Memory and Regulatory T Cells Following T Cell Depletion in Transplantation

描述（由申请人提供）：几十年来，移植界一直在寻求实现对同种异体移植物的免疫耐受，这既是为了患者的健康利益，也是一种节约成本的措施。在与ITN的合作中，我们正在进行一项使用人类T细胞耗竭的耐受性试验。通过了解免疫抑制（IS）是否可以完全撤销的机制分析，这项研究将大大增强。我们的目标是使用临床前非人灵长类动物（NHP）肾移植模型作为验证1）耐受诱导方案和2）临床适用的免疫测定的手段，以帮助确定患者是否可以安全地从IS中退出。在本申请中，我们假设同种异体特异性记忆应答在耗尽后可以被充分阻断，并且调节性T细胞活性增强，使得长期移植的T细胞可以被激活。 NHP肾移植受者将实现耐受。此外，我们假设耐受性的获得和丧失是可测量和可预测的。我们根据以下观察结果提出这些假设。首先，在NHP或人类中单独的淋巴细胞耗竭通常不会导致耐受性诱导，但确实增加了具有效应子（TEFF）或记忆（TMEM）表型的T细胞的相对频率。第二，随着短期IS的加入，NHP中的T细胞耗竭已被证明可延长同种异体移植物存活并产生供体特异性无反应性。第三，调节性T细胞（Treg）活性可以通过在啮齿动物模型中使用常见的IS药物在体内被允许，甚至增强。最后，我们开发了一种人体体外试验，在啮齿动物中进行测试时，准确反映了对移植物的免疫状态。具体目标1是表征T细胞耗尽的NHP肾移植受者中的TMEM细胞扩增以及他克莫司（TAC）对控制TMEM群体的影响。我们将a）在接受维持生命的肾同种异体移植物的恒河猴中，在抗CD 3免疫毒素（IT）介导的T细胞耗竭后，检查TAC与或不与西罗莫司（SRL）一起对TMEM细胞恢复的影响，B）在T细胞恢复期间检查TMEM细胞的体外活性，c）监测影响T细胞耗竭和恢复的因素。对于特定目标2，我们将移植第二组猴子，并用IT/类固醇/TAC/SRL治疗，以测试一组新的免疫耐受性测定。我们将a）在接受者NHP接受SRL单一疗法时确定同种异体移植物耐受状态，B）将猴分配到耐受组或非耐受组，随后撤回SRL，c）用供体皮肤移植物测试耐受性，和d）监测TREG细胞频率和活性。在具体目标3中，我们将确定TREG细胞活性是否与慢性移植物肾病（CAN）的进展呈负相关。我们将a）监测TREG活性随CAN耐受性丧失/进展的变化，B）确定恢复SRL治疗是否阻断或延迟由于CAN导致的肾功能进一步丧失。

DESCRIPTION (provided by applicant): The transplant community has sought to achieve immune tolerance to allografts for decades, both for the health benefit of the patient and as a cost saving measure. In collaboration with ITN, we are conducting a tolerance trial using T-cell depletion in humans. This study would be augmented greatly by knowing whether immunosuppression (IS) can be withdrawn completely based on mechanistic assays. Our goal here is to use a preclinical non-human primate (NHP) renal transplant model as a means of validating 1) tolerance induction regimens and 2) clinically applicable immune assays to assist in determining whether patients can be safely withdrawn from IS. In this application, we hypothesize that the allo-specific memory response can be blocked sufficiently after depletion, and regulatory T cell activity enhanced, such that long term transplant tolerance in NHP renal transplant recipients will be achieved. Further, we hypothesize that the acquisition and loss of tolerance is measurable and predictable by the proposed assays. We base these hypotheses on the following observations. First, lymphocyte depletion alone in NHP or in humans does not generally result in tolerance induction but does increase the relative frequency of T cells with an effector (TEFF) or memory (TMEM) phenotype. Second, with the addition of short-term IS, T-cell depletion in NHPs has been shown to prolong allograft survival and to create donor-specific unresponsiveness. Third, regulatory T-cell (TREG) activity can be permitted, even enhanced, in vivo through the use of common IS drugs in rodent models. Lastly, we have developed an in vitro assay in humans, which when tested in rodents accurately reflects the immune status toward the graft. Specific Aim 1 is to characterize TMEM cell expansion in T-cell depleted NHP renal transplant recipients and the impact tacrolimus (TAC) on controlling the TMEM population. We will a) examine the effects of TAC with or without sirolimus (SRL) on TMEM cell recovery following anti-CD3 immunotoxin (IT)-mediated T-cell depletion in rhesus macaques receiving a life sustaining renal allograft, b) examine TMEM cell activity in vitro during T cell recovery, c) monitor factors that influence T cell depletion and recovery. For Specific Aim 2, we will transplant a second set of monkeys and treat with IT/steroid/TAC/SRL to test a novel set of immune tolerance assays. We will a) determine allograft tolerance status while recipient NHPs are on SRL monotherapy, b) assign monkeys to either tolerant or non-tolerant groups and subsequently withdraw SRL, c) test for tolerance with a donor skin graft, and d) monitor TREG cell frequency and activity. In Specific Aim 3 we will determine whether TREG cell activity inversely correlates with the progression of chronic allograft nephropathy (CAN). We will a) monitor changes in TREG activity with the loss of tolerance/progression of CAN, b) determine whether resumption of SRL treatment blocks or delays further loss of kidney function due to CAN.