Solving the elusive mechanism of rapamycin action in lymphocytes

描述（由申请人提供）：mTOR是一种普遍表达的丝氨酸/苏氨酸激酶。mTOR酶存在于两种细胞复合物中，TORC 1和TORC 2。mTOR是基于其与药物雷帕霉素的相互作用而被发现和命名的。该化合物是一种变构抑制剂，部分阻断TORC 1活性，通常不抑制TORC 2功能。雷帕霉素对T和B淋巴细胞的治疗可显著阻断由抗原和其他有丝分裂原引发的增殖，雷帕霉素（西罗莫司; Rapamune（R））是FDA批准的免疫抑制剂。然而，雷帕霉素和类似物对包括癌细胞在内的其他细胞系统中的增殖具有较小的影响。尽管雷帕霉素对淋巴细胞的影响已经研究了20年，但仍然不清楚为什么该化合物选择性地诱导淋巴细胞增殖的深度阻断。有趣的是，一类新的ATP竞争性mTOR激酶抑制剂（TOR-KI）比雷帕霉素引起更少的免疫抑制，尽管对TORC 1和TORC 2都引起更完全的抑制。该项目的目标是确定雷帕霉素选择性阻断正常淋巴细胞增殖的机制。根据初步数据，我们提出了两个假设，形成了单独的具体目标的基础。目的1将检验雷帕霉素对S6激酶（TORC 1的关键底物）产生更完全和持续抑制的假设。有两种S6激酶，S6 K1和S6 K2，但它们在淋巴细胞活化和增殖中的作用尚未报道。我们将使用一种新的小鼠模型，这将允许一种化学遗传方法，以特异性抑制正常T和B细胞中的S6 K活性。我们将使用该模型比较S6 K抑制与雷帕霉素和TOR-KI的作用。目的2将检验雷帕霉素抑制mTOR的激酶非依赖性功能的假设。我们将产生一种新的小鼠品系，其中激酶死亡的mTOR可以以细胞特异性方式表达。这将使我们能够测试表达激酶死亡mTOR的淋巴细胞将保留对雷帕霉素敏感的残余增殖的预测。我们还将开始一种候选方法来测试可能的激酶非依赖性mTOR功能。 公共卫生相关性：一种名为雷帕霉素的药物用于抑制免疫系统，防止器官移植排斥反应。然而，目前尚不清楚为什么雷帕霉素与其他细胞类型相比选择性地阻断免疫细胞的激活。这项建议旨在解决这个长期存在的问题。

DESCRIPTION (provided by applicant): mTOR is a serine/threonine kinase that is expressed ubiquitously. The mTOR enzyme is present in two cellular complexes, TORC1 and TORC2. mTOR was discovered and named based on its interaction with the drug rapamycin. This compound is an allosteric inhibitor that partially blocks TORC1 activity and generally does not inhibit TORC2 function. Rapamycin treatment of T and B lymphocytes profoundly blocks proliferation triggered by antigen and other mitogens, and rapamycin (sirolimus; Rapamune(R)) is an FDA-approved immunosuppressant. However, rapamycin and analogs have lesser effects on proliferation in other cellular systems including cancer cells. Although the effects of rapamycin on lymphocytes have been studied for 20 years, it remains unclear why the compound induces a profound block of proliferation selectively in lymphocytes. Paradoxically, a novel class of ATP-competitive mTOR kinase inhibitors (TOR-KIs) causes less immunosuppression than rapamycin despite causing more complete inhibition of both TORC1 and TORC2. The goal of this project is to define mechanisms by which rapamycin selectively blocks proliferation of normal lymphocytes. Based on preliminary data, we propose two hypotheses that form the basis of separate specific aims. Aim 1 will test the hypothesis that rapamycin causes more complete and sustained inhibition of S6 kinases, key substrates of TORC1. There are two S6 kinases, S6K1 and S6K2, but their roles in lymphocyte activation and proliferation have not been reported. We will use a novel mouse model that will allow a chemical genetic approach to specifically inhibit S6K activity in normal T and B cells. We will use this model to compare the effects of S6K inhibition with the effects of rapamycin and TOR-KIs. Aim 2 will test the hypothesis that rapamycin inhibits kinase-independent functions of mTOR. We will generate a novel mouse strain in which kinase-dead mTOR can be expressed in a cell-specific manner. This will allow us to test the prediction that lymphocytes expressing kinase-dead mTOR will retain residual proliferation that remains sensitive to rapamycin. We will also begin a candidate approach to test possible kinase-independent mTOR functions. PUBLIC HEALTH RELEVANCE: A drug called rapamycin is used to suppress the immune system and prevent rejection of organ transplants. However, it is not known why rapamycin selectively blocks the activation of immune cells, compared to other cell types. This proposal seeks to solve this long-standing question.