DIRECTING T CELL RESPONSES DURING IMMUNE RECONSTITUTION

在免疫重建过程中指导 T 细胞反应

• 批准号: 6290854
• 负责人: CRYSTAL L MACKALL
• 金额: --
• 依托单位: DIVISION OF CLINICAL SCIENCES - NCI
• 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/6290854
• 关键词: CD95 molecule; Ewing's tumor; T lymphocyte; antigen presenting cell; apoptosis; cellular immunity; clinical research; clinical trials; human subject; human therapy evaluation; immunoregulation; interleukin 2; neoplasm /cancer chemotherapy; neoplasm /cancer immunotherapy; polymerase chain reaction; rhabdomyosarcoma; tissue /cell culture; tumor antigens; western blottings

The objectives of this project are to study the biology of T cell regeneration and to develop therapeutic approaches which could be used in T cell depleted hosts to improve overall host immunocompetence and to direct T cell responses toward tumor-specific antigens in the setting of minimal residual neoplastic disease. Using a murine model of HY skin graft rejection, following T cell depletion, we have shown that thymectomized mice which have undergone immune reconstitution via peripheral expansion are unable to reject HY disparate skin grafts while hosts reconstituted via thymic-dependent pathways reject grafts normally. Rejection is accomplished in thymectomized mice if sufficient numbers of T cells are provided which we have shown requires approximately 10% of the normal T cell repertoire. In these experiments, we have also shown that IL-7 potently enhances thymic-independent T cell regeneration and that IL-7 therapy restores the capacity to reject HY disparate grafts in thymectomized hosts reconstituted with insufficient T cell numbers. Mechanistically, this appears to be related, at least in part, to IL-7?s ability to inhibit programmed cell death since inhibition of programmed cell death (PCD) by the use of bcl-2 transgenic T cell inocula also enhances immune competence in this model. Therefore, these results suggest that PCD may serve to limit host immune competence during peripheral expansion and raise the possibility that modulation of PCD in vivo may serve as a useful approach for enhancing immune competence in thymic-deficient TCD hosts. This model has also emphasized the critical role of IL-7 in modulating the process of peripheral expansion in vivo. Based upon these results, we have hypothesized that endogenous IL-7 production could contribute to T cell homeostasis by enhancing the process of peripheral expansion in the setting of T cell depletion. In order to test this hypothesis, we sought to evaluate serum IL-7 levels in children with T cell depletion due to HIV infection. We have observed a striking inverse correlation between serum IL-7 levels and T cell counts in children with HIV associated CD4+ depletion. Statistically, the serum IL-7 levels appear to be related to both CD4+ and CD8+ T cell counts, but not B cell counts. Current work is underway to address whether similar relationships exist in adults with T cell depletion and in T cell depletion related to cytotoxic therapy. These studies may serve to identify a new role for IL-7 in maintenance of T cell homeostasis.With regard to thymic-dependent pathways of T cell regeneration, we are currently investigating the relative importance of age- or disease-associated thymic dysfunction in limiting T cell receptor repertoire diversity following T cell depletion in humans. In order to evaluate the ability of low level thymic function to diversify the T cell receptor (TCR) repertoire over time in humans, current work is focused upon evaluation of TCR repertoire diversity in a series of pediatric patients treated with highly active antiretroviral therapy for HIV infection. We have hypothesized that even suboptimal thymopoiesis in this patient population may lead to diversification of the TCR repertoire over time. In an attempt to measure such changes in TCR diversity, we have developed a semi-quantitative approach using complementarity determining region 3 (CDR3) size analysis of serially diluted purified CD4+ T cells. Interestingly, preliminary results using this approach in normal individuals have shown significant age related changes in T cell receptor repertoire diversity. Whereas cord blood shows a diverse repertoire with as few as 10e5 input CD4 cells, young adults do not show a diverse repertoire until 5 x 10e5 input CD4 cells are utilized. Older adults (40-50 years) do not show a diverse repertoire until 1 x 10e6 input CD4+ T cells are utilized. These results suggest that measurable declines in TCR repertoire diversity occur throughout the normal human lifespan. With regard to children treated with highly active antiretroviral therapy, we have observed significantly reduced TCR repertoire diversity in children with CD4+ T cell counts <200 cells/mcl. In contrast, preliminary results in children with higher CD4+ T cell counts (e.g. >400 cells/mcl) reveal essentially normal TCR repertoire diversity. We have also used this approach to compare TCR repertoire diversity in CD45RA+ vs. CD45RO+ CD4+ T cells from patients with HIV infection. Thus far, no significant differences in repertoire diversity have been observed. Future plans involve the use of this technique to monitor changes in TCR repertoire diversity in cancer patients treated with immunorestorative therapies in the context of our ongoing clinical trials. AIDS RELATED 100%

本项目的目标是研究T细胞再生的生物学，并开发可用于T细胞耗竭宿主的治疗方法，以提高宿主的整体免疫能力，并在微小残留肿瘤疾病的背景下引导T细胞对肿瘤特异性抗原的反应。利用HY皮肤移植排斥反应的小鼠模型，在T细胞耗尽后，我们发现通过外周扩张进行免疫重建的胸腺切除小鼠不能排斥不同的HY皮肤移植，而通过胸腺依赖途径重建的宿主正常地排斥移植物。在胸腺切除的小鼠中，如果提供足够数量的T细胞，就可以实现排斥反应，我们已经证明，这需要大约正常T细胞库的10%。在这些实验中，我们还证明了IL-7有效地促进了胸腺非依赖性T细胞的再生，并且IL-7治疗在T细胞数量不足的胸腺切除宿主中恢复了排斥HY不同移植物的能力。从机制上讲，这可能至少部分与IL-7抑制细胞程序性死亡的能力有关，因为在该模型中，使用bcl2转基因T细胞接种抑制细胞程序性死亡也提高了免疫活性。因此，这些结果表明，PCD可能在外周扩张时限制宿主的免疫能力，并增加了体内PCD的调节可能成为增强胸腺缺陷TCD宿主免疫能力的有效途径的可能性。该模型还强调了IL-7在体内调节外周扩张过程中的关键作用。基于这些结果，我们假设内源性IL-7的产生可能通过在T细胞耗竭的情况下促进外周扩张过程来促进T细胞的动态平衡。为了验证这一假设，我们试图评估因HIV感染而导致T细胞耗竭的儿童的血清IL-7水平。我们观察到HIV相关的CD4+耗竭儿童的血清IL-7水平和T细胞计数之间存在显著的负相关。统计学上，血清IL-7水平似乎与CD4+和CD8+T细胞计数有关，但与B细胞计数无关。目前正在进行的工作是解决在成人T细胞耗竭和与细胞毒治疗相关的T细胞耗竭中是否存在类似的关系。这些研究可能有助于确定IL-7在维持T细胞稳态中的新作用。关于胸腺依赖的T细胞再生途径，我们目前正在研究年龄或疾病相关的胸腺功能障碍在限制人类T细胞耗竭后T细胞受体谱系多样性方面的相对重要性。为了评估低水平胸腺功能随着时间的推移而使T细胞受体(TCR)谱多样化的能力，目前的工作集中在评估一系列接受高效抗逆转录病毒治疗的儿童患者的TCR谱多样性。我们假设，即使这些患者群体的胸腺生成能力不佳，随着时间的推移，也可能导致TCR谱系的多样化。为了测量TCR多样性的这种变化，我们开发了一种半定量方法，使用序列稀释的纯化的CD4+T细胞的互补决定区3(CDR3)大小分析。有趣的是，在正常人中使用这种方法的初步结果显示，与年龄相关的T细胞受体谱系多样性的显著变化。脐带血只有10e5输入的CD4细胞，而年轻人只有在5x10e5输入的CD4细胞被利用后才能表现出多样化的谱系。老年人(40-50岁)在1x10e6输入的CD4+T细胞被利用之前不会表现出多样化的能力。这些结果表明，在正常的人类寿命中，TCR曲目多样性出现了可测量的下降。对于接受高效抗逆转录病毒治疗的儿童，我们观察到CD4+T细胞计数为200个/mCL的儿童TCR谱系多样性显著降低。相比之下，在CD4+T细胞计数较高的儿童中(例如，400细胞/MCL)的初步结果显示，TCR谱系的多样性基本上正常。我们还使用这种方法比较了HIV感染患者CD45RA+和CD45RO+CD4+T细胞中TCR谱系的多样性。到目前为止，还没有观察到曲目多样性方面的显著差异。未来的计划包括在我们正在进行的临床试验的背景下，使用这项技术来监测接受免疫修复治疗的癌症患者TCR谱系多样性的变化。与艾滋病100%相关