Development of Prion Protein-Lowering Divalent siRNA Therapy for Prion Disease

开发针对朊病毒病的降低朊病毒蛋白的二价 siRNA 疗法

• 批准号: 10549815
• 负责人: Eric Vallabh Minikel
• 金额: $ 38.77万
• 依托单位: BROAD INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-01 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10549815
• 关键词: Ablation; Aftercare; Agreement; Animal Model; Animals; Antisense Oligonucleotides; Award; Bilateral; Biochemical; Bioinformatics; Biological Assay; Biological Markers; Bolus Infusion; Brain; Brain Diseases; Cell Culture Techniques; Cells; Chemicals; Chronic; Clinic; Closure by clamp; Collaborations; Data Collection; Development; Diagnosis; Disease; Dose; Etiology; Genes; Genetic; Genetic Risk; Goals; Heterozygote; Human; Human Cell Line; Huntington gene; Individual; Infection; Knock-out; Link; Modality; Modeling; Molecular Conformation; Mouse Cell Line; Mus; Natural History; Neurodegenerative Disorders; Neurons; Pathogenicity; Pathologic; Pathology; Pharmaceutical Preparations; Phase; PrP; PrP gene; Predisposition; Prevention; Preventive treatment; Prion Diseases; Prions; Process; Property; Proteins; RNA; Rodent; Schedule; Small Interfering RNA; Symptoms; Technology; Terminal Disease; Testing; Therapeutic; Time; Transgenic Mice; Transgenic Organisms; Wild Type Mouse; advanced disease; clinical subtypes; clinically relevant; cohort; cross reactivity; experimental study; humanized mouse; in vivo; in vivo evaluation; knock-down; model organism; mouse model; neuropathology; non-genetic; nonhuman primate; novel; patient registry; pharmacokinetics and pharmacodynamics; pharmacologic; prevent; prion seeds; prophylactic; protein expression; research clinical testing; response; therapeutic siRNA; translational pipeline

Development of prion protein-lowering divalent siRNA therapy for prion disease Project Summary / Abstract Prion disease is a fatal, untreatable, rapidly progressive neurodegenerative disease caused by a single protein, the prion protein or PrP. In disease, PrP converts into a pathogenic “prion” conformation that spreads across the brain by conformationally corrupting other PrP molecules. This process is strictly reliant on PrP expression, and reduction of PrP, either genetically or pharmacologically, dose-dependently prion delays disease in animals. Critically, even with sustained 50% knockdown of PrP, all animals eventually succumb to fatal disease. PrP knockout models indicate that total ablation of PrP is fully protective against prion disease, and that even transient clamp suppression of >90% could allow the brain to clear prions, resulting in a one-time “cure.” To date, no drug modality has been available that is a) capable of reducing levels of a single pathogenic protein, b) active in, and realistically deliverable to, the whole brain, and c) potent enough to test the above hypothesis pharmacologically. However, recently described divalent si-RNAs (di-siRNAs) now offer the first such modality. Di-siRNAs are chemically modified linked siRNA duplexes capable of reaching the whole rodent and non-human primate brain following delivery into CSF, and reducing levels of specific disease-causing proteins such as Huntingtin by >99%. Di-siRNAs preliminarily pre-screened against both mouse and human PrP RNA will be synthesized and provided by our collaborator Dr. Anastasia Khvorova at UMass. In the R61 award phase, we will: 1) Validate potency of PrP-targeting di-siRNA molecules in cells, by characterizing PrP RNA and protein knockdown, and dose-responsiveness of effect, in mouse and human cells. 2) Characterize leading di-siRNA in vivo PK/PD properties in uninfected mice, by assessing in vivo potency, tolerability, correlation between RNA and protein reduction, dose-responsiveness, brain distribution, and time to washout in wild-type and “humanized” (human-PrP expressing) transgenic mice. 3) Characterize time to pathology and terminal disease in humanized mice, by infecting a novel transgenic humanized mouse model with human prions and tracking time to biomarker changes, symptoms and endpoint. In the R33 award phase, we will perform survival experiments in mice infected with both mouse and human prions to assess multiple clinically relevant treatment paradigms. 1) Survival following prophylactically initiated chronic dosing. We will model preventive treatment of individuals at risk for genetic prion disease by initiating chronic di-siRNA dosing before prion infection, and assess impact on survival; 2) Survival following a single dose at a pre-pathological timepoint. We will assess the impact of a single bolus of di-siRNA early in prion infection on both brain prion titer and survival. 3) Survival following a single dose at a symptomatic timepoint. We will model symptomatic-stage treatment by delivering a single bolus dose of di-siRNA at an advanced disease timepoint, and will assess impact on prion titer and survival. Together, these aims will determine whether PrP- lowering di-siRNAs should be advanced to the clinic for prevention and/or treatment of human prion disease.

PrP病的二价siRNA治疗研究进展 项目摘要/摘要 Pron病是一种致命的、无法治愈的、进展迅速的神经退行性疾病，由单一蛋白质引起， Prion蛋白质或PrP。在疾病中，PrP转化为一种致病的“普里恩”构象，并在 通过构象破坏其他PrP分子来影响大脑。这个过程严格依赖于PrP的表达， 而PrP的减少，无论是在遗传上还是在药物上，都以剂量依赖的方式延缓了疾病的发生 动物。关键是，即使PrP持续50%被击倒，所有动物最终都会死于 疾病。PrP基因敲除模型表明，完全消融PrP对PrP疾病具有完全保护作用，并且 即使是短暂的抑制90%的钳制也可以允许大脑清除普恩病毒，导致一次性 “治愈。”到目前为止，还没有能够降低单一致病物质水平的药物模式可用 蛋白质，b)活跃在整个大脑中，并可实际输送到整个大脑，以及c)足够强大，足以测试上述 药理学假说。然而，最近描述的二价siRNA(di-siRNA)现在提供了第一个 这样的情态。Di-siRNA是一种化学修饰的连接的siRNA双链，能够到达整个啮齿动物 和非人类灵长类动物的大脑进入脑脊液，并降低特定致病水平 亨廷顿等蛋白质含量高达99%。小鼠和人双小干扰RNA的初步筛选 PRP RNA将由我们的合作者阿纳斯塔西娅·赫沃洛娃博士在马萨诸塞州大学合成并提供。在R61公路上 获奖阶段，我们将：1)验证PrP靶向细胞中di-siRNA分子的效力，通过表征 在小鼠和人类细胞中，PRP RNA和蛋白质被击倒，以及效应的剂量-反应。2) 通过评估体内效力，鉴定未感染小鼠体内领先的di-siRNA PK/PD特性， 耐受性、RNA和蛋白质减少之间的相关性、剂量反应、脑分布和时间 在野生型和人源化(表达人PrP的)转基因小鼠中被淘汰。3)描述时间以 通过感染新型转基因人源化小鼠，人源化小鼠的病理和终末期疾病 用人类普恩病毒建立模型，跟踪生物标记物的变化、症状和终点的时间。在R33奖项中 阶段，我们将在感染了小鼠和人类普恩病毒的小鼠身上进行存活实验，以评估 多种临床相关的治疗范例。1)预防启动慢性阻塞性肺疾病后的生存 给药。我们将通过启动慢性慢性病来模拟对有遗传性普恩病毒疾病风险的个人进行预防性治疗 Pron感染前的Di-siRNA剂量，并评估对生存的影响；2)单次注射后的生存 一个病态前的时间点。我们将评估单剂双干扰核糖核酸在普恩病毒感染早期的影响。 对脑蛋白滴度和存活率都有影响。3)在出现症状的时间点单次给药后的存活率。我们 将模拟症状阶段的治疗，在晚期疾病中提供单剂量的di-siRNA 时间点，并将评估对病毒滴度和存活率的影响。这些目标加在一起将决定PrP是否- 应将降低disiRNAs应用于预防和/或治疗人类Pron病的临床。