CRISPR/Cas9-based gene therapy for Angelman syndrome
基于 CRISPR/Cas9 的 Angelman 综合征基因疗法
基本信息
- 批准号:10490828
- 负责人:
- 金额:$ 54.07万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-09-15 至 2024-08-31
- 项目状态:已结题
- 来源:
- 关键词:AgeAllelesAngelman SyndromeAntisense OligonucleotidesBehavioralBindingBiodistributionBiologyBrainCRISPR/Cas technologyCellsChildChildhoodClinicClustered Regularly Interspaced Short Palindromic RepeatsDataDependovirusFunctional disorderGene ExpressionGene Transduction AgentGenesGenetic TranscriptionGuide RNAHumanInjectionsInterventionIntronsKnowledgeLightLongevityMediatingMedicalModelingMusMutagenesisMutationNeurodevelopmental DisorderNeuronsPhenotypePilot ProjectsPre-Clinical ModelPrimatesRNAResearchSiteSmall Nucleolar RNAStaphylococcus aureusStreptococcus pyogenesTestingTherapeuticTimeTopoisomerase InhibitorsTopotecanToxic effectTranslatingUBE3A geneUntranslated RNAVariantadeno-associated viral vectorautism onsetautism spectrum disorderbasebehavioral phenotypingdesigneffective therapyefficacy evaluationefficacy validationgene repressiongene therapyin vivoinnovationmammalian genomemouse modelnovel strategiespre-clinicalrelating to nervous systemside effecttherapeutic candidateubiquitin-protein ligase
项目摘要
PROJECT SUMMARY
Angelman syndrome (AS) is a severe neurodevelopmental disorder caused by deletion or mutation of the
maternal allele of UBE3A. UBE3A is biallelically expressed in nearly all cells of the body except in mature
neurons, where the paternal allele is silenced by an extremely long non-coding RNA called UBE3A-ATS. In
light of this biology, the most direct way to treat behavioral dysfunctions associated with AS is to unsilence the
intact paternal UBE3A allele. CRISPR/Cas9 technology can be used to target specific regions of the
mammalian genome for mutagenesis or transcriptional repression. In unpublished studies, we generated
hundreds of S. pyogenes (Sp)Cas9 guide RNAs (gRNAs) that target regions throughout UBE3A-ATS. Several
of these gRNAs, when transfected along with SpCas9, potently unsilenced paternal Ube3a in cultured mouse
cortical neurons. Some of our most effective gRNAs targeted a region of Ube3a-ATS that is conserved
between mice and humans, making it possible to translate our findings to human neurons. Here, we will test
the central hypothesis that CNS-directed delivery of Cas9 and a gRNA that targets Ube3a-ATS can
enduringly unsilence paternal UBE3A and treat behavioral phenotypes associated with Angelman
syndrome. We will use adeno-associated virus (AAV) for delivery because it can drive gene expression for
years in the brain. Pilot studies with S. aureus (Sa)Cas9, a smaller Cas9 variant, suggest that our gene
therapy approach can be used to unsilence paternal Ube3a in mice for at least three months. To advance this
innovative gene therapy towards the clinic, we will evaluate efficacy, on- and off-target effects, and mechanism
of action of candidate therapeutic SaCas9 gRNAs that target Ube3a-ATS. We will use cultured neurons from
AS model mice and AS-derived human neurons. We will package SaCas9 and an optimized gRNA into a
single AAV vector, and then evaluate unsilencing efficacy and longevity for up to two years in mice, as well as
biodistribution and toxicity. Lastly, we will evaluate the extent to which AAV-mediated delivery of this
CRISPR/Cas9-based gene therapy treats behavioral phenotypes in AS model mice.
项目总结
Angelman综合征(AS)是由基因缺失或突变引起的一种严重的神经发育障碍
UBE3A的母系等位基因。UBE3A在体内除成熟细胞外的几乎所有细胞中双等位表达
其中父亲的等位基因被一种名为UBE3A-ATS的超长非编码RNA沉默。在……里面
从这一生物学角度来看,治疗与强直性脊柱炎相关的行为障碍的最直接方法是解除沉默
完整的父系UBE3A等位基因。CRISPR/Cas9技术可用于针对
用于突变或转录抑制的哺乳动物基因组。在未发表的研究中,我们产生了
数百个化脓性链球菌(Sp)Cas9指导RNAs(GRNAs),靶向整个UBE3A-ATS区域。几个
在这些gRNA中,当与SpCas9一起转染时,在培养的小鼠中有效地沉默了父亲的Ube3a
皮质神经元。我们的一些最有效的gRNA针对的是Ube3a-ATS的保守区域
在老鼠和人类之间,使我们的发现有可能转化为人类神经元。在这里,我们将测试
中枢假设,即CNS导向的Cas9和靶向Ube3a-ATS的gRNA可以
持久不让父亲UBE3A沉默并治疗与Angelman相关的行为表型
综合症。我们将使用腺相关病毒(AAV)进行传递,因为它可以驱动基因表达
在大脑里呆了几年。对金黄色葡萄球菌(Sa)Cas9的初步研究表明,我们的基因
治疗方法可用于解除小鼠父亲Ube3a至少三个月的沉默。为了推进这一进程
创新的基因治疗将走向临床,我们将评估疗效、靶上和靶外效应以及机制
靶向Ube3a-ATS的候选治疗性SaCas9 gRNA的作用。我们将使用培养的神经元
作为模型小鼠和AS来源的人类神经元。我们将把SaCas9和优化后的gRNA打包成
单个AAV载体,然后评估小鼠的去沉默效果和寿命长达两年,以及
生物分布和毒性。最后,我们将评估AAV介导的交付在多大程度上
基于CRISPR/Cas9的基因疗法治疗AS模型小鼠的行为表型。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Mark J. Zylka其他文献
The environmental neuroactive chemicals list of prioritized substances for human biomonitoring and neurotoxicity testing: A database and high-throughput toxicokinetics approach
- DOI:
10.1016/j.envres.2024.120537 - 发表时间:
2025-02-01 - 期刊:
- 影响因子:
- 作者:
Julia E. Rager;Lauren E. Koval;Elise Hickman;Caroline Ring;Taylor Teitelbaum;Todd Cohen;Giulia Fragola;Mark J. Zylka;Lawrence S. Engel;Kun Lu;Stephanie M. Engel - 通讯作者:
Stephanie M. Engel
Correction to: Chd8 haploinsufficiency impairs early brain development and protein homeostasis later in life
- DOI:
10.1186/s13229-021-00438-6 - 发表时间:
2021-05-08 - 期刊:
- 影响因子:5.500
- 作者:
Jessica A. Jiménez;Travis S. Ptacek;Alex H. Tuttle;Ralf S. Schmid;Sheryl S. Moy;Jeremy M. Simon;Mark J. Zylka - 通讯作者:
Mark J. Zylka
Mark J. Zylka的其他文献
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{{ truncateString('Mark J. Zylka', 18)}}的其他基金
Development of a deep neural network to measure spontaneous pain from mouse facial expressions
开发深度神经网络来测量小鼠面部表情的自发疼痛
- 批准号:
10094266 - 财政年份:2020
- 资助金额:
$ 54.07万 - 项目类别:
Development of a deep neural network to measure spontaneous pain from mouse facial expressions
开发深度神经网络来测量小鼠面部表情的自发疼痛
- 批准号:
10579988 - 财政年份:2020
- 资助金额:
$ 54.07万 - 项目类别:
Development of a deep neural network to measure spontaneous pain from mouse facial expressions
开发深度神经网络来测量小鼠面部表情的自发疼痛
- 批准号:
10717670 - 财政年份:2020
- 资助金额:
$ 54.07万 - 项目类别:
Development of a deep neural network to measure spontaneous pain from mouse facial expressions
开发深度神经网络来测量小鼠面部表情的自发疼痛
- 批准号:
10349447 - 财政年份:2020
- 资助金额:
$ 54.07万 - 项目类别:
CRISPR/Cas9-based gene therapy for Angelman syndrome
基于 CRISPR/Cas9 的 Angelman 综合征基因疗法
- 批准号:
10237150 - 财政年份:2019
- 资助金额:
$ 54.07万 - 项目类别:
Environmental-use chemicals that target pathways linked to autism and other neurodevelopmental disorders
针对与自闭症和其他神经发育障碍相关途径的环境使用化学品
- 批准号:
10402265 - 财政年份:2019
- 资助金额:
$ 54.07万 - 项目类别:
Environmental-use chemicals that target pathways linked to autism and other neurodevelopmental disorders
针对与自闭症和其他神经发育障碍相关途径的环境使用化学品
- 批准号:
10618242 - 财政年份:2019
- 资助金额:
$ 54.07万 - 项目类别:
CRISPR/Cas9-based gene therapy for Angelman syndrome
基于 CRISPR/Cas9 的 Angelman 综合征基因疗法
- 批准号:
10011898 - 财政年份:2019
- 资助金额:
$ 54.07万 - 项目类别:
Identification of candidate environmental risks for autism
识别自闭症的候选环境风险
- 批准号:
9525549 - 财政年份:2017
- 资助金额:
$ 54.07万 - 项目类别:
Lipid kinase regulation of pain signaling and sensitization
脂质激酶对疼痛信号传导和敏化的调节
- 批准号:
9279273 - 财政年份:2013
- 资助金额:
$ 54.07万 - 项目类别:
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