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

• 批准号: 8575057
• 负责人: Marina Gorbatyuk
• 金额: $ 24.13万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-03-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8575057
• 关键词: Unfolded; Protein; Response; Therapeutic; Target

Unfolded Protein Response as a Therapeutic Target for ADRP Animal Models 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.

展开的蛋白质反应是ADRP动物模型的治疗靶标 该项目的重点是阐明未折叠蛋白反应（UPR）在常染色体中的作用 基于基因治疗的显性视网膜炎色素（ADRP）的发病机理和基于基因治疗的发育 UPR信号标记的调制。色素性视网膜炎（RP）是最常见的遗传形式 失明，全世界所有种族中每4000人中约有1人。 RP可以传输 作为常染色体显性剂（ADRP），常染色体隐性（ARRP）或X连锁性状。超过100个突变 视视占ADRP病例的约30％，视觉障碍的严重程度不同。错误地折 OPSIN干扰野生型Rhodopsin的贩运，积聚在内质网（ER）和 刺激称为展开的蛋白质反应（UPR）的信号转导级联。如果没有选中，这 途径触发了光感受器死亡，大概是通过细胞凋亡。虽然补充维生素A 在某些情况下，可能是有益的，目前尚无对ADRP的有效药理疗法。所以， 该提案的主要目的是确定是否基于重新编程的基因疗法 异常的视紫红蛋白引起的ER应力反应是一种可行的治疗方法，无限。 视紫红质突变（p23h和t17m）。 在ADRP的两个小鼠模型中，我们计划通过病毒输送来重新编程ER应力信号传导 分子伴侣GRP78/BIP和针对caspase-7，caspase-12的小干扰siRNA的递送 促凋亡的CHOP/GADD153 mRNA可降低ADRP光感受器的凋亡水平。为了 每个ADRP模型，我们都计划：（1）调节UPR，以超过 - BIP蛋白的表达； （2）通过降低活化的caspase-7和caspase-12抑制凋亡 （3）通过靶向CHOP mRNA抑制与斩波相关的凋亡。我们将监控生存 使用视网膜图和形态计量学的感光体，将测量ER应力的激活 使用特定抗体和RT-PCR的凋亡。我们还将使用 验光，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，向量剂量的适当组合， 伴侣BIP的光感受器特异性启动子和优化表达。而AAV介导的基因 正在开发转移用于治疗RP，抑制ER应力和使用的凋亡 伴侣是新颖的。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细胞死亡的途径。