Leukemia stem cell regulation and resistance

项目概要/摘要 慢性粒细胞白血病（CML）是由BCR-1基因的造血干细胞（HSC）转化引起的。 ABL酪氨酸激酶。酪氨酸激酶抑制剂（TKI）在诱导缓解和延长生存期方面有效 在慢性粒细胞白血病患者，但未能消除原始白血病干细胞（LSC），可以再生疾病。最 患者需要持续TKI治疗以维持缓解，并保持毒性、经济困难和 不遵守。我们研究的长期目标是提高对LSC机制的理解 对治疗的耐药性，以支持制定有效和安全的LSC靶向策略，并加强 CML患者的无治疗缓解的可能性。线粒体代谢在细胞内起着重要的调节作用， 正常HSC功能的作用。CML LSC显示线粒体氧化磷酸化增加 （OXPHOS）与正常HSC中的低OXPHOS相比。然而，线粒体也发挥重要作用， 代谢过程，包括脂肪酸、谷氨酰胺和葡萄糖氧化，以及 生物合成中间体我们研究的基本原理是， 导致LSC生长和TKI耐药性改变的原因尚不清楚。我们的初步研究显示 TKI治疗后CML LSC中的OXPHOS增加，但随后OXPHOS恢复，脂肪酸增加 氧化（FAO），并继续处理。SIRT 1、P53和MYC调控网络在这一过程中发挥重要作用。 在LSC传播中。我们发现SIRT 1及其靶点PGC-1α在OXPHOS增加中起重要作用。 CML LSC。PPARa是一种PGC-1α共激活的转录因子，是FAO的关键调节因子， TKI治疗后CML LSC中的OXPHOS表达增加，并有助于增加OXPHOS、增殖和存活。 我们将探讨BCR-ABL激酶抑制后FAO增加的假设， 维持高水平的OXPHOS、糖酵解和β-氨基丁酸解有助于CML中的TKI耐药性 LSC，代谢调节机制是消除TKI治疗CML的潜在靶点 LSC。在具体目标1中，我们将使用基因表达、细胞外通量、代谢物谱分析和 体外和体内代谢标记研究TKI治疗对CML LSC中线粒体代谢的影响， 研究SIRT 1、PGC 1a和PPARa在代谢改变中的作用，并研究MYC和p53的相互作用 线粒体代谢的调控网络。在具体目标2中，我们将研究增加 OXPHOS和FAO促进CML LSC的TKI耐药性。骨髓微环境小生境发挥着 在维持静止的TKI耐药LSC群体中起关键作用。然而，微环境的作用 在LSC生长的代谢调节中的作用尚不清楚，将在此进行评价。这些研究意义重大 因为他们预期鉴定CML LSC中TKI抗性的代谢调节机制， 在CML LSC中建立代谢和其他调节机制之间的联系，并识别新的 治疗的目标这里发展的概念将对其他恶性肿瘤产生广泛的影响。

PROJECT SUMMARY/ABSTRACT Chronic myelogenous leukemia (CML) results from hematopoietic stem cell (HSC) transformation by the BCR- ABL tyrosine kinase. Tyrosine kinase inhibitors (TKI) are effective in inducing remission and prolonging survival in CML patients, but fail to eliminate primitive leukemia stem cells (LSC) that can regenerate disease. Most patients need ongoing TKI treatment to maintain remission, and remain at risk of toxicity, financial hardship and non-adherence. The long-term goal of our research is to improve understanding of mechanisms of LSC resistance to treatment, to support development of effective and safe strategies for LSC targeting, and enhance possibilities of treatment-free remissions in CML patients. Mitochondrial metabolism plays a critical regulatory role in normal HSC function. CML LSC demonstrate increased mitochondrial oxidative phosphorylation (OXPHOS) compared to low OXPHOS in normal HSC. However, mitochondria also play important roles in metabolic processes besides OXPHOS, including fatty acid, glutamine and glucose oxidation, and generation of biosynthetic intermediates. The rationale for our studies is that specific mitochondrial metabolic alterations that contribute to altered LSC growth and TKI resistance are not known. Our preliminary studies show initial inhibition of OXPHOS in CML LSC after TKI treatment, but subsequent restoration of OXPHOS, and increased fatty acid oxidation (FAO), with continued treatment. A SIRT1, P53 and MYC regulatory network plays an important role in LSC propagation. We show that SIRT1 and its target PGC-1α play an important role in increased OXPHOS in CML LSC. PPARa, a PGC-1α-coactivated transcription factor and a key regulator of FAO, shows increased expression in CML LSC after TKI treatment, and contributes to increased OXPHOS, proliferation and survival. We will explore the hypothesis that increased FAO following BCR-ABL kinase inhibition, together with maintenance of high levels of OXPHOS, glycolysis and glutaminolysis, contributes to TKI resistance in CML LSC, and that metabolic regulatory mechanisms represent potential targets for elimination of TKI-treated CML LSC. In Specific Aim 1 we will use a combination of gene expression, extracellular flux, metabolite profiling and in vitro and in vivo metabolic labeling to study effects of TKI treatment on mitochondrial metabolism in CML LSC, examine the role of SIRT1, PGC1a and PPARa in metabolic alterations, and study interactions of MYC and p53 regulatory networks with mitochondrial metabolism. In Specific Aim 2 we will investigate the role of increased OXPHOS and FAO in promoting TKI resistance in CML LSC. Bone marrow microenvironment niches play a critical role in maintaining quiescent, TKI-resistant LSC populations. However, the role of the microenvironment in metabolic regulation of LSC growth is not known, and will be evaluated here . These studies are significant since they are expected to identify mechanisms of metabolic regulation underlying TKI resistance in CML LSC, establish connections between metabolism and other regulatory mechanisms in CML LSC, and identify new targets for therapy. The concepts developed here will have broad implications for other malignancies.