Improving AAV-based gene therapy for Krabbe Disease

改进基于 AAV 的克拉伯病基因疗法

• 批准号: 10512038
• 负责人: Gregory J Heller
• 金额: $ 5.18万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10512038
• 关键词: 3 year old; Adult; Affect; Aftercare; Age; Animals; Architecture; Behavioral; Biochemical; Birth; Brain; Capsid Proteins; Catabolism; Cell membrane; Cells; Cessation of life; Characteristics; Clinical; Clinical Trials; Complementary DNA; DNA; Data; Demyelinations; Dependovirus; Development; Developmental Delay Disorders; Diffuse; Disease; Disease Progression; Dose; Effectiveness; Elderly; Enzymes; Episome; Failure to Thrive; Fluorescent in Situ Hybridization; Genes; Globoid cell leukodystrophy; Goals; Hematopoietic Stem Cell Transplantation; Histologic; Histopathology; Immunohistochemistry; Impairment; Infant; Inherited; Intrathecal Injections; Intravenous; Investigation; Lesion; Life; Life Expectancy; Longevity; Measures; Mendelian disorder; Mission; Modality; Modification; Mus; Mutation; Neonatal; Nerve Degeneration; Neurons; Oligodendroglia; Paralysed; Pathologic; Patients; Protocols documentation; Psychosine; Quality of life; Recurrence; Research Personnel; Signal Pathway; Teenagers; Testing; Therapeutic; Time; Tissues; Treatment Efficacy; Treatment Protocols; United States National Institutes of Health; Viral Genome; Wild Type Mouse; adeno-associated viral vector; aged; base; burden of illness; cell type; chronic neurologic disease; clinically relevant; design; dosage; end of life; exhaustion; experimental study; functional restoration; galactosylceramidase; gene therapy; immunoreaction; improved; leukodystrophy; mouse genome; mouse model; oligodendrocyte precursor; preclinical trial; prevent; stem cells; success; treatment duration; vector; viral DNA; white matter

Abstract Krabbe’s Disease (KD) is a rare inherited leukodystrophy caused by mutations in the GALC gene encoding the lysosomal enzyme, galactosylceramidase (GALC). Lack of functional GALC leads to global demyelination and neurodegeneration. Untreated KD presents in infants as developmental delay and regression, failure to thrive, and ultimately death by 2-3 years of age. KD is treated with hematopoietic stem cell transplantation (HSCT) which slows the progression of disease and prolongs life expectancy into the teenage years, but it is not a cure. Adeno-associated viruses (AAV) have been utilized successfully as gene therapy treatment vectors in numerous pre-clinical and clinical trials. The initial success of AAV gene therapy in diseases primarily affecting neuronal cells led to investigations of their efficacy in monogenic diseases affecting multiple cells types such as KD. The Bongarzone lab previously developed a treatment protocol using AAV9- GALC to correct the deficiency in the Twitcher murine model of KD. This treatment completely prevented disease development for the first 6-8 months of life, such that AAV-treated mice were nearly indistinguishable from wild-type mice. Despite these promising results, our study also revealed a slow decline in treatment efficacy over time, including development of disease signs and focal demyelinating plaques in mice of advanced age; similar findings have been noted by other investigators as well. No alternative modifications to improve gene therapy have been proposed thus far. The overarching goal of this application is to establish whether gene therapy efficacy is declining due to exhaustion of AAV DNA in the adult brain and how to optimize gene therapy treatment efficacy and duration in Twitcher mice. As AAV-GALC DNA exists as a non-replicating extra-chromosomal episome after entering a cell, we hypothesize the decline in treatment efficacy is caused by loss of therapeutic vector in replicating cells, particularly oligodendrocyte precursors, in the Twitcher brain, which leads to a decrease in the average AAV- GALC episomes per cell over time. This hypothesis will be investigated by treating mice shortly after birth with AAV9-GALC at decreasing dosages and then observing how long the treatment is efficacious based on clinical score, survival, as well as biochemical and histological analyses. We will then examine if redosing utilizing oligodendrocyte targeted AAV (AAV-001) later in life, but prior to disease sign onset, delays or prevents development of disease signs. Finally, we will determine if utilizing AAV-001 singly will increase the efficacy and duration of treatment in comparison to AAV9. This study is important because we need to better understand what happens to episomal AAV DNA long term before attempting to use AAV based therapies to treat KD patients. Furthermore, our findings will have the intrinsic impact of better characterizing how episomal AAV DNA behaves in similar monogenic disorders.

抽象的 克拉伯氏病 (KD) 是一种罕见的遗传性脑白质营养不良，由 GALC 基因突变引起 编码溶酶体酶，半乳糖神经酰胺酶（GALC）。缺乏功能性 GALC 导致全球 脱髓鞘和神经变性。未经治疗的川崎病在婴儿中表现为发育迟缓和 退化，发育迟缓，最终在 2-3 岁时死亡。造血干治疗川崎病 细胞移植（HSCT）可减缓疾病进展并延长预期寿命 青少年时期，但这不是治愈方法。腺相关病毒（AAV）已成功用作基因 大量临床前和临床试验中的治疗载体。 AAV基因治疗取得初步成功 主要影响神经元细胞的疾病导致了对其在影响单基因疾病中的功效的研究 多种细胞类型，例如 KD。 Bongarzone 实验室之前开发了一种使用 AAV9 的治疗方案- GALC 可纠正 Twitcher 小鼠 KD 模型中的缺陷。这种治疗完全避免了 生命前 6-8 个月的疾病发展，使得 AAV 治疗的小鼠几乎无法区分 来自野生型小鼠。尽管取得了这些有希望的结果，但我们的研究也显示治疗效果缓慢下降 随着时间的推移，疗效，包括小鼠疾病体征和局灶性脱髓鞘斑块的发展 高龄;其他研究人员也注意到了类似的发现。没有替代修改 迄今为止已提出改进基因治疗。 本申请的总体目标是确定基因治疗功效是否因以下原因而下降： 成人大脑中 AAV DNA 的耗尽以及如何优化基因治疗的疗效和持续时间 抽搐老鼠。由于AAV-GALC DNA进入细胞后作为非复制的染色体外附加体存在 细胞，我们假设治疗效果的下降是由于复制细胞中治疗载体的丢失造成的， 特别是 Twitcher 大脑中的少突胶质细胞前体，这会导致平均 AAV- 减少 随着时间的推移，每个细胞的 GALC 附加体。这一假设将通过在出生后不久对小鼠进行治疗来进行研究 AAV9-GALC 逐渐减少剂量，然后根据临床观察治疗有效时间 评分、生存率以及生化和组织学分析。然后我们将检查是否使用重剂量 少突胶质细胞在生命后期、但在疾病症状出现之前靶向 AAV (AAV-001)，延迟或预防 疾病体征的发展。最后，我们将确定单独使用AAV-001是否会提高功效 以及与 AAV9 相比的治疗持续时间。这项研究很重要，因为我们需要更好地 在尝试使用基于 AAV 的疗法之前，了解游离型 AAV DNA 长期会发生什么 治疗 KD 患者。此外，我们的研究结果将对更好地描述游离态如何产生内在影响。 AAV DNA 在类似的单基因疾病中表现良好。