Protection from GVHD with Gene-Modified Donor T Cells

描述（由申请人提供）：该项目的总体目标是开发一种有效的策略来管理严重的移植物抗宿主病（GVHD），同时保持供体T细胞的活性。同种异体造血干细胞移植（HSCT）后，供体T细胞有助于预防移植物排斥和复发（GVH效应）以及免疫恢复，但导致GVHD。目前的GVHD管理战略没有充分处理这些相互竞争的终点。我们提出，修饰表达单纯疱疹病毒/胸苷激酶基因（HSV/TK；自杀基因）的同种反应性供体T细胞可以维持体内功能（GVH效应），并被更昔洛韦诱导“自毁”，以防止严重GVHD的发展。对表达HSV/TK基因的转基因小鼠进行的HSCT临床前研究表明，受体接受更昔洛韦治疗后，GVHD得到控制，移植物排斥反应未发生。由于不分裂的非同种异体反应性T细胞不被诱导凋亡，免疫重建得到改善。然而，迄今为止，转基因小鼠的研究尚未反映出临床经验。在临床试验中，人类T细胞必须在体外用有丝分裂性CD3单克隆抗体扩增后用逆转录病毒载体转导。在这些试验中，同种异体反应性的显著丧失否定了任何潜在的益处，并阻止了关于该策略控制GVHD有效性的任何结论。在本提案中，将在临床前的dla -单倍体相同供体的HSCT狗模型中进行用病毒载体转导的T细胞研究。同种异体反应性T细胞的功能是移植所必需的，在这个模型中可以预测可能致命的GVHD。在目的1中，受体特异性细胞毒性T淋巴细胞（CTL）基因修饰（GM）与逆转录病毒载体表达HSV/TK基因将被添加回T-枯竭的骨髓，以评估同种异体反应功能。由于T细胞在被激活进行逆转录病毒转导时可能会失去同种异体反应性，因此在该模型中也将研究慢病毒载体，因为不需要激活，CD4和CD8 T细胞都被转导。在目标2中，将评估更昔洛韦诱导GM T细胞凋亡和随后控制GVHD的有效性。通过保留供体T细胞的作用，但防止严重GVHD的发展，同种异体造血干细胞移植的发病率和死亡率将会降低，并有可能将同种异体造血干细胞移植的“治疗”效益扩大到更广泛的非恶性疾病患者。

DESCRIPTION (provided by applicant): The overall goal of this project is to develop an effective strategy for management of severe graft-versus-host disease (GVHD) yet preserves donor T cell activity. After allogeneic hematopoietic stem cell transplantation (HSCT), donor T cells contribute to the prevention of graft rejection and relapse (GVH effect) as well as immune recovery but cause GVHD. Current strategies for the management of GVHD do not adequately deal with these competing endpoints. It is proposed that alloreactive donor T cells, modified to express the herpes simplex virus/thymidine kinase gene (HSV/TK; suicide gene) can maintain in vivo function (GVH effect) and be induced to "self-destruct" with ganciclovir to prevent the development of severe GVHD. Preclinical studies of HSCT from transgenic mice that express the HSV/TK gene show that GVHD was controlled and graft rejection did not occur after treatment of the recipient with ganciclovir. Since nondividing nonalloreactive T cells were not induced to apoptose, reconstitution of immunity was improved. However studies in transgenic mice have not reflected the clinical experience to date. In clinical trials, human T cells must be transduced with retroviral vectors after ex vivo expansion with a mitogenic CD3 monoclonal antibody. In these trials, there has been a significant loss of alloreactivity which negated any potential benefit and prevented any conclusions regarding the effectiveness of this strategy for controlling GVHD. In this proposal, studies will be conducted on T cells transduced with viral vectors in a preclinical dog model of HSCT from DLA-haploidentical donors. Functioning alloreactive T cells are required for engraftment and potentially fatal GVHD is predictable in this model. In aim 1, recipient-specific cytotoxic T lymphocytes (CTL) genetically-modified (GM) with retroviral vectors to express the HSV/TK gene will be added back to T-depleted marrow to assess alloreactive function. Since T cells may lose alloreactivity when activated for retroviral transduction, lentiviral vectors will also be studied in this model since activation is not required and both CD4 and CD8 T cells are transduced. In aim 2, the effectiveness of ganciclovir on the induction of apoptosis of GM T cells and the subsequent control of GVHD will be assessed. By preserving the effects of the donor T cells but preventing the development of severe GVHD, the morbidity and the mortality of allogeneic HSCT will be decreased and it will be possible to extend the "curative" benefits of allogeneic HSCT more broadly to include patients with nonmalignant diseases.