Gene Therapy For Immune Deficiencies

免疫缺陷的基因治疗

• 批准号: 7194079
• 负责人: Harry L Malech
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7194079
• 关键词: CD34 molecule; T cell receptor; chemokine receptor; chronic granulomatous disease; clinical trials; gene therapy; graft versus host disease; hematopoietic stem cells; hematopoietic tissue transplantation; homologous transplantation; human subject; human therapy evaluation; immunomagnetic separation; immunopathology therapy; inborn immunodeficiency; laboratory mouse; lymphocyte proliferation; method development; patient oriented research; polymerase chain reaction; receptor expression; restriction fragment length polymorphism; severe combined immunodeficiency; stem cell transplantation; tissue /cell culture; transfection /expression vector

This project involves the conduct of therapeutic clinical trials for the treatment of inherited immune deficiencies using hematopoietic stem cell therapies including allogeneic transplantation and autologous blood stem cell targeted gene therapy. This project also includes associated studies of the diagnostic procedures (including genetic diagnosis), and treatment modalities that are alternatives to transplantation and gene therapy for those patients with the inherited immune deficiencies that are the target diseases for our transplant and gene therapy program. About four years ago we completed a clinical trial of gene therapy for the inherited deficiency of the phagocytic cell immune system known as the X-linked form of chronic granulomatous disease (X-CGD). Patients with CGD have defective circulating blood neutrophils that fail to produce microbicidal hydrogen peroxide. They suffer from recurrent life threatening infections and premature mortality. Six years ago we completed a phase I study of gene therapy for the p47phox deficient autosomal recessive form of CGD (AR-CGD). In that study we demonstrated that a single cycle of gene therapy targeting cytokine mobilized and purified stem cells could result in production of small numbers (.004% to .05%) functionally corrected neutrophils in the peripheral blood that persisted for 2 to 6 months. Following completion of this first trial, we incorporated a number of technical improvements in the gene transfer technology into a modification of the gene therapy clinical trial and focused on gene therapy for the X-linked, gp91phox deficient form of CGD. An amended gene therapy protocol for CGD began to enroll patients in April 1998 incorporating these enhancements and enrollment and treatments were completed in March 2000 and results were reported. In some of the gene therapy treated patients up to 1 in 400 circulating neutrophils in the peripheral blood demonstrated functional correction following the gene therapy. This peak level of correction occurred at 3 to 6 weeks after therapy and the effect could be sustained for over a year in three of five patients treated with multiple infusions of autologous ex vivo gene corrected CD34+ progenitor cells. These gene therapy studies demonstrate that it is possible to provide a low level partial and transient correction of the CGD defect in patients by gene therapy. While the level of correction is not a cure and may not be at a level that provides clinical benefit, it represents a very important demonstration of the principle that gene therapy can correct the biochemical defect of CGD in the patient. It may be that even low level and transient correction might provide clinical benefit in the setting of severe recurrent infections. The current study has achieved its scientific goal of demonstrating feasibility. We have continued to follow these patients for several years and have noted the disappearance of marked cells by about a year after the last treatment. No adverse effects of the gene therapy have been observed more than three to five years after the gene therapy. Recently it has become possible to enumerate and determine the genome location through sequencing of retrovirus insertion sites using a novel method called linear amplification modified polymerase chain reaction (LAM-PCR). We also are in the process of delineated insertion site in archived blood samples from the CGD clinical trial. We are in the planning stage of a study to determine if non-ablative marrow conditioning might enhance the level and durability of the effect of gene therapy for CGD. One of the important questions is whether in a clinical setting it is possible to use non-ablative marrow cytoreduction to more safely achieve the engraftment of stem cells. While this question has an impact on achieving success in gene therapy, advances in the use of non-ablative conditioning for allogeneic transplantation have alllowed us to explore the potential of this approach to achieve curative allogeneic transplantation for CGD. An ongoing clinical trial was initiated and recently completed in which patient with CGD undergo non-ablative marrow conditioning with immune suppression achieved with a combination of cyclophosphamide, fludarabine and anti-thymocyte globulin. The patients then received a transplant with purified CD34+ peripheral blood stem cells mobilized from a fully 6/6 HLA matched sibling of the CGD patient. The graft is depleted of most lymphocytes and donor lymphocytes are infused at later time points after transplant to help to establish the donor graft. 5 adults and 5 children were transplanted. 4 of 5 children achieved stable long term engraftment that appears to provide a significant level of protection from infection, and all 5 children are alive and well. While 4 of 5 adults achieved long term engraftment, there were three deaths, one from complications of graft versus host disease, one from pneumococcal pneumonia at 1 year post transplant, and one from complications from a second fully ablative salvage transplant procedure. Of the two adult CGD transplant patients who are fully engrafted one remains infection-free for 3 years without any complications and appears cured of his CGD. The other fully engrafted adult transplant patient is also infection-free and appears to be cured of his CGD, but continues to have mild and well controlled graft versus host disease. We conclude that non-ablative matched related allogeneic transplant is a reasonable option in pediatric patients with CGD and a high risk of mortality from recurrent infection. Currently adults appear to have a higher risk of complications from graft versus host disease and delayed recovery of lymphocyte immunity. Because of that we have begun to study the potential of extracorporeal photophoresis to treat chronic graft versus host disease. If graft versus host disease risks can be reduced it would then reduce one of the risks of allogeneic transplantation for inherited immune deficiencies. A follow up trial of non-ablative allogeneic transplantation in children with high risk CGD has begun using modified procedures to enhance engraftment and reduce the risks of graft versus host disease. A similar transplant trial has been developed as a salvage therapy for CGD patients with incurable infection. In order to reduce graft versus host disease we have begun a clinical trial to study the effect of extracorporeal photopheresis (ECP) on treatment of graft versus host disease. We have also define novel immunologic changes in ratios of central and effector memory T-cells that correct following over three months of ECP. An important goal is to also develop gene therapy for X-linked severe combined immune deficiency. However, within the past year there has been a report of two out of 9 infants with XSCID cured with gene therapy in France who at 2 to 3 years after the gene therapy developed lympocytic leukemia as a result in insertion al mutagenesis in the LMO-2 oncogene. Regulatory bodies such as the Food and Drug Administration and the Recombinant DNA Advisory Committee of the Office of Biotechnology Activities have determined that gene therapy for XSCID should be restricted to patients who do not have other therapeutic alternative. We have studied a series of patients with XSCID without a matched sibling who have failed to achieve significant benefit from the standard therapy of a haploidentical marrow transplant from a parent. These patients have recurrent infections and extreme failure to grow normally expressing significant pulmonary impairment. We have begun a clinical trial of ex vivo gene therapy for this group of XSCID patients who are doing poorly and lack other therapeutic alternatives, with early evidence of enhanced marking and correction of T lymphocytes in this clinical trial in 3 patients.

该项目涉及进行治疗性临床试验，以使用造血干细胞疗法（包括同种异体移植和自体血干细胞靶向基因疗法）治疗遗传性免疫缺陷。该项目还包括对诊断程序（包括遗传诊断）的相关研究，以及对于那些具有遗传性免疫缺陷患者的移植和基因治疗的治疗方式，这些遗传性缺乏症是我们移植和基因疗法计划的靶向疾病。大约四年前，我们完成了一项基因治疗的临床试验，用于遗传性吞噬细胞免疫系统，称为X连锁形式的慢性肉芽肿性疾病（X-CGD）。 CGD患者的血液中性粒细胞有缺陷，无法产生过氧化杀菌氢。他们遭受危及生命感染和过早死亡的反复发作。六年前，我们完成了对CGD（AR-CGD）的P47Phox缺乏常染色体隐性形式的基因治疗的I期研究。在这项研究中，我们证明了靶向细胞因子动员和纯化的干细胞的单个基因治疗周期可能导致少量（.004％至0.05％）在功能矫正的外周血中持续2到6个月的中性粒细胞的产生。在第一次试验完成后，我们将基因转移技术中的许多技术改进纳入了基因治疗临床试验的修饰中，并着重于X连锁的GP91PHOX缺乏CGD的基因治疗。 1998年4月，CGD的一项经过修改的基因治疗方案开始招募患者，并于2000年3月完成了这些增强和入学率和治疗方法，并报告了结果。在某些基因治疗中，在外周血中，在400个循环中性粒细胞中最多有1例在基因治疗后表现出功能矫正。校正峰值水平发生在治疗后3至6周时，在五名患者中，有三名用多种自体外基因校正了CD34+祖细胞治疗的五名患者中的三名可以持续一年多。这些基因治疗研究表明，通过基因治疗，可以对患者的CGD缺陷进行低水平的部分和短暂校正。虽然校正水平不能治愈，也可能无法提供临床益处的水平，但它代表了基因治疗可以纠正患者CGD生化缺陷的原理的一个非常重要的证明。在严重复发感染的情况下，即使是低水平和瞬时校正也可能会提供临床益处。当前的研究实现了证明可行性的科学目标。我们已经继续关注这些患者已有几年了，并注意到最后一次治疗后大约一年的标记细胞消失。基因治疗后三到五年未观察到基因治疗的不良影响。最近，使用一种称为线性扩增修饰的聚合酶链反应（LAM-PCR）的新方法，通过对逆转录病毒插入位点进行测序来枚举和确定基因组位置。我们也在CGD临床试验中的存档血液样本中划定的插入部位。我们正处于研究的计划阶段，以确定非燃烧性骨髓调节是否可以提高基因治疗对CGD的影响的水平和耐用性。重要的问题之一是，在临床环境中，是否可以使用非燃烧的骨髓细胞减少来更安全地植入干细胞。尽管这个问题对在基因疗法方面取得成功有影响，但使用非燃烧性调理进行同种异体移植的进步使我们探索了这种方法的潜力，以实现CGD的治愈性同种异体移植。开始了一项正在进行的临床试验并最近完成，其中CGD患者接受了非燃烧性骨髓调节，并结合了环磷酰胺，氟达拉滨和抗心理细胞球蛋白，并结合使用了免疫抑制作用。然后，患者接受了纯化的CD34+外周血干细胞的移植，从完全6/6 HLA匹配的CGD患者的兄弟姐妹动员。移植物耗尽大多数淋巴细胞，供体淋巴细胞在移植后的稍后时间注入以帮助建立供体移植物。 5名成人和5名儿童被移植。 5个儿童中有4个实现了稳定的长期植入，似乎可以提供很大的保护免受感染的保护，所有5名儿童都活着且良好。虽然5名成年人中有4例长期植入，但有3例死亡，一次是由移植物与宿主疾病的并发症发生的，其中一名来自移植后1年的肺炎球菌性肺炎，另一项来自第二个完全消融的抢救移植程序的并发症。在两名完全植入的成年CGD移植患者中，没有任何并发​​症没有任何并发​​症，并且似乎治愈了他的CGD。另一个完全植入的成人移植患者也没有感染，并且似乎已治愈了他的CGD，但仍具有轻度且控制良好的移植物与宿主疾病。我们得出的结论是，与CGD的儿科患者相关的非抗性相关同种异体移植是一个合理的选择，并且复发感染的死亡率很高。目前，成年人似乎患有移植物与宿主疾病的并发症风险更高，淋巴细胞免疫的恢复延迟。因此，我们已经开始研究体外光电治疗慢性移植物与宿主疾病的潜力。如果可以降低移植物与宿主疾病的风险，那么它将降低遗传免疫缺陷的同种异体移植风险之一。 CGD高风险儿童非燃烧性同种异体移植的随访试验已开始使用改良程序来增强植入术并降低移植物与宿主疾病的风险。针对患有无法治愈的CGD患者的打捞疗法已经开发了类似的移植试验。为了减少移植物与宿主疾病，我们已经开始了一项临床试验，研究体外光寄生发生（ECP）对治疗移植物与宿主疾病的影响。我们还定义了中央和效应子记忆T细胞比率的新型免疫学变化，这些变化在三个月以后的ECP之后纠正了。 一个重要的目标是为X连锁的严重合并免疫缺陷开发基因治疗。然而，在过去的一年中，有9名在法国XSCID治疗基因治疗的婴儿中有2例在基因治疗后2至3年中出现了淋巴细胞性白血病，从而导致LMO-2 Oncogene插入诱变。食品药物管理局和生物技术活动办公室的重组DNA咨询委员会等监管机构已确定XSCID的基因治疗应仅限于没有其他治疗替代方案的患者。我们已经研究了一系列XSCID的患者，没有匹配的兄弟姐妹，这些患者未能通过父母的单倍性骨髓移植的标准疗法获得可观的好处。这些患者复发感染和极端失败，正常表达明显的肺部受损。我们已经开始对这组XSCID患者进行过体内基因治疗的临床试验，这些XSCID患者的表现不佳且缺乏其他治疗替代方案，在这项临床试验中，早期证据表明对T淋巴细胞的标记和矫正增强。