Sleeping Beauty-mediated Gene Therapy of X-linked SCID

睡美人介导的 X 连锁 SCID 基因治疗

• 批准号: 7908974
• 负责人: Kendra A. Hyland
• 金额: $ 27.07万
• 依托单位: DISCOVERY GENOMICS, INC.
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7908974
• 关键词: Address; Allogenic; Animal Model; Autologous; B-Lymphocytes; Bone Marrow; CD34 gene; Candidate Disease Gene; Cells; Clinical Trials; Cytokine Signaling; DNA; Data; Development; Disease; Effectiveness; Electroporation; Engraftment; Evaluation; Excision; Fetal Liver; Flow Cytometry; Gene Expression; Gene Transfer; Genes; Genomics; Goals; Graft Rejection; Hematological Disease; Hematopoietic; Hematopoietic stem cells; Hereditary Disease; Human; Immune; Immunity; Immunologic Deficiency Syndromes; Inherited; Interleukin 2 Receptor; Interleukin 2 Receptor Gamma; Interleukin-15; Interleukin-4; Interleukin-7; Interleukin-9; Interruption; Laboratories; Lead; Link; Lymphoid; Lymphoid Cell; Marrow; Mediating; Modeling; Molecular; Monitor; Mus; Mutation; Pathway interactions; Patients; Phase; Physiologic pulse; Plasmids; Preclinical Testing; Proteins; Recovery; Reporting; Research Personnel; Retroviridae; Serious Adverse Event; Signal Transduction; Site; Sleeping Beauty; Small Business Innovation Research Grant; Source; Stem cell transplant; Syndrome; System; T-Cell Leukemia; Testing; Transplantation; Transposase; Viral; Viral Genes; Viral Vector; Virus; Work; X-Linked Severe Combined Immunodeficiency; allotransplant; base; cellular targeting; gene therapy; gene therapy clinical trial; genotoxicity; graft vs host disease; immune function; interest; lymphoblastoid cell line; pre-clinical; public health relevance; research study; restoration; therapeutic gene

DESCRIPTION (provided by applicant): Primary immune deficiencies comprise a group of inherited genetic disorders caused by interruption of normal lymphoid development. These diseases are considered prime candidates for gene therapy by introduction of the missing gene into hematopoietic stem cells that can then differentiate into lymphoid cells, and thus restore immunity. X-linked severe combined immunodeficiency (X-SCID), caused by a mutation in the gene encoding the common ( chain gene ((c) of the receptors for IL-2, IL-4, IL-7, IL-9, IL-15, and IL-213, is one of the more common of the primary immunodeficiencies. Clinical gene therapy trials for X-SCID have focused on the use of retroviruses as an integrating viral vector for introduction of the (c gene, resulting in restoration of immunity in patients that have engrafted with transduced, autologous hematopoietic stem cells. However, serious adverse events have emerged in these clinical trials, namely a T-cell leukemia-like syndrome developing in 5 of 21 treated patients. Therefore, safer, alterative therapies are needed. At Discovery Genomics, Inc. (DGI), we are working on developing the Sleeping Beauty (SB) transposon system for gene therapy, including the targeting of hematopoietic stem cells for treatment of primary immunodeficiencies. The two-component SB system consists of a transposon (inverted repeats (IR's) flanking a therapeutic gene of interest) and a transposase that catalyzes excision of the transposon at the ends of the IRs and then integration into host cell chromosomal sequence. As a lead immunodeficiency, here we propose that X-SCID may be treatable without the use of a virus by combining the power of electroporation for introduction of DNA into cells, along with use of DGI's Sleeping Beauty transposon system to achieve integration and long-term (c gene expression. In this Phase I application, the overall goal is to establish conditions for SB-mediated gene therapy for X-SCID. In this regard, there are two key questions that will need to be addressed: (i) Will SB-mediated transposition of the (c gene allow functional correction of lymphoid cells? (ii) What is the effectiveness of SB-mediated (c gene insertion in the treatment of an animal model of X-SCID? There are two Specific Aims: Aim 1. Evaluate the effectiveness of Sleeping Beauty transposons for correction of (c chain deficiency in a lymphoblastoid cell line derived from an X-linked SCID patient. Aim 2. Evaluate Sleeping Beauty-mediated transposition and long term expression of (c gene in hematopoietic stem cells derived of X-SCID mice, as a model for SB-mediated gene therapy of X-SCID. Successful accomplishment of these goals will provide evidence for the effectiveness of the SB system to mediate non-viral gene transfer in murine hematopoietic stem cells (previously undemonstrated), as well as provide key preclinical data for the development of SB transposons for X-SCID as a lead primary immunodeficiency. PUBLIC HEALTH RELEVANCE: Results from these experiments will provide the technical basis for achieving transposon-mediated integration and long-term expression in hematopoietic stem cells, the cellular target inr gene therapy of primary immunodeficiencies. In addition, it will provide the basis for the development of the SB transposon system for treatment of other hematologic diseases.

描述（由申请人提供）：原发性免疫缺陷包括一组由正常淋巴发育中断引起的遗传性遗传疾病。这些疾病被认为是基因治疗的主要候选者，通过将缺失的基因引入造血干细胞，然后造血干细胞可以分化为淋巴样细胞，从而恢复免疫力。X连锁严重联合免疫缺陷（X-SCID）是由编码IL-2、IL-4、IL-7、IL-9、IL-15和IL-213受体的共同链基因（（c））的基因突变引起的，是较常见的原发性免疫缺陷之一。X-SCID的临床基因治疗试验集中在使用逆转录病毒作为整合病毒载体，用于引入c基因，从而恢复移植了转导的自体造血干细胞的患者的免疫力。然而，在这些临床试验中出现了严重的不良事件，即21名接受治疗的患者中有5名出现T细胞白血病样综合征。因此，需要更安全的替代疗法。 在Discovery Genomics，Inc. (DGI)目前，我们正致力于开发用于基因治疗的睡美人（SB）转座子系统，包括靶向造血干细胞治疗原发性免疫缺陷。双组分SB系统由转座子（位于感兴趣的治疗基因侧翼的反向重复序列（IR））和转座酶组成，所述转座酶催化在IR末端切除转座子，然后整合到宿主细胞染色体序列中。作为一种主要的免疫缺陷，在这里我们提出，X-SCID可能是可治疗的，而不使用病毒，通过结合电穿孔的力量，将DNA引入细胞，沿着使用DGI的睡美人转座子系统，以实现整合和长期的基因表达。在该I期申请中，总体目标是建立SB介导的X-SCID基因治疗的条件。在这方面，有两个关键的问题，将需要解决：（i）是否SB介导的转座的（c）基因允许淋巴样细胞的功能校正？(ii)SB介导的c基因插入治疗X-SCID动物模型的有效性如何？有两个具体目标：目标1。评价睡美人转座子用于校正来自X连锁SCID患者的淋巴母细胞系中的c链缺陷的有效性。目标2.评价Sleeping Beauty介导的（c）基因在X-SCID小鼠造血干细胞中的转座和长期表达，作为SB介导的X-SCID基因治疗的模型。这些目标的成功实现将为SB系统介导小鼠造血干细胞中非病毒基因转移的有效性提供证据（以前未证实），并为开发用于X-SCID的SB转座子作为主要的原发性免疫缺陷提供关键的临床前数据。 公共卫生相关性：这些实验结果将为实现转座子介导的整合和在造血干细胞中的长期表达，原发性免疫缺陷的细胞靶向inr基因治疗提供技术基础。此外，它将为开发SB转座子系统用于治疗其他血液病提供基础。