Proj 4: Chemical Chaperone Therapy of Batten Disease

项目 4：Batten 病的化学伴侣疗法

基本信息

批准号：
8150188
负责人：
Glyn Dawson
金额：
$ 20.23万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/8150188
关键词：
Address Adolescent Age Amino Acids Biological Biological Models Blood - brain barrier anatomy Blood capillaries Borates Brain CLN1 gene CLN2 gene Cells Central cord canal structure Chemicals Chickens Child Cholesterol Collaborations Complex Cultured Cells Defect Disease Drug Delivery Systems Embryo Endosomes Enzymes Fibroblasts Glass Glutamine Hereditary Disease Heterozygote Hippocampus (Brain)Histidine Human Hydrolase Hydrolysis Individual Infantile neuronal ceroid lipofuscinosis Injection of therapeutic agent Instruction Label Ligands Lysosomes Mediating Membrane Membrane Microdomains Mental Retardation Methods Methyl Green Missense Mutation Modeling Molecular Chaperones Monoglycerides Mutation Neonatal Nerve Degeneration Neuroglia Neurons Patients Peptide Hydrolases Peptides Pharmaceutical Preparations Point Mutation Proline Protein Region Proteins Psyche structure Quantum Dots Rat-1 Rattus Residual state Rhodamine Seizures Signal Transduction Slice Sphingolipids Spielmeyer-Vogt Disease Spinal Cord Structure Sulfhydryl Compounds Surface System Testing Therapeutic Tissues Toxic effect Work base capillary design disease-causing mutation enzyme activity esterase feeding fluorophore in vitro activity inhibitor/antagonist inorganic phosphate link protein lymphoblast lysyl-aspartyl-glutamyl-leucine novel novel strategies null mutation palmitoyl-protein hydrolase postnatal protein aminoacid sequence protein misfolding rabies virus glycoprotein G research study thioester trafficking tripeptidyl aminopeptidase tripeptidyl-peptidase I uptake

项目摘要

PROJECT SUMMARY (See instructions): Pharmacological chaperones (eg: AcGDap(Palm)VKIKK)can be internalized by cells and re-fold misfolded proteins to an active configuration, but need to cross the blood brain barrier, enter neurons and escape endosomes. We have shown that the palmitoylated peptide motif is uniquely able to allow drugs to escape endosomes and that such chaperones can reactivate misfolded proteins such as palmitoyhprotein thioesterase (PPTl). We now propose to design sequences to permit the chaperones to cross the blood brain barrier by either attaching a fluorophore and target motif (eg: rabies virus glycoprotein coat peptide (RVG)) or coating on 6-1 Onm quantum dots. We will test short sequences of proline and histidine, with a glutamine spacer to attach to the surface of 635 nm red QDs capped with the 4-thiol PEG ligand. We will test this in cultured postnatal neurons, and lymphoblasts from patients with defined point mutations in PPTl origin, and then use either the E3 embryonic chick spinal cord injection system, in collaboration with Project I, or the rat hippocampal slice system through collaboration with Project II. The three model systems represent embryonic brain, neonatal brain and postnatal brain and the collaborations will allow us to better assess the toxicity (if any) of these drugs, their efficacy and their ultimate cellular distribution by adding our chaperones into their experimental systems. We will also test the idea that the palmitoylated peptide motif works by specifically localizing to lipid rafts, microdomains in membranes which are greatly enriched in cholesterol and sphingolipids and appear to be used to assemble signaling complexes. Chaperones can only treat 20-50% of the mutations so for the remaining 50-80% of INCL patients we propose that hydrophobic thiols such as thiocholesterol could chemically facilitate hydrolysis of the storage material itself. Finally we will extend our approach to the most common form of Batten disease caused by mutations in the CLN2 gene (tripepfidyl-peptidase-1). Compound heterozygote patients with milder disease should benefit from chaperone therapy based on borate complexes of the inhibitor (AAFX) delivered to the CNS with our unique peptide sequences either directly or coated on quantum dots.

项目摘要（参见说明）：药理学伴侣（例如：AcGDap（Palm）VKIKK）可以被细胞内化并将错误折叠的蛋白质重新折叠成活性构型，但需要穿过血脑屏障，进入神经元并逃逸内体。我们已经证明，棕榈酰化肽基序独特地能够允许药物逃逸内体，并且这种伴侣可以重新激活错误折叠的蛋白质，例如棕榈酰蛋白硫酯酶(PPT1)。我们现在建议设计序列，通过附加荧光团和目标基序（例如：狂犬病病毒糖蛋白外壳肽（RVG））或涂覆在 6-1 Onm 量子点上，允许伴侣穿过血脑屏障。我们将测试脯氨酸和组氨酸的短序列，谷氨酰胺间隔基附着到用 4-硫醇 PEG 配体封端的 635 nm 红色量子点的表面。我们将在培养的出生后神经元和来自 PPT1 起源中具有明确点突变的患者的淋巴母细胞中对此进行测试，然后使用与 Project I 合作的 E3 胚胎鸡脊髓注射系统，或与 Project I 合作的大鼠海马切片系统二.这三个模型系统代表胚胎大脑、新生儿大脑和出生后大脑，通过将我们的伴侣添加到他们的实验系统中，合作将使我们能够更好地评估这些药物的毒性（如果有的话）、它们的功效和它们的最终细胞分布。我们还将测试这样的想法，即棕榈酰化肽基序通过特异性定位于脂筏、膜中的微结构域而发挥作用，这些微结构域富含胆固醇和鞘脂，似乎用于组装信号复合物。分子伴侣只能治疗 20-50% 的突变，因此对于剩余 50-80% 的 INCL 患者，我们建议疏水性硫醇（如硫代胆固醇）可以通过化学方式促进储存材料本身的水解。最后，我们将把我们的方法扩展到由 CLN2 基因（tripepfidyl-peptidase-1）突变引起的最常见的巴顿病。患有较轻疾病的复合杂合子患者应该受益于基于抑制剂硼酸盐复合物（AAFX）的伴侣治疗，通过我们独特的肽序列直接或涂覆在量子点上递送至中枢神经系统。