Protection from GVHD with Gene-Modified Donor T Cells

描述（由申请人提供）：本项目的总体目标是开发一种有效的策略，用于管理严重的移植物抗宿主病（GVHD），同时保留供体T细胞活性。异基因造血干细胞移植（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，但会导致GVHD。目前的GVHD管理策略没有充分处理这些竞争性终点。有人提出，同种异体反应性供体T细胞，修饰表达单纯疱疹病毒/胸苷激酶基因（HSV/TK;自杀基因），可以保持在体内功能（GVH效应），并诱导“自毁”与更昔洛韦，以防止严重GVHD的发展。从表达HSV/TK基因的转基因小鼠进行的HSCT的临床前研究表明，GVHD得到控制，接受更昔洛韦治疗后未发生移植物排斥反应。由于未分裂的非同种异体反应性T细胞未被诱导为果糖，因此免疫重建得到改善。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T细胞必须在用促有丝分裂CD 3单克隆抗体离体扩增后用逆转录病毒载体转导。在这些试验中，同种异体反应性显著丧失，这否定了任何潜在的益处，并阻止了关于该策略用于控制GVHD的有效性的任何结论。在该提案中，将在来自DLA-半相合供体的HSCT临床前犬模型中对用病毒载体转导的T细胞进行研究。功能性同种异体反应性T细胞是植入所必需的，并且在该模型中可预测潜在致命的GVHD。在目标1中，将用逆转录病毒载体遗传修饰（GM）以表达HSV/TK基因的致敏特异性细胞毒性T淋巴细胞（CTL）添加回T细胞耗竭的骨髓中，以评估同种异体反应性功能。由于T细胞在被激活用于逆转录病毒转导时可能失去同种异体反应性，因此也将在该模型中研究慢病毒载体，因为不需要激活并且CD 4和CD 8 T细胞都被转导。在目标2中，将评估更昔洛韦对GM T细胞的凋亡诱导和随后的GVHD控制的有效性。通过保留供体T细胞的作用但防止严重GVHD的发展，异基因HSCT的发病率和死亡率将降低，并且将有可能将异基因HSCT的“治愈”益处更广泛地扩展到包括患有非恶性疾病的患者。

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.