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.

开发用于治疗朊病毒病的降低朊病毒蛋白的二价 siRNA 疗法 项目概要/摘要 朊病毒病是一种致命的、无法治愈的、快速进展的神经退行性疾病，由单一蛋白质引起， 朊病毒蛋白或 PrP。在疾病中，PrP 转化为致病性“朊病毒”构象，并在各处传播 通过破坏其他 PrP 分子的构象来破坏大脑。这个过程严格依赖于 PrP 表达， PrP 的减少，无论是遗传上还是药理学上，剂量依赖性朊病毒都可以延缓疾病的发生 动物。至关重要的是，即使 PrP 持续降低 50%，所有动物最终也会死于致命的疾病。 疾病。 PrP 敲除模型表明，PrP 的完全消除可完全预防朊病毒病，并且 即使瞬时钳位抑制>90%也可以让大脑清除朊病毒，从而导致一次性 “治愈。”迄今为止，还没有可用的药物方式 a) 能够降低单一致病菌的水平 蛋白质，b) 活跃于整个大脑，并可实际递送至整个大脑，以及 c) 足以测试上述内容 药理学假设。然而，最近描述的二价 si-RNA (di-siRNA) 现在提供了第一个 这样的方式。 Di-siRNA 是经过化学修饰的连接 siRNA 双链体，能够到达整个啮齿动物 和非人类灵长类动物大脑在输送到脑脊液后，并降低特定致病水平 亨廷顿蛋白等蛋白质的含量>99%。针对小鼠和人类的 Di-siRNA 进行初步预筛选 PrP RNA 将由我们在麻省大学的合作者 Anastasia Khvorova 博士合成和提供。在R61中 在授予阶段，我们将： 1) 通过表征来验证细胞中 PrP 靶向 di-siRNA 分子的效力 小鼠和人类细胞中 PrP RNA 和蛋白质的敲低以及效果的剂量反应性。 2） 通过评估体内效力，表征未感染小鼠体内主要的 di-siRNA 体内 PK/PD 特性， 耐受性、RNA 和蛋白质减少之间的相关性、剂量反应性、大脑分布和时间 在野生型和“人源化”（表达人 PrP）转基因小鼠中进行清除。 3) 表征时间 通过感染新型转基因人源化小鼠，研究人源化小鼠的病理学和绝症 使用人类朊病毒建立模型并跟踪生物标志物变化、症状和终点的时间。在R33奖中 在这一阶段，我们将在感染小鼠和人类朊病毒的小鼠中进行生存实验，以评估 多种临床相关的治疗范例。 1) 预防性慢性病后的存活率 剂量。我们将通过启动慢性治疗来模拟有遗传性朊病毒病风险的个体的预防性治疗 在朊病毒感染前给予di-siRNA，并评估对生存的影响； 2) 单次剂量后的存活率 病理前时间点。我们将评估在朊病毒感染早期单次注射双 siRNA 的影响 对脑朊病毒滴度和存活率都有影响。 3) 在有症状的时间点单次给药后的存活率。我们 将通过在晚期疾病中提供单次推注剂量的 di-siRNA 来模拟症状阶段的治疗 时间点，并将评估对朊病毒滴度和生存的影响。这些目标共同决定 PrP- 降低di-siRNAs应推进临床用于预防和/或治疗人类朊病毒病。