Survival Mechanisms in Leukemic NK Cells

描述（由申请人提供）：本提案的长期目标是了解NK白血病的发病机制。白血病NK细胞扩增的机制尚不清楚。该建议的中心假设是NK白血病是由细胞凋亡失调引起的。白血病NK细胞相对抵抗Fas介导的死亡，尽管表达高水平的Fas受体和Fas配体。IL-2激活后Fas耐药性的逆转提示白血病NK细胞中Fas信号的抑制。特异性目的1的实验旨在确定白血病NK细胞中fas耐药的机制。最初的实验将集中在新的，抑制性诱饵Fas受体的作用，我们已经从白血病NK细胞克隆。MAPK信号传导有助于Fas抗性的假设也将在特定目标1中进行检验。白血病NK细胞显示MAPK的组成性激活；此外，抑制MEK/MAPK可逆转Fas抗性。牛痘病毒中MEK/MAPK和显性阴性（DN） MEK蛋白表达的药理抑制将用于研究MAPK下游信号导致Fas抗性的机制。我们假设白血病NK细胞是激活的细胞毒性细胞，依赖于体内靶识别产生的生存信号。初步数据表明，在NK细胞系中，连接未知的NK激活受体可诱导MAPK激活并调节Fas敏感性。特异性目的2的实验将研究MAPK激活上游NK受体信号传导的机制。特异性NK受体负责保护白血病NK细胞免于fas诱导的死亡将被确定。具体目的3的实验将检验组成型STAT信号调节凋亡抵抗的假设。初步数据显示，组成型STAT在白血病NK细胞中活化；此外，抑制STAT激活直接诱导白血病NK细胞凋亡，并赋予fas敏感性。实验策略将利用酪氨酸激酶的特定抑制剂来调节STAT激活以及DN- STAT蛋白。这些研究对了解NK白血病的发病机制具有重要意义。这些研究的结果应该确定MAPK和STAT信号通路中的分子靶点，这对血液系统恶性肿瘤的治疗发展很重要。

DESCRIPTION (provided by applicant): The broad long-term objective of this proposal is to understand the pathogenesis of NK leukemia. The mechanisms responsible for expansion of leukemic NK cells are not known. The central hypothesis of this proposal is that NK leukemia results from dysregulated apoptosis. Leukemic NK cells are relatively resistant to Fas-mediated death despite expressing high levels of both Fas receptor and Fas ligand. Reversal of Fas resistance after IL-2 activation suggests inhibition of Fas signaling in leukemic NK cells. Experiments in specific aim 1 are directed at determining mechanisms of Fas-resistance in leukemic NK cells. Initial experiments will focus on the role of novel, inhibitory decoy Fas receptors, which we have cloned from leukemic NK cells. The hypothesis that MAPK signaling contributes to Fas resistance will also be examined in specific aim 1. Leukemic NK cells show constitutive activation of MAPK; furthermore inhibition of MEK/MAPK reverses Fas resistance. Pharmacological inhibition of MEK/MAPK and dominant negative (DN) MEK proteins expressed in vaccinia virus will be utilized to examine mechanisms of MAPK downstream signaling leading to Fas resistance. We hypothesize that leukemic NK cells are activated cytotoxic cells dependent on survival signals resulting from target recognition in vivo. Preliminary data show that ligation of an unknown NK activating receptor induces MAPK activation and modulates Fas sensitivity in an NK cell line. Experiments in specific aim 2 will examine mechanisms of NK receptor signaling upstream of MAPK activation. Specific NK receptors responsible for protecting leukemic NK cells from Fas-induced death will be identified. Experiments in specific aim 3 will examine the hypothesis that constitutive STAT signaling regulates apoptotic resistance. Preliminary data show constitutive STAT activation in leukemic NK cells; furthermore, inhibition of STAT activation directly induces apoptosis in leukemic NK cells as well as conferring Fas-sensitivity. The experimental strategy will utilize specific inhibitors of tyrosine kinases that regulate STAT activation as well as DN- STAT proteins. These studies are important for understanding the pathogenesis of NK leukemia. Results of these studies should identify molecular targets in MAPK and STAT signaling pathways important for therapeutic development in hematologic malignancies.