Flavopiridol and differentiation-inducers in leukemia

黄酮吡醇和白血病分化诱导剂

• 批准号: 6835626
• 负责人: Steven Grant
• 金额: $ 24.94万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-01-01 至 2007-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6835626
• 关键词: apoptosis; biological signal transduction; cell differentiation; cell line; clinical research; cyclin dependent kinase; enzyme inhibitors; flavopiridol; human subject; leukemia; mitogen activated protein kinase; oncoprotein p21; protein localization; transfection

DESCRIPTION (provided by applicant): The goal of this proposal is to develop a rational basis for a novel antileukemic strategy combining pharmacologic cyclin-dependent kinase (CDK) inhibitors with clinically relevant differentiation-inducers (DIs). This approach has been prompted by the discovery that flavopiridol (FP; NSC 649890), a potent CDK ATP-binding domain inhibitor, fails to lower the maturation threshold for human leukemia cells (e.g., U937, HL-60) despite promoting cell cycle arrest; instead, it interacts synergistically with a variety of DIs, e.g., PMA, bryostatin 1, and multiple histone deacetylase inhibitors (HDIs), including sodium butyrate (SB) and SAHA, to trigger mitochondrial damage and apoptosis. Combination of FP with DIs is associated with dysregulation of multiple signaling and cell cycle regulatory pathways, including interference with the induction of the endogenous CDK inhibitor p21 CIPI, degradation of p27KIP1, accelerated dephosphorylation and proteolysis of pRb, derepression of E2F, and enhanced activation of p42/44 MAPK. The specific aims of this proposal are (1) to test the hypothesis, using p21 CIP1 antisense, nuclear localization signal and CDK binding domain mutants, as well as inducible constructs, that dysregulation of this CDKI contributes functionally to FP/DI-induced apoptosis; (2) to determine what role, if any, dysregulation of pRb dephosphorylation, cleavage, and E2F activation play in potentiation of DI-associated apoptosis by FP; and (3) to investigate, using stable transfectants ectopically expressing Bcl-2/Bcl-xL as well as phosphorylation-deficient mutants, mechanisms by which FP and DIs circumvent the block to mitochondrial damage and apoptosis conferred by these proteins. For each of these studies, stable cell lines inducibly expressing Raf-1 or MEK1 will be employed to define the role of the PKC/Raf/MEKIMAPK cascade in FP/DI-mediated lethality. Lastly, the effect of combinations of FP with various DIs will be compared in primary human leukemic blasts and normal progenitors (e.g., CD34+, DR-, 71-) to establish whether a basis for therapeutic selectivity exists. It is hoped that information derived from this proposal will lay the foundation for the implementation of a novel therapeutic strategy combining pharmacologic CDK inhibitors with DIs in the treatment of leukemia and possibly other hematologic malignancies.

描述（由申请人提供）：本提案的目标是开发一个 一种新的抗白血病策略的合理基础， 具有临床相关性的细胞周期蛋白依赖性激酶（CDK）抑制剂 分化诱导剂（DI）。这种做法是由 发现了一种有效的CDK ATP结合结构域flavopiridol（FP; NSC 649890）， 抑制剂，未能降低人类白血病细胞的成熟阈值 (e.g., U937，HL-60），尽管促进细胞周期阻滞;相反，它与细胞周期阻滞剂相互作用。 与各种DI协同，例如，PMA、苔藓抑素1和多种 组蛋白脱乙酰酶抑制剂（HDIs），包括丁酸钠（SB）和SAHA， 引发线粒体损伤和细胞凋亡FP与DI的组合是 与多种信号转导和细胞周期调节的失调有关 途径，包括干扰内源性CDK的诱导 抑制剂p21 CIPI，降解p27 KIP 1，加速去磷酸化， pRb的蛋白水解，E2 F的去抑制，以及p42/44的增强激活 MAPK。本建议的具体目的是（1）检验假设，使用 p21 CIP 1反义、核定位信号和CDK结合结构域突变体， 以及诱导型结构，这种CDKI的失调有助于 功能上与FP/DI诱导的细胞凋亡有关;（2）确定FP/DI在细胞凋亡中的作用，如果有的话， pRb去磷酸化、切割和E2 F激活的失调在 FP对DI相关凋亡的增强作用;以及（3）使用 异位表达Bcl-2/Bcl-xL以及 磷酸化缺陷突变体，FP和DI规避的机制 这些蛋白质对线粒体损伤和凋亡的阻断。 对于这些研究中的每一个，诱导表达Raf-1或MEK 1的稳定细胞系 将被用来定义PKC/Raf/MEKIMAPK级联反应的作用， FP/DI介导的致死率。最后，FP与各种药物组合的效果 将在原代人白血病原始细胞和正常祖细胞中比较DI (e.g., CD 34+、DR-、71-），以确定是否为治疗性 存在选择性。希望从这一建议中得到的信息 将为新的治疗策略的实施奠定基础 联合药物CDK抑制剂和DI治疗白血病 可能还有其他血液恶性肿瘤