Unfolded Protein Response as a Therapeutic Target for ADRP Animal Models

未折叠蛋白反应作为 ADRP 动物模型的治疗靶点

基本信息

批准号：
7948809
负责人：
Marina Gorbatyuk
金额：
$ 21.75万
依托单位：
UNIVERSITY OF NORTH TEXAS HLTH SCI CTR
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-30 至 2015-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7948809
关键词：
11 cis Retinal Accounting Affect Animal Model Antibodies Apoptosis Apoptotic Biological Preservation Blindness Caspase Cell Death Cessation of life Comparative Study Contrast Sensitivity Data Deterioration Development Disease Electroretinography Endoplasmic Reticulum Ethnic group Evaluation GADD45 Gene Transfer Genes Genetic Variation Goals Golgi Apparatus Inherited Investigational Drugs Knockout Mice Lead Location Measures Mediating Messenger RNA Methods Modeling Molecular Molecular Chaperones Monitor Mus Mutation Nucleosomes Opsin Optometry Pathogenesis Pathway interactions Patients Photoreceptors Proteins Rattus Resistance Retina Retinal Retinal Degeneration Retinitis Pigmentosa Reverse Transcriptase Polymerase Chain Reaction Rhodopsin Role Serotyping Severities Signal Transduction Small Interfering RNA Small RNA Structure Supplementation Techniques Testing Therapeutic Transgenic Mice Viral Virus Vision Visual impairment Vitamin A X inherited trait base biological adaptation to stress caspase 12 caspase-7 defined contribution design dosage drug discovery endoplasmic reticulum stress gene therapy improved functioning molecular chaperone GRP78 morphometry mouse model mutant novel programs promoter protein misfolding protein transport response success therapeutic target trafficking transcription factor CHOP vector

项目摘要

DESCRIPTION (provided by applicant): This project is focused on the elucidation of the role of the Unfolded Protein Response (UPR) in autosomal dominant retinitis pigmentosa (ADRP) pathogenesis and development of the gene therapy based on modulation of the UPR signaling markers. Retinitis pigmentosa (RP) is the most common inherited form of blindness, affecting about 1 in every 4000 people in all ethnic groups worldwide. RP can be transmitted either as an autosomal dominant (ADRP), autosomal recessive (ARRP), or X-linked trait. More than 100 mutations in rhodopsin account for approximately 30% of ADRP cases with varying severity of visual impairment. Misfolded opsin interferes with the trafficking of wild-type rhodopsin, accumulates in the endoplasmic reticulum (ER) and stimulates a signal transduction cascade known as the Unfolded Protein Response (UPR). If unchecked, this pathway triggers photoreceptor death, presumably through apoptosis. Although supplementation with vitamin A may be beneficial in some cases, currently, there is no effective pharmacological therapy for ADRP. Therefore, the major objective of this proposal is to determine whether the gene therapy based on the re-programming of the ER stress response caused by aberrant rhodopsin is a viable treatment, unlimited by different localizations of rhodopsin mutations (P23H and T17M). In two mouse models of ADRP, we plan to reprogram the ER stress signaling by viral delivery of the molecular chaperone GRP78/BiP and delivery of small interfering siRNAs targeting caspase-7, caspase-12 and pro-apoptotic CHOP/GADD153 mRNAs to diminish the level of apoptosis in ADRP photoreceptors. For each ADRP model, we plan to: (1) modulate the UPR in favor of activation of pro-survival pathway by over-expression of BiP protein; (2) suppress apoptosis by diminishing levels of activated caspase-7 and caspase-12 and (3) inhibit the CHOP-associated apoptosis by targeting CHOP mRNA. We will monitor survival of photoreceptors using electroretinography and morphometry and will measure the activation of the ER stress and apoptosis using specific antibodies and RT-PCR. We will also measure improvement in vision using Optometry, a technique that can evaluate both acuity and contrast sensitivity in mice. We anticipate that the success of this approach will also require the appropriate combination of AAV serotype, vector dosage, photoreceptor specific promoter and optimized expression for the chaperone BiP. While AAV mediated gene transfer is being developed for treatment of RP, the suppression of ER stress and of apoptosis using chaperones is novel. This approach may overcome the genetic diversity of this disease and reveal the pathways of cell death that lead from mutation to retinal degeneration. PUBLIC HEALTH RELEVANCE: Our goal is to design gene therapy for blinding disorder known as Autosomal Dominant Retinitis Pigmentosa (ADRP). We plan to study cellular mechanisms involved in photoreceptor death and develop a gene therapy based on the blockage of apoptosis in ADRP photoreceptors. Using a harmless virus, we plan to deliver molecular chaperon BiP and small RNA molecules to improve function and structure of photoreceptors and also intend to study the mechanism by which these therapeutic molecules provide therapy.

描述（由申请人提供）：该项目的重点是阐明未折叠的蛋白质反应（UPR）在基因治疗基于UPR信号标记的调节基础上，基因治疗的发育和发育。色素性视网膜炎（RP）是最常见的失明形式，在全球所有种族中每4000人中约有1人。 RP可以作为常染色体显性剂（ADRP），常染色体隐性（ARRP）或X连锁性状传输。在视紫红质中，超过100个突变占ADRP病例的30％，视力障碍严重程度。错误折叠的OPSIN干扰了野生型视紫红质的运输，积聚在内质网中（ER），并刺激被称为展开的蛋白质反应（UPR）的信号转导级联。如果不选中，该途径会触发感光器死亡，大概是通过细胞凋亡。尽管在某些情况下补充维生素A可能是有益的，但目前尚无有效的ADRP药理疗法。因此，该提案的主要目的是确定基于异常的视紫红蛋白引起的ER应激反应的基因治疗是否是可行的治疗方法，是一种可行的治疗方法，无限制地被视紫红质突变的不同局部化（p23h和t17m）。在ADRP的两种鼠标模型中，我们计划通过病毒递送分子伴侣GRP78/BIP以及针对Caspase-7的小型干扰siRNA，CASPASE-12，CASPASE-12和Pro-Apoptotic Chop/gadd153 mRNAS的小型干扰siRNA的递送来重新编程ER应力信号传导。对于每个ADRP模型，我们计划：（1）通过过表达BIP蛋白来调节UPR，以激活促生物途径；（2）通过降低活化的caspase-7和caspase-12的水平来抑制凋亡，以及（3）通过靶向CHOP mRNA抑制与斩波相关的凋亡。我们将使用电视图和形态计量光感受器的生存率，并使用特定的抗体和RT-PCR测量ER应力和凋亡的激活。我们还将使用验光测量视力的改进，该技术可以评估小鼠的敏锐度和对比度灵敏度。我们预计，这种方法的成功还将需要AAV血清型，载体剂量，感光受体特异性启动子和优化伴侣BIP的表达的适当组合。虽然正在开发AAV介导的基因转移以治疗RP，但使用伴侣蛋白抑制ER应激和凋亡的抑制是新颖的。这种方法可能会克服该疾病的遗传多样性，并揭示从突变到视网膜变性导致的细胞死亡途径。公共卫生相关性：我们的目标是针对被称为常染色体显性视网膜炎色素炎（ADRP）设计基因疗法。我们计划研究涉及光感受器死亡的细胞机制，并基于ADRP光感受器的细胞凋亡的阻塞而发展基因治疗。使用无害病毒，我们计划提供分子伴侣BIP和小的RNA分子，以改善感光体的功能和结构，并打算研究这些治疗分子提供治疗的机制。