Site-Specific Correction of Sickle Cell Disease Using Acoustofludic Gene Delivery

使用声流控基因传递对镰状细胞病进行位点特异性校正

基本信息

批准号：
10023174
负责人：
Jason Nathaniel Belling
金额：
$ 3.23万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
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：
2019-09-16 至 2021-06-10
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10023174
关键词：
Acoustics Address Adult Affect Autoimmune Diseases Autologous Autologous Transplantation Biological Assay CD34 gene CRISPR/Cas technology Cell Line Cell Membrane Permeability Cell Therapy Cell membrane Cell model Cell physiology Cells Clinical Clinical Management Clustered Regularly Interspaced Short Palindromic Repeats Collaborations Complex DNA Sequence Alteration Development Devices Disease Electroporation Engraftment Erythrocytes Erythroid Flow Cytometry Foundations Frequencies Gene Cluster Gene Delivery Gene-Modified Generations Genes Goals Heart Diseases Hematological Disease Hematopoietic Stem Cell Transplantation Hematopoietic stem cells Hemoglobin Hemoglobinopathies Hereditary Disease High Pressure Liquid Chromatography Human Cell Line Improve Access Intervention K-562 Lung diseases Mechanics Mediating Medical Methods Microfluidics Modeling Mutation Output Patients Peripheral Blood Mononuclear Cell Permeability Polymerase Chain Reaction Process Production Property Quality of life Reaction Recovery Research Ribonucleoproteins Risk Running Sickle Cell Sickle Cell Anemia Site Stem cell transplant Structure System T-Lymphocyte Techniques Technology Testing Therapeutic Toxic effect Transfection Translations Xenograft procedure base beta Globin bioinformatics tool cell injury clinical practice clinical translation clinically relevant cost design gene correction gene therapy graft vs host disease immunogenic innovation insertion/deletion mutation interdisciplinary collaboration lipofection mouse model new technology next generation sequencing processing speed repaired stem stem cell population stem cell therapy stem cells targeted nucleases uptake vector voltage

项目摘要

Project Summary Sickle cell disease (SCD) is among the most common monogenetic inherited disorders. Clinical management of SCD is primarily supportive. However, in the most severe cases, the only definitive curative option for patients suffering from SCD is an allogeneically matched hematopoietic stem cell transplant. This hemoglobinopathy directly affects the structure and function of hemoglobin, leading to deficiencies of β-globin chains in the development of functional adult hemoglobin. Furthermore, the lack of fully matched donors for patients to receive a stem cell transplant runs the risk of adverse immunogenic reactions, such as auto-immune disorders or graft-versus-host disease. Recent efforts to address this disease and its clinical sequela have focused on gene therapies based on the transplantation of autologous gene-modified hematopoietic stem & progenitor cells (HSPC), where a patient's own cells are corrected and reinfused to enable production of fully functioning erythrocytes. However, non-viral strategies for the batch processing of stem cell gene therapies are known to be inefficient and are unable to meet clinical demands. We hypothesize that the optimization of an acoustofluidic therapeutic platform that physically permeabilizes cells for the delivery of CRISPR-Cas9 biomolecules will address this technologic gap. This high-throughput gene-delivery strategy will enable our long-term goal to generate gene-modified stem cell therapies quickly and efficiently for curing sickle cell disease. This physical permeabilization process renders target cells transiently permeable, enabling vector uptake while minimizing damage to the cell membrane and maintaining high levels of viability. In order to achieve our clinical target, our proposed specific aims include: 1) optimize acoustofluidic gene delivery in model cell lines harboring the sickle cell mutation and 2) evaluate site-specific correction of the sickle cell disease mutation in hematopoietic stem and progenitor cells. Given the utility of this acoustofluidic technology, there is a wide range of heart, lung, and blood disorders that can be addressed, overcoming the state of the art for gene delivery. We expect the generation of rapid and safe gene-modified stem cell therapies using our acoustofludic technology will greatly improve access to these medical interventions and the quality of life for patients with the most severe cases of SCD.

项目概要镰状细胞病（SCD）是最常见的临床单基因遗传性疾病之一。 SCD 的治疗主要是支持性的，但在最严重的情况下，唯一确定的方法是支持性治疗。 SCD 患者的治疗选择是同种异体匹配的造血干细胞这种血红蛋白病直接影响血红蛋白的结构和功能，导致移植。功能性成人血红蛋白发育过程中β-珠蛋白链的缺陷。缺乏完全匹配的供体供患者接受干细胞移植会带来不良风险免疫原性反应，例如自身免疫性疾病或移植物抗宿主病。解决这种疾病及其临床后遗症的重点是基于自体基因修饰造血干细胞和祖细胞（HSPC）移植，其中患者自身的细胞被纠正并重新注入，以产生功能齐全的红细胞。然而，众所周知，用于批量处理干细胞基因疗法的非病毒策略是效率低下，无法满足临床需求。声流控治疗平台，可对细胞进行物理渗透以输送 CRISPR-Cas9 生物分子将弥补这一技术差距。该战略将使我们的长期目标能够快速产生基因修饰的干细胞疗法这种物理透化过程可有效治疗镰状细胞病。瞬时渗透，实现载体摄取，同时最大限度地减少对细胞膜的损害保持高水平的活力为了实现我们的临床目标，我们提出了具体的建议。目标包括：1）优化携带镰刀的模型细胞系中的声流控基因传递细胞突变和 2) 评估镰状细胞病突变的位点特异性校正鉴于这种声流控技术的实用性，造血干细胞和祖细胞。可以解决多种心脏、肺和血液疾病，克服现有技术我们期望使用快速且安全的基因修饰干细胞疗法。我们的声流控技术将极大地改善这些医疗干预措施的可及性和质量最严重的 SCD 病例患者的生命。