Role of Redox Dependent Signaling in Leukemia

氧化还原依赖性信号传导在白血病中的作用

• 批准号: 7939273
• 负责人: Joya Chandra
• 金额: $ 12.46万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7939273
• 关键词: Antioxidants; BCR/ABL1; Bcr-Abl tyrosine kinase; Binding Sites; Biological Assay; Blast Phase; Bone Marrow; Cell Proliferation; Cells; Chronic; Chronic Myeloid Leukemia; Chronic Phase of Disease; Clinical; DNA Damage; Data; Disease; Disease Progression; Employee Strikes; Family member; Feedback; Genomic Instability; Goals; Growth; Imatinib; Imatinib mesylate; In Vitro; Link; Measures; Mediating; Messenger RNA; Modality; Modeling; Mus; Mutation; Nature; Oncogenes; Onset of illness; Oxidants; Oxidation-Reduction; Pathway interactions; Patients; Peroxides; Phase; Phosphotransferases; Play; Point Mutation; Production; Proteins; Reactive Oxygen Species; Refractory; Relative (related person); Reporter Genes; Research Personnel; Resistance; Role; Signal Transduction; Specimen; Testing; Therapeutic; Tissue Microarray; Transcriptional Regulation; Transducers; Transplantation; Up-Regulation; bcr-abl Fusion Proteins; cell growth; cell injury; cellular transduction; chemical genetics; deletion analysis; human FRAP1 protein; in vivo; insight; leukemia; overexpression; oxidative damage; prevent; programs; promoter; response; small hairpin RNA; therapeutic target; transcription factor

DESCRIPTION (provided by applicant): CML (chronic myeloid leukemia) is unique in that the BCR/ABL oncogene drives disease onset and progression. Therapies inhibiting the BCR/ABL kinase, such as imatinib mesylate, have dramatically altered the treatment of chronic phase disease, but are less effective in blast crisis and against specific point mutations in BCR/ABL. Blast crisis is a very aggressive phase of the disease characterized by genomic instability and secondary mutations. These features have been linked to DNA damage caused by reactive oxygen species (ROS). We and others find that overexpression of p210 BCR/ABL alone causes an increase in levels of ROS. Our preliminary data reveal that a consequence of this increased level of ROS is upregulation of Fyn, a Src kinase family member. We hypothesize that ROS dependent signaling transduced through Fyn contributes to progression to blast crisis. BCR/ABL overexpressing cells display a four-fold upregulation of Fyn protein. This upregulation is blocked by antioxidants, thus linking increased ROS levels and Fyn expression. Point mutations in BCR/ABL that confer resistance to imatinib mesylate retain upregulation of Fyn, suggesting that Fyn may be an appropriate therapeutic target in refractory patients. Knockdown of Fyn using shRNA slows leukemia cell growth in vitro and in vivo, inhibits clonogenic growth by 45% and causes increased sensitivity to imatinib. These striking changes indicate that Fyn is important for leukemia cell proliferation and sensitivity to imatinib. Blockade of Fyn also lowers the overall levels of intracellular peroxides, suggesting that Fyn can amplify ROS levels, thus contributing to subsequent DNA damage and secondary mutations. These preliminary data place Fyn both downstream and upstream of increased ROS, suggesting the presence of a positive feedback loop, whereby Fyn activation through BCR/ABL leads to increased ROS potentially playing an important role in blast crisis. This proposal will elucidate the mechanism(s) and consequences of oxidant-dependent Fyn upregulation and validate these findings in murine models and in clinical specimens. The specific aims of this proposal are to: #1: Determine the mechanism by which BCR/ABL and ROS levels upregulate Fyn. # 2: Evaluate the effects of Fyn upregulation on ROS dependent proliferative signals and DNA damage. #3: Test the relative contributions of BCR/ABL initiated ROS and Fyn upregulation on progression in CML murine models and patient specimens. These data will provide insight into one significant consequence of BCR/ABL-mediated ROS alterations: Fyn upregulation. Our long-term goal is to understand oxidant-dependent signaling in leukemia in order to devise strategies to prevent progression. Tactics that quelch ROS production, or block downstream signaling through Fyn in BCR/ABL containing cells may provide promising therapeutic modalities.

描述（由申请人提供）：CML（慢性髓样白血病）是独一无二的，因为BCR/ABL癌细胞驱动疾病的发作和进展。抑制BCR/ABL激酶（例如伊马替尼）的疗法已极大地改变了慢性期疾病的治疗，但在BLY危机中的有效性较小，并且对BCR/ABL的特定点突变有效。爆炸危机是以基因组不稳定性和继发性突变为特征的疾病的非常激进的阶段。这些特征与由活性氧（ROS）引起的DNA损伤有关。我们和其他人发现，单独的P210 BCR/ABL的过表达会导致ROS水平增加。我们的初步数据表明，这种ROS水平升高的结果是SRC激酶家族成员Fyn的上调。我们假设通过FYN转导的ROS依赖性信号传导有助于发展爆炸危机。 BCR/ABL过表达的细胞显示FYN蛋白的四倍上调。该上调被抗氧化剂阻断，从而将增加的ROS水平和FYN表达联系起来。 BCR/ABL中赋予伊马替尼甲酸酯抗性的点突变保留了FYN的上调，这表明FYN可能是难治性患者的适当治疗靶标。使用shRNA敲低FYN会在体外和体内减慢白血病细胞的生长，抑制克隆生长的生长45％，并导致对伊马替尼的敏感性提高。这些引人注目的变化表明，FYN对于白血病细胞增殖和对伊马替尼的敏感性很重要。 FYN的阻断还降低了细胞内过氧化物的总体水平，这表明FYN可以扩大ROS水平，从而导致随后的DNA损伤和继发突变。这些初步的数据位置在下游和上游ROS增加，这表明存在正反馈循环，从而通过BCR/ABL激活FYN会导致ROS增加在爆炸危机中可能发挥重要作用。该建议将阐明氧化剂依赖性FYN上调的机制和后果，并在鼠模型和临床标本中验证这些发现。该提案的具体目的是：＃1：确定BCR/ABL和ROS水平上调FYN的机制。 ＃2：评估FYN上调对ROS依赖性增殖信号和DNA损伤的影响。 ＃3：测试BCR/ABL启动ROS和FYN上调对CML鼠模型和患者标本中进展的相对贡献。这些数据将为BCR/ABL介导的R​​OS改变的一个重要结果提供洞察力：FYN上调。我们的长期目标是了解白血病中氧化剂依赖性信号传导，以制定防止进展的策略。在含有BCR/ABL的细胞中通过FYN阻止产生或通过FYN阻止下游信号传导的策略可能会提供有希望的治疗方式。