Targeting DNA repair to eradicate TKI-refractory/resistant CML

靶向 DNA 修复以根除 TKI 难治性/耐药性 CML

• 批准号: 8702641
• 负责人: TOMASZ SKORSKI
• 金额: $ 53.77万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-12 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8702641
• 关键词: ABL1 gene; Affect; Aggressive-Phase Myeloid Leukemia; Apoptotic; Autophagocytosis; BRCA1 gene; Blast Phase; Blood; CD34 gene; Cells; Chronic Myeloid Leukemia; Chronic Phase; Clinical Trials; Cytogenetics; DNA Binding; DNA Double Strand Break; DNA Repair; DNA lesion; DNA-PKcs; Dasatinib; Defect; Diagnosis; Disease; Double Strand Break Repair; Environment; Erinaceidae; Generations; Genetic Models; Goals; Growth; Health; Hematopoietic stem cells; Histones; Imatinib; Knockout Mice; Malignant Neoplasms; Mediating; Modality; Molecular; Mutation; Nonhomologous DNA End Joining; Normal Cell; Normal tissue morphology; Oncogenes; Other Genetics; Outcome; Pathway interactions; Patients; Peptide aptamers; Phosphotransferases; Population; Population Study; Proliferating; Protein Phosphatase 2A Regulatory Subunit PR53; Proteins; Reactive Oxygen Species; Recurrent disease; Refractory; Reporting; Resistance; Running; Source; Staging; Stem cells; Testing; Therapeutic; Time; Tissues; Transgenic Mice; Tyrosine Kinase Inhibitor; clinical application; cytokine; homologous recombination; improved; inhibitor/antagonist; leukemia; leukemic stem cell; mutant; novel; recombinational repair; repaired; response; small molecule; stem; tumor; tyrosine kinase ABL1

DESCRIPTION (provided by applicant): Chronic myeloid leukemia in chronic phase (CML-CP) is initiated by t(9;22) encoding for p210BCR-ABL1 tyrosine kinase that transforms hematopoietic stem cells (HSCs). CML-CP is leukemia stem cells (LSCs) - derived disease, but deregulated growth of LSCs-derived leukemia progenitor cells (LPCs) leads to the manifestation of the disease. CML-CP may progress to more advanced accelerated phase (CML-AP), and subsequently to a very aggressive blast phase (CML-BP). Most CML-CP patients are currently treated with tyrosine kinase inhibitors (TKIs) such as imatinib, dasatinib and nilotinib. However, it is unlikely that TKIs will "cure" CML patients due to the presence of TKI- refractory cells (e.g, quiescent LSCs) and TKI-resistant cells (e.g., proliferating LSCs/LPCs carrying BCR- ABL1 kinase T315I mutant). In addition, population studies revealed that overall only 51% of CML-CP patients respond favorably to TKI treatment. Therefore, novel treatment modalities are needed to eradicate TKI- refractory/resistant CML cells in the responding patients and also to treat patients who do not respond favorably to TKIs. To cure CML these strategies should simultaneously target two fundamentally different leukemia cell populations: TKI-refractory quiescent LSCs and TKI-resistant/refractory proliferating LSCs/LPCs. We found that CML LSCs and LPCs, including quiescent LSCs accumulate 2-4 times more reactive oxygen species (ROS)-induced DNA double strand breaks (DSBs) than normal counterparts (Cramer et al., Cancer Res., 2008; Nieborowska-Skorska et al., Blood, 2012; Bolton-Gillespie et al., Blood, 2013). DSBs are the most lethal DNA lesions. We reported that CML cells can tolerate high numbers of DSBs because two major repair mechanisms, homologous recombination repair (HRR) and non-homologous end-joining (NHEJ) are hyper- activated (Slupianek et al., Mol. Cell, 2001; Oncogene, 2005; DNA Repair, 2006; Cancer Res., 2011; Blood, 2011; Nowicki et al., Blood, 2005). CML cells are "addicted" to these pathways to survive pro-apoptotic challenge from high numbers of lethal DSBs. However, there are critical differences between DSB repair in normal and CML cells. Proliferating LSCs/LPCs employ RAD52-dependent HRR, in contrast to BRCA1-mediated HRR in normal counterparts. Quiescent LSCs use PARP1-mediated NHEJ instead of DNA-PKcs -dependent NHEJ, which is predominant in normal quiescent HSCs. We will explore these differences to target leukemia-specific DNA repair mechanisms simultaneously in quiescent LSCs and proliferating LSCs/LPCs to achieve "dual synthetic lethality", with negligible effect on normal cells and tissues. According to our best knowledge the concept of "dual synthetic lethality" was not tested before. "Dual synthetic lethality" will be induced in TKI-treated CML-CP/AP cells by simultaneous targeting of RAD52 and PARP1 using recently identified candidate small molecule inhibitors interrupting key functions of RAD52 and PARP1: RAD52 DNA binding activity and stimulation of PARP1 by histone 4. In addition, using CML-CP -like transgenic mice, Rad52-/-Parp1-/- double knockout mice, mutagenic approach, peptide aptamers, and CML-CP/AP primary cells we will determine if other RAD52 and/or PARP1 activities could be targeted to trigger more efficient "dual synthetic lethality" simultaneously in TKI-refractory quiescent LSCs and TKI-resistant proliferating LSCs/LPCs. Our long-term plan is to run a clinical trial testing the possibility to eradicate CML-CP/AP by induction of "dual synthetic lethality" in TKI-treated patients.

描述（由申请人提供）：慢性期慢性粒细胞白血病 (CML-CP) 由编码 p210BCR-ABL1 酪氨酸激酶的 t(9;22) 引发，该激酶可转化造血干细胞 (HSC)。 CML-CP 是白血病干细胞 (LSC) 衍生的疾病，但 LSC 衍生的白血病祖细胞 (LPC) 生长失调会导致该疾病的表现。 CML-CP 可能进展到更高级的加速期 (CML-AP)，随后进入非常侵袭性的急变期 (CML-BP)。大多数 CML-CP 患者目前接受酪氨酸激酶抑制剂 (TKI) 治疗，例如伊马替尼、达沙替尼和尼罗替尼。然而，由于 TKI 难治性细胞（例如静止的 LSC）和 TKI 耐药细胞（例如携带 BCR-ABL1 激酶 T315I 突变体的增殖性 LSC/LPC）的存在，TKI 不太可能“治愈”CML 患者。此外，人群研究显示，总体而言，只有 51% 的 CML-CP 患者对 TKI 治疗有良好反应。因此，需要新的治疗方式来根除有反应的患者中 TKI 难治/耐药的 CML 细胞，并治疗对 TKI 没有良好反应的患者。为了治愈 CML，这些策略应同时针对两种根本不同的白血病细胞群：TKI 难治性静态 LSC 和 TKI 耐药/难治性增殖 LSC/LPC。我们发现，CML LSC 和 LPC（包括静止 LSC）积累的活性氧 (ROS) 诱导的 DNA 双链断裂 (DSB) 比正常对应物多 2-4 倍（Cramer 等人，Cancer Res.，2008；Nieborowska-Skorska 等人，Blood，2012；Bolton-Gillespie 等人，Blood，2013）。 DSB 是最致命的 DNA 损伤。我们报道CML细胞可以耐受大量的DSB，因为两种主要的修复机制，同源重组修复（HRR）和非同源末端连接（NHEJ）被过度激活（Slupianek et al., Mol. Cell, 2001; Oncogene, 2005; DNA Repair, 2006; Cancer Res., 2011; Blood, 2011; Nowicki et al.,血，2005）。 CML 细胞对这些途径“上瘾”，以在来自大量致命 DSB 的促凋亡挑战中生存。然而，正常细胞和 CML 细胞中的 DSB 修复之间存在重大差异。增殖的 LSC/LPC 采用 RAD52 依赖性 HRR，这与正常对应物中 BRCA1 介导的 HRR 不同。静态 LSC 使用 PARP1 介导的 NHEJ，而不是 DNA-PKcs 依赖性 NHEJ，后者在正常静态 HSC 中占主导地位。我们将探索这些差异，以同时针对静止LSC和增殖LSC/LPC中的白血病特异性DNA修复机制，以实现“双重合成致死”，而对正常细胞和组织的影响可以忽略不计。据我们所知 以前没有测试过“双重合成致死率”的概念。通过同时靶向 RAD52 和 PARP1，使用最近确定的干扰 RAD52 和 PARP1 关键功能的候选小分子抑制剂，将在 TKI 处理的 CML-CP/AP 细胞中诱导“双重合成致死”：RAD52 DNA 结合活性和组蛋白 4 对 PARP1 的刺激。此外，使用 CML-CP 样转基因小鼠， Rad52-/-Parp1-/- 双敲除小鼠、诱变方法、肽适体和 CML-CP/AP 原代细胞，我们将确定是否可以针对其他 RAD52 和/或 PARP1 活性，在 TKI 难治性静态 LSC 和 TKI 抗性增殖 LSC/LPC 中同时触发更有效的“双重合成致死”。我们的长期计划是开展一项临床试验，测试通过在 TKI 治疗的患者中诱导“双重合成致死率”来根除 CML-CP/AP 的可能性。