浓度低，而脱氢胆固醇（DHC）浓度高。尽管 DHCR7 基因的突变已得到很好的描述，但疾病特征的致病发展尚不清楚。在代谢遗传性错误中，SLOS 相对常见，新的诊断方法有助于其产前检测。该提案的长期目标是为 SLOS 的治疗奠定基础。在实验中，将使用基因转移技术和模仿人类 SLOS 的小鼠模型。提出了三个具体目标：（目标 1）明确产后系统基因治疗的局限性。 DHC/C 的比率是 SLOS 严重程度的指标。到目前为止，我们已经通过基因治疗改善了这一比例，但尚未完全正常化。我们现在寻求增强这种基因疗法的有效性。我们的假设是能够表达引入的 DHCR7 基因的肝细胞数量是限制因素。我们将通过改变携带标记 DHCR7 基因的载体的剂量和类型来测试这一点，通过显微镜鉴定表达标记酶 (FLAG-DHCR7) 的细胞来跟踪转导程度，并与 DHC/C 进行比较。（目标 2）通过治疗新生儿使中枢神经系统中的胆固醇正常化。中枢神经系统中的胆固醇代谢必须单独处理，因为它通过血脑屏障与全身胆固醇隔离。基因转移载体将被鞘内注射到新生 SLOS 小鼠的脑脊液中。在不同时间对大脑进行甾醇、DHCR7 DNA 和 mRNA 以及表达 FLAG-DHCR7 细胞的分布进行分析。大脑中 DHC/C 的持续正常化将促进行为测试，以了解学习、记忆和神经肌肉性能是否得到改善。（目标 3）纠正胎儿发育期间的胆固醇水平。这里的重点是预防性治疗以及确定发育过程中何时发生不可逆转的损害。生化标记、畸形特征和生理指标（例如存活率、体重增加和光敏性）都将受到监测。如果有显着改善，我们将再次探讨神经肌肉和行为表现。对于全身、中枢神经系统和胎儿这三种治疗类型，还将监测长期后果、安全性和潜在不良反应。由于胆固醇具有多种重要作用，尽管 SLOS 的遗传起源相对简单，但它是一种复杂且具有破坏性的疾病。在北美，SLOS 的发生率估计约为六万分之一，尽管许多病例由于产前致死率或轻微表型而被认为从未被诊断出来。通过所提出的使用基因转移和 SLOS 小鼠的实验，我们希望更多地了解疾病发展的时间和生物化学，并为人类的最终治疗奠定基础。