Novel platform for optimizing AAV transgene expression to improve efficacy of ocular gene therapies

用于优化 AAV 转基因表达以提高眼部基因治疗功效的新平台

• 批准号: 10385010
• 负责人: Matthew Louis Hirsch
• 金额: $ 26.34万
• 依托单位: EPIGENOS BIOSCIENCE, INC.
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10385010
• 关键词: Binding; Biological Sciences; Biomedical Engineering; CXCR4 gene; Cell Culture Techniques; Cells; Chemicals; Chimeric Proteins; Collaborations; Competitive Binding; Cornea; Couples; Dependovirus; Disease; Dose; Ensure; Enzymes; Epigenetic Process; Episome; FDA approved; FK506; Feasibility Studies; Formulation; Future; Gene Delivery; Gene Expression; Genetic Diseases; Genetic Transcription; Genome; Graft Rejection; Histones; Human; In Vitro; Keratoplasty; Licensing; Mediating; Methods; Modeling; Pathway interactions; Phase; Prevention; Production; Proteins; Regulation; System; Tacrolimus Binding Proteins; Technology; Therapeutic; Tissues; Transcription Coactivator; Transcription Repressor; Transcriptional Regulation; Transgenes; Transgenic Organisms; Transplantation Surgery; Validation; Zinc Fingers; adeno-associated viral vector; base; clinical application; design; epigenetic regulation; epigenetic silencing; experimental study; gene repression; gene therapy; improved; inhibitor; lead candidate; monomer; nervous system disorder; novel; novel therapeutics; promoter; recruit; small molecule; therapeutic gene; therapy development; transgene expression; vector; vector genome

Abstract Gene therapy applications based on adeno-associated virus (AAV) have demonstrated promise in clinical applications for the treatment of diverse genetic diseases highlighted by the recent FDA approval of AAV vector formulations as new drugs for ocular and neurological diseases. In these instances, along with all applications of clinical AAV gene therapy to date, constitutive transcription of the therapeutic cassette is employed without any safeguards in place to modulate transgene production in the targeted tissue. At the mechanistic level, the AAV vector transduction pathway is not well understood. The episomal AAV vector genomes form circular monomers and concatemers and a limited number of studies have demonstrated their associations with histones, transcriptional activators and repressors implying an untapped level of control related to overall transgene production. These observations suggest that AAV episomes may be restricted for maximal expression and allude to the ability for targeted approaches for modulating their epigenetic composition to enhance and/or repress AAV vector transduction. We recently developed a technology that couples a dCas9-based protein targeting system at a promoter with a small bifunctional molecule to control gene expression. Spefically, the dCas9-FKBP system was used to successfully activate epigenetically-silenced endogenous loci (e.g., MyoD1 and CXCR4) in HEK293 cells. Epigenos Biosciences proposes adding a new functionality, referred as CEMtrol, to AAV modules that will be designed to mitigate epigenetic dampening of transgene expression to ensure the efficacy of gene therapies will not be subject to uncontrollable epigenetic regulation. In collaboration with Dr. Matthew Hirsch, who is developing therapies for ocular diseases, Phase I will focus on developing an AAV that maximizes transgene expression in cell culture and explant cornea models as a proof-of-concept feasibility study.

摘要 基于腺相关病毒的基因治疗在临床上显示出良好的应用前景 FDA最近批准AAV载体在治疗多种遗传病方面的应用 作为治疗眼科和神经系统疾病的新药的制剂。在这些情况下，以及所有应用程序 到目前为止，在临床AAV基因治疗中，治疗盒的结构性转录没有被使用 是否有任何适当的保障措施来调节目标组织中的转基因生产。在机械层面上， AAV载体转导途径尚不清楚。表型AAV载体基因组呈环状 单体和串联体和有限数量的研究已经证明了它们与组蛋白的关联， 转录激活因子和抑制因子暗示与整体转基因相关的未被利用的控制水平 制作。这些观察表明，AAV的表型可能受到最大限度的表达和暗示的限制 有针对性地调节其表观遗传成分以增强和/或抑制AAV的能力 载体转导。 我们最近开发了一种技术，将启动子上基于dCas9的蛋白质靶向系统与 控制基因表达的双功能小分子。具体地说，dCas9-FKBP系统用于 在HEK293细胞中成功激活表观遗传沉默的内源性基因座(如MyoD1和CXCR4)。 Eigenos Biosciences建议向AAV模块添加一种名为CEMtrol的新功能，该功能将被 旨在减轻表观遗传抑制转基因表达，以确保基因治疗的有效性 不受不可控的表观遗传调控。与马修·赫希博士合作，马修·赫希博士正在开发 眼科疾病的治疗，第一阶段将专注于开发一种最大化转基因表达的AAV 细胞培养和角膜移植模型作为概念验证的可行性研究。