Epigenetic Targeted Therapy for Chronic Lymphocytic Leu*

描述(申请人提供)：关键的肿瘤抑制基因的表观遗传转录沉默在许多恶性肿瘤中的重要性已经得到了很好的证实。最近几年，人们开始了解导致这种转录沉默的分子机制，导致了DNA甲基化与核组蛋白修饰以动态方式相互作用的概念，从而抑制或增强转录。我们的染色质重塑团队积极参与这项提议，在慢性淋巴细胞白血病(CLL)的临床前工作已经证明了这些基因沉默的机制具有临床意义。具体地说，我们发现与正常B细胞相比，CLL患者样本中异常甲基化的数量显著变化，特定基因甲基化增加了1%到6%。此外，我们还证明了一些特定的基因，如真皮表达-1(dermo-1)、twist和代谢型谷氨酸受体7(GRM7)，在来自慢性淋巴细胞性白血病患者的原代肿瘤细胞中存在差异甲基化。此外，dermo-1和twist与特定的VH基因亚型和既往治疗状态有关。此外，去甲基化药物地西他滨处理体外培养的CLL细胞可促进DNA上DNA甲基转移酶1(DNMT1)的捕获和游离DNMT1蛋白的耗尽，随后基因重新表达和caspase依赖的细胞凋亡。基于这一点以及我们之前对包括丙戊酸在内的组蛋白脱乙酰酶抑制剂所做的工作，我们试图验证这样一个总体假设，即在CLL中应用针对染色质的表观遗传治疗将解除肿瘤抑制基因的异常转录抑制，恢复细胞增殖、分化和凋亡的正常模式，并最终使CLL患者临床受益。这项建议的具体目的是：1)在氟达拉滨耐药的慢性淋巴细胞白血病患者中进行地西他滨的最小有效药理剂量(MEPD)研究，然后地西他滨与丙戊酸联合使用，以及2)作为目标1临床试验的一部分，同时进行MEPD指导的研究和详细的药理学和药效学研究。这项研究将提供足够的初步数据，作为单独应用的一部分，稍后进行随机II期研究，以确定这两种治疗方法在氟达拉滨难治性CLL中的临床和基因再表达有效性。此外，这项建议的完成将提供关于CLL中DNMT1抑制的动力学和相关基因重新表达的知识，以便在未来寻求表观遗传靶向治疗的替代组合。

DESCRIPTION (provided by applicant): The importance of epigenetic transcriptional silencing of key tumor suppressor genes in many malignancies has been well established. The molecular mechanisms leading to this transcriptional silencing has recently begun to be understood over the last few years, leading to the concept that DNA methylation interacts in a dynamic way with nuclear histone modifications to either repress or enhance transcription. Pre-clinical work in chronic lymphocytic leukemia (CLL) by our chromatin remodeling team actively involved in this proposal has demonstrated these mechanisms of gene silencing is clinically relevant. Specifically, we have demonstrated marked variation in the amount of aberrant methylation in CLL patient samples ranging from 1% to 6% increase in specific gene methylation as compared to normal B cells. In addition, we have demonstrated that selected genes such as Dermis expressed-1 (DERMO-1), TWIST, and the metabotrobic glutamate receptor 7 (GRM7) are differentially methylated in primary tumor cells derived from patients with CLL. In addition, DERMO-1 and TWIST are associated with specific VH gene subtypes and prior treatment status. Furthermore, treatment of CLL cells with the hypomethylating agent decitabine promotes trapping of DNA methyltransferase 1 (DNMT1) on DNA and depletion of free DNMT1 protein with subsequent gene re-expression and caspase dependent apoptosis in CLL cells in vitro. Based upon this and our previous work with histone deacetylase inhibitors including valproic acid, we seek to test the overall hypothesis that application of epigenetic therapy targeting chromatin in CLL will relieve aberrant transcriptional repression of tumor suppressor genes, restore normal patterns of cell proliferation, differentiation and apoptosis and ultimately result in clinical benefit to patients with CLL. The specific aims of this proposal are: 1) to perform a minimal effective pharmacologic dose (MEPD) finding study of decitabine and then decitabine combined with valproic acid in fludarabine-refractory CLL patients, and 2) to perform the MEPD-directed studies concurrent with detailed pharmacologic and pharmacodynamic studies as part of the aim 1 clinical trial. This study will provide sufficient preliminary data to later pursue, as part of a separate application, a randomized phase II study to determine the clinical and gene re-expression efficacy of these two therapeutic approaches in fludarabine-refractory CLL. In addition, completion of this proposal will afford knowledge of the kinetics of DNMT1 inhibition in CLL and associated gene re-expression to allow pursuit of alternative combinations of epigenetically targeted therapies in the future.