Survival Mechanisms in Leukemic NK Cells

白血病 NK 细胞的生存机制

• 批准号: 8828338
• 负责人: Thomas Patrick Loughran
• 金额: $ 25.09万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8828338
• 关键词: Animal Model; Apoptosis; Apoptotic; Autophagocytosis; Binding; CD94 Antigen; Cell Death; Cell Survival; Cells; Cellular biology; Ceramides; Chronic; Color; Data; Disease; Disease remission; Drug Formulations; Enzymes; Equilibrium; Funding; Genetic; Goals; Immune system; In Vitro; Inbred F344 Rats; Induction of Apoptosis; Infection; Killer Cells; Lead; Leukemic Cell; Lipids; Mediating; Metabolism; Modeling; N-caproylsphingosine; Natural Killer Cells; Pathogenesis; Pathway interactions; Phenotype; Phosphorylation; Production; Proteins; ROCK1 gene; Research; Shapes; Signal Induction; Signal Transduction; Sphingolipids; Sphingosine-1-Phosphate Receptor; System; Testing; Therapeutic; Therapeutic Intervention; Treatment Efficacy; Up-Regulation; Work; design; extracellular; galactosylgalactosylglucosylceramidase; in vivo; inhibition of autophagy; leukemia; nanoliposome; novel strategies; novel therapeutic intervention; overexpression; response; sphingosine 1-phosphate; sphingosine-1-phosphate lyase

DESCRIPTION (provided by applicant): The long term goal of this project is to develop better therapeutics for NK-LGL leukemia. The proposal represents a new approach to leukemia pathogenesis by positing that imbalances in the sphingolipid rheostat confer the leukemia phenotype. This model postulates that cell fate is determined by the balance between pro- survival (sphingosine-1-phosphate, S1P) and pro-apoptotic (ceramide) lipids. Preliminary data demonstrate altered sphingolipid metabolism in this disease, including increased levels of S1P and decreased levels of ceramide in leukemic LGL; increased expression of acid ceramidase; and findings that stable knockdown of acid ceramidase enhances production of long chain ceramide and results in decreased viability of leukemic LGL. Furthermore, we show that targeting sphingolipid signaling represents a new avenue for therapeutic intervention in this disease. We achieve complete remission of aggressive NK-LGL leukemia by utilizing nanoliposomal formulation of short chain C6 ceramide or the S1P receptor antagonist, FTY720, in the Fischer rat animal model. We will test the hypothesis that S1PR5 signaling mediates survival of leukemic NK cells (Specific Aim 1). Knockdown of S1PR5, the predominant S1P receptor expressed on leukemic NK cells, inhibits constitutive ERK phosphorylation and induces apoptosis. Constitutive overexpression of Rac1, PAK3, RhoB and ROCK1, potential downstream effectors of S1PR5, is observed in leukemic NK cells. Pharmacological inhibition of Gi, RAC1 and ROCK also results in cell death in leukemic NK cells, demonstrating that these targets may be involved in S1PR5-dependent survival pathways. Specific Aim 2 will test the hypothesis that FTY720 induces both apoptosis and autophagy in leukemic NK cells by targeting sphingolipid signaling. We show that pharmacological inhibition of autophagy increases apoptosis induced by FTY720, indicating that blockade of autophagy potentiates FTY720 therapeutic efficacy in leukemic NK cells. FTY720 treatment of leukemic LGL leads to accumulation of intracellular S1P, suggesting that autophagy induced by FTY720 might occur through inhibition of S1P lyase. In addition, we postulate that leukemic NK cells are dependent on survival signaling resulting from NK receptor target recognition in vivo. Convincing preliminary data indicate that sphingolipid survival pathways are activated in normal NK cells after target binding in vitro. Findings similar to those seen in NK-LGL leukemia include upregulation of S1PR5, identification of RhoB, ROCK1, Rac1 and PAK3 as downstream components of S1PR5 signaling, and induction of apoptosis by FTY720. We anticipate that the proposed research will have significant impact on the field of NK cell biology. Our work will define new pathways important for survival of both leukemic and activated normal NK cells. We expect that understanding sphingolipid survival signaling will lead to novel therapeutic approaches for an incurable illness.

描述（由申请人提供）：该项目的长期目标是开发更好的 NK-LGL 白血病治疗方法。该提案提出了一种研究白血病发病机制的新方法，认为鞘脂变阻器的不平衡会导致白血病表型。该模型假设细胞命运由促生存（1-磷酸鞘氨醇，S1P）和促凋亡（神经酰胺）脂质之间的平衡决定。初步数据表明，这种疾病中的鞘脂代谢发生了改变，包括白血病 LGL 中 S1P 水平升高和神经酰胺水平降低；酸性神经酰胺酶表达增加；研究结果表明，酸性神经酰胺酶的稳定敲低可增强长链神经酰胺的产生，并导致白血病 LGL 的活力下降。此外，我们表明靶向鞘脂信号传导代表了治疗干预该疾病的新途径。我们在 Fischer 大鼠动物模型中利用短链 C6 神经酰胺或 S1P 受体拮抗剂 FTY720 的纳米脂质体制剂，实现了侵袭性 NK-LGL 白血病的完全缓解。 我们将检验 S1PR5 信号传导介导白血病 NK 细胞存活的假设（具体目标 1）。 S1PR5（白血病 NK 细胞上表达的主要 S1P 受体）的敲低可抑制 ERK 的持续磷酸化并诱导细胞凋亡。在白血病 NK 细胞中观察到 Rac1、PAK3、RhoB 和 ROCK1（S1PR5 的潜在下游效应子）的组成型过度表达。 Gi、RAC1 和 ROCK 的药理学抑制也会导致白血病 NK 细胞的细胞死亡，表明这些靶标可能参与 S1PR5 依赖性生存途径。具体目标 2 将测试 FTY720 通过靶向鞘脂信号传导诱导白血病 NK 细胞凋亡和自噬的假设。我们发现，自噬的药理抑制会增加 FTY720 诱导的细胞凋亡，表明自噬的阻断增强了 FTY720 对白血病 NK 细胞的治疗功效。 FTY720 治疗白血病 LGL 会导致细胞内 S1P 积累，表明 FTY720 诱导的自噬可能是通过抑制 S1P 裂合酶而发生的。此外，我们假设白血病 NK 细胞依赖于体内 NK 受体靶标识别产生的生存信号。令人信服的初步数据表明，在体外靶标结合后，正常 NK 细胞中的鞘脂存活途径被激活。与 NK-LGL 白血病相似的发现包括 S1PR5 上调，RhoB、ROCK1、Rac1 和 PAK3 被鉴定为 S1PR5 信号下游成分，以及 FTY720 诱导细胞凋亡。 我们预计拟议的研究将对 NK 细胞生物学领域产生重大影响。我们的工作将定义对白血病细胞和活化的正常 NK 细胞的生存都很重要的新途径。我们期望了解鞘脂存活信号传导将为治疗不治之症带来新的治疗方法。