Glia Exclusive Gene Therapy

胶质细胞独家基因疗法

• 批准号: 10739502
• 负责人: Or Shemesh
• 金额: $ 23.85万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-15 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10739502
• 关键词: 3' Untranslated Regions; 5' Untranslated Regions; Acute; Address; Affect; Alzheimer' s Disease; Antibodies; Astrocytes; Binding; Biomedical Engineering; Brain; Brain Diseases; Cells; Cholesterol; Clinical; Code; Dependovirus; Development; Disease; Engineering; Ependymal Cell; Exhibits; Gene Delivery; Gene Expression; Gene Targeting; Gene Transduction Agent; Genes; Green Fluorescent Proteins; Hela Cells; Human; Image; Immune response; Intervention; Knock-out; Lentivirus; Lipids; Luciferases; Luminescent Proteins; Malignant Neoplasms; Malignant neoplasm of brain; Medical; Methods; Microglia; Modeling; Modification; Molecular; Mood Disorders; Mus; Mutagenesis; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neuroglia; Neurons; Oligodendroglia; Optics; Pain; Pain Disorder; Pathology; Play; Poly(A) Tail; Process; Proteins; RNA; RNA Stability; Rapid screening; Reporter Genes; Research; Role; Specificity; Surface Antigens; Tail; Technology; Testing; Therapeutic; Time; Vaccination; Vascular Diseases; Viral Vector; Virus; analog; aptamer; autism spectrum disorder; cancer therapy; cell type; clinical application; delivery vehicle; design; gene therapy; healing; immunogenicity; in vivo; interest; lipid nanoparticle; nervous system disorder; non-viral gene delivery; novel; prevent; screening; targeted delivery; transmission process; vector

Abstract Glia are supportive cells in the human brain, comprising microglia, oligodendrocytes, astrocytes, and ependymal cells. Glia are deeply involved in diseases of the nervous system such as Alzheimer’s (AD), autism, pain, affective disorders, and cancers. Different glial cell types play different mechanistic roles in disease formation, driven by specific genes. Modulating glial gene expression via a process called gene therapy could thus be studied as a means of preventing deleterious effects of glia in the brain. However, while significant progress has been made in delivering genes exclusively to neurons, such capabilities are lacking for glia, despite their demonstrated role in disease formation, posing a critical medical need. Although gene delivery to neurons can be achieved using viral vectors, their use to transmit genes to glia in-vivo has been unsuccessful. Here, we propose to design a novel nonviral gene delivery vector targeting microglia or astrocytes exclusively by bioengineering Modified RNAs (ModRNAs). ModRNAs are synthetic RNA molecules known not to trigger an immune response and are strongly expressed in target cells. Currently, ModRNAs enable only days- long expression, impeding long-duration medical applications and lacking cell specificity to glia types. We will engineer glia-type-specific, ModRNAs-based constructs, GliaRNAs, as a platform for glia-exclusive gene therapy, with a customizable expression duration. First, ModRNAs that enable robust and prolonged expression (7-14 days) will be developed (Aim 1). For this purpose, existing ModRNA will be altered, through modifications and by inflicting random mutations of structural components of the molecule, including CAP analog, 3’ untranslated region, coding region, 5’ untranslated region, and the poly-A tail. We will test the expression of these novel GliaRNAs in glial cultures from mice. Next, the vector specificity will be optimized (Aim 2). We will screen for molecular manipulations that enable robust and specific delivery of the GliaRNAs into either microglia or astrocytes (GliaRNA-vectors) and select the best gene delivery vectors, specifically either lipid nanoparticles, antibody-lipid conjugates, or aptamers. As a proof of concept, we will use the new GliaRNA-vector technology to express the green fluorescent protein (GFP) in either astrocytes or microglia in mice brains. The GliaRNA-vector platform will pave the way for genetically healing and modifying different types of glia, opening multiple therapeutic and research avenues in humans by targeting neurodegeneration, autism, pain disorders, mood disorders, and brain cancers.

抽象的 神经胶质是人脑中受支持的细胞，完成了小胶质细胞，少突胶质细胞，星形胶质细胞和室心脑室细胞。 神经胶质在神经系统的疾病中深处参与，例如阿尔茨海默氏症（AD），自闭症，疼痛，情感障碍， 和癌症。不同的神经胶质细胞类型在由特定基因驱动的疾病形成中起不同的机械作用。 因此，可以通过称为基因治疗的过程调节神经胶质基因表达 神经胶质在大脑中的有害作用。但是，尽管在传递基因方面取得了重大进展 对于神经元，缺乏神经胶质的能力，从而在疾病形成中表现出了作用，提出了关键 医疗需求。尽管可以使用病毒载体来实现至神经元的基因，但它们用于将基因传输到神经胶质 体内未成功。在这里，我们建议设计一种新型的非病毒基因递送载体，靶向小胶质细胞或 星形胶质细胞专门由生物工程修饰的RNA（MODRNA）。 ModRNA是已知的合成RNA分子 不触发免疫响应，并且在靶细胞中强烈表达。目前，莫德纳斯仅启用几天 - 长期表达，阻碍长期医学应用，缺乏对胶质类型的细胞特异性。我们将设计 胶质型特异性的基于modrnas的构建体Gliarnas，作为胶质排放基因疗法的平台，具有A 可自定义的表达持续时间。首先，可以实现稳健和延长表达（7-14天）的modrnas是 开发（目标1）。为此目的，将通过修改和随机造成现有的modrna 分子结构成分的突变，包括盖类似物，3“未翻译区域，编码区，5' 未翻译的区域和poly-a尾巴。我们将在小鼠的神经胶质培养物中测试这些新型Gliarnas的表达。 接下来，将优化向量特异性（AIM 2）。我们将筛选出可以鲁棒性和的分子操作 特异性递送Gliarnas到小胶质细胞或星形胶质细胞（Gliarna-vector）中，然后选择最佳基因递送 载体，特别是脂质纳米颗粒，抗体 - 脂质结合物或适体。作为概念证明，我们将使用 在星形胶质细胞或小胶质细胞中表达绿色荧光蛋白（GFP）的新型Gliarna-Vector技术 小鼠大脑。 Gliarna-Vector平台将为基因康复和修改不同类型的方式铺平道路 神经胶质，通过靶向神经变性，自闭症，开放人类多种治疗和研究途径 疼痛障碍，情绪障碍和大脑癌。