Peripheral T Cell tolerance by targeting AKT

描述(申请人提供)：我们实验室的目标是开发诱导外周T细胞耐受和预防人类造血细胞移植(HCT)后移植物抗宿主病(GVHD)的方法。移植可以治愈多种良性和恶性的造血系统疾病，但移植物抗宿主病仍然是移植相关的发病率、致残率和死亡率的主要病例，从而限制了HCT的使用。干细胞接种中包括的供者T细胞识别受体同种异体抗原，介导移植物抗宿主病。预防GVHD的理想方法是在移植后早期用较短的疗程选择性地清除同种异体激活的T细胞，而保留负责免疫重建和病原体控制的其他T细胞。如果调节性T细胞(Treg)也能幸免，耗尽同种异体反应性T细胞可能会使平衡倾向于调节和耐受。我们评估了TCN对T细胞活化的影响，发现TCN在体外抑制DNA合成、增殖和促进细胞凋亡。根据这些初步数据和文献中的证据，AKT促进T细胞的激活、存活、Th1细胞因子的产生，但抑制Treg的产生，我们推测TCN的治疗将阻止异基因骨髓移植后GVHD的发展。这笔赠款将为在接受异基因骨髓移植的患者中测试TCN预防GVHD所需的初步数据。如果建议的方法在预防人类移植物抗宿主病方面是有效的，更多的患者将受益于骨髓移植并实现治愈。公共卫生相关性：AKT是一种依赖于磷脂酰肌醇3-激酶(PI3K)的丝氨酸-苏氨酸激酶，促进葡萄糖代谢和激活T细胞的存活，并限制调节性T细胞(Tregs)的分化。虽然AKT对胸腺细胞的发育至关重要，但一些作者认为AKT对静息T细胞的生存并不是必不可少的。AKT的这些功能使其成为控制GVHD的一个有吸引力的靶点。这项建议的目的是通过靶向AKT途径来控制异基因骨髓移植(BMT)临床前模型中的急性GVHD。小分子AKT抑制剂TCN可抑制AKT的磷酸化，从而抑制其活性。用TCN治疗可以有效地抑制过表达AKT的人类癌细胞在裸鼠体内的生长。我们计划开发足够的临床前数据，以便设计一项针对人类的临床GVHD预防试验。我们的中心假设是，TCN的治疗抑制了T细胞的激活、扩增和存活，同时促进了体内对同种异体抗原的Treg生成，从而防止了异基因骨髓移植后GVHD的发生。为了验证这一假说，我们将追求以下两个具体目标：1)确定TCN对异基因骨髓移植后GVHD、免疫重建和GVL活性的影响。我们的初步数据表明，TCN在体外抑制T细胞对同种异体抗原的反应，促进抗原激活的T细胞的凋亡，提示TCN的治疗可能预防体内异基因骨髓移植后的GVHD。为此，我们将在临床前的小鼠骨髓移植模型上测试TCN对GVHD的预防效果。此外，我们还将评估TCN对骨髓重建和GVL活性的影响，以及供者T细胞的有益作用。2)明确TCN预防移植物抗宿主病的作用机制。TCN预防移植物抗宿主病可能涉及多种机制。在这个特定的目标中，我们将评估TCN对T细胞增殖、凋亡、细胞因子的产生、运输和Treg产生的潜在影响，这些都是GVHD发生的关键因素。为了实现这些目标，我们的团队拥有免疫学、骨髓移植和药物治疗方面的专业知识。独特的转基因、敲除和敲入小鼠可用于所有计划中的实验。这些研究将肯定评估TCN在临床相关骨髓移植模型中控制GVHD的有效性和机制，并可能为临床预防GVHD提供一种新的、更具选择性的策略。

DESCRIPTION (provided by applicant): The goal of our laboratory is to develop methods for the induction of peripheral T cell tolerance and prevention of graft-versus-host disease (GVHD) after human hematopoietic cell transplantation (HCT). Transplantation can cure a variety of benign and malignant hematopoietic disorders, but GVHD remains the primary case of transplant-related morbidity, disability and mortality, thereby limiting the use of HCT. Donor T cells that are included in the stem cell inoculum and recognize recipient alloantigens mediate GVHD. An ideal approach to prevent GVHD is to selectively deplete the alloactivated T cells with a short course of treatment early after transplantation, while sparing other T cells that would be responsible for immune reconstitution and control of pathogens. Depleting alloreactive T cells may tip the balance towards regulation and tolerance, if regulatory T cells (Tregs) are also spared. We have evaluated the effects of TCN on T cell activation and found that TCN suppresses DNA synthesis, proliferation and promotes apoptosis in vitro. Based on these preliminary data and the evidence in literature that AKT promotes T cell activation, survival, Th1 cytokine production but suppresses Treg generation, we reason that treatment of TCN would prevent the development of GVHD after allogeneic BMT. This grant would generate the required preliminary data for testing TCN for GVHD prevention in patients undergoing allogeneic BMT. If the proposed approach is effective in preventing GVHD in humans, more patients would benefit from BMT and achieve cure. PUBLIC HEALTH RELEVANCE: AKT, a phosphatidylinositol 3-kinase (PI3K)-dependent serine-threonine kinase, promotes glucose metabolism and survival of activated T cells and limits differentiation of regulatory T cells (Tregs). While AKT is critical for thymocyte development, some authors have proposed that AKT is not essential for survival of resting T cells. These functions of AKT make it an attractive target in controlling GVHD. The objective of this proposal is to control acute GVHD in pre- clinical models of allogeneic bone marrow transplantation (BMT) by targeting the AKT pathway. A small molecule AKT inhibitor, triciribine (TCN), suppress the phosphorylation of AKT thereby inhibiting its activity. Treatment with TCN potently inhibits growth in nude mice of human cancer cells that over-express AKT. We plan to develop sufficient preclinical data to allow the design of a clinical GVHD prevention trial in humans. Our central hypothesis is that treatment of TCN suppresses T cell activation, expansion and survival while promoting Treg generation in response to alloantigen in vivo, and thus prevents GVHD development after allogeneic BMT. To test this hypothesis, we will pursue the following two Specific Aims: 1) To determine the effects of TCN on GVHD, immune reconstitution and GVL activity after allogeneic BMT. Our preliminary data indicate that TCN suppresses T cell response to alloantigen and promotes apoptosis of antigen-activated T cells in vitro, suggesting that treatment of TCN may prevent GVHD after allogeneic BMT in vivo. In this Specific Aim, we will test the efficacy of TCN in the prevention of GVHD in pre-clinical murine models of BMT. Furthermore, we will evaluate the effects of TCN on marrow reconstitution and GVL activity, the beneficial effects of donor T cells. 2) To define the mechanisms of TCN in the prevention of GVHD. Multiple mechanisms may be involved in the prevention of GVHD by TCN. In this Specific Aim, we will evaluate the potential effects of TCN on T cell expansion, apoptosis, cytokine production, trafficking and Treg generation, which are critical factors in the development of GVHD. To accomplish these objectives, our team has expertise in immunology, bone marrow transplantation and drug therapy. Unique transgenic, knock-out and knock-in mice are available for all the planned experiments. The proposed studies will definitely evaluate the efficacy and mechanisms of TCN in controlling GVHD in clinically relevant models of BMT, and may point to a novel and more selective strategy to prevent GVHD in the clinic.