Genomic instability causes imatinib resistance in CML

描述（由申请人提供）：Bcr/abl基因来源于t（9;22）相互易位，存在于大多数慢性髓性白血病（CML）和急性淋巴细胞白血病（ALL）患者中。BCR/ABL致癌酪氨酸激酶调节对DNA损伤的反应，诱导对基因毒性治疗的抗性。此外，BCR/ABL刺激基因组不稳定，这可能导致BCR/ABL激酶突变，从而对甲磺酸伊马替尼产生耐药性。我们假设：BCR/ABL升高活性氧（ROS）的水平，从而诱导“自发”DNA损伤（例如尿嘧啶残基），其不可靠的修复引入了BCR/ABL激酶结构域的氨基酸取代，从而导致对IM的抗性。在CML造血干细胞（HSC）、普通髓系祖细胞（CMP）和粒细胞/巨噬细胞祖细胞（GMP）中，利用抗氧化方法和体外和体内BCR/ABL白血病发生模型，研究ROS在氧化性DNA损伤导致突变和甲磺酸伊马替尼耐药中的作用。BCR/ABL白血病细胞中ros依赖性氧化DNA损伤机制的效率和保真度将通过研究碱基切除修复（BER）来确定，重点是UDG糖基化酶去除尿嘧啶残基。这些反应将使用定义良好的报告/底物系统和不同方法的组合进行检查，包括免疫荧光，磷酸化和相互作用缺失突变体，转基因小鼠和测序。BCR/ABL-UDG功能相互作用将通过诱变和适体靶向研究来抑制对甲磺酸伊马替尼的耐药性。甲磺酸伊马替尼（IM）彻底改变了BCR/ abl阳性白血病的治疗，如慢性髓性白血病（CML）和一系列急性淋巴细胞白血病（ALL）。不幸的是，活性氧（ROS）会导致BCR/ABL激酶结构域的氨基酸取代，从而导致对IM的抗性。我们将确定抗氧化剂是否能够减少突变和IM耐药性。此外，我们将研究碱基切除修复在这些突变产生中的作用。我们的研究结果可能会导致更好的治疗BCR/ abl阳性白血病。

DESCRIPTION (provided by applicant): Bcr/abl gene is derived from the t(9;22) reciprocal translocation and is present in most of chronic myelogenous leukemia (CML) and a cohort of acute lymphoblastic leukemia (ALL) patients. BCR/ABL oncogenic tyrosine kinase modulates response to DNA damage inducing resistance to genotoxic therapies. In addition BCR/ABL stimulates genomic instability, which may lead to mutations in BCR/ABL kinase causing resistance to imatinib mesylate. We hypothesize that: BCR/ABL elevates the levels of reactive oxygen species (ROS) which induce "spontaneous" DNA lesions (for example uracil residues), whose unfaithful repair introduces amino acid substitutions in the BCR/ABL kinase domain causing resistance to IM. The role of ROS in generation of oxidative DNA damage leading to mutagenesis and resistance to imatinib mesylate will be studied in CML hematopoietic stem cells (HSC), common myeloid progenitor cells (CMP), and granulocyte/macrophage progenitor cells (GMP) using anti-oxidant approaches and in vitro and in vivo models of BCR/ABL leukemogenesis. The efficiency and fidelity of the mechanisms processing ROS-dependent oxidative DNA damage in BCR/ABL leukemia cells will be determined by studying base excision repair (BER), focusing on UDG glycosylase removing uracil residues. These reactions will be examined using well-defined reporter/substrate systems and a combination of different approaches including immunofluorescence, phosphorylation-less and interaction- deprived mutants, transgenic mice, and sequencing. BCR/ABL-UDG functional interaction will be investigated by mutagenesis and targeted by aptamers to inhibit resistance to imatinib mesylate. Imatinib mesylate (IM) revolutionized the treatment of BCR/ABL-positive leukemias such as chronic myelogenous leukemia (CML) and a cohort of acute lymphocytic leukemias (ALL). Unfortunately, reactive oxygen species (ROS) cause amino acid substitutions in the BCR/ABL kinase domain resulting in resistance to IM. We will determine if anti-oxidants are able to reduce mutations and IM resistance. In addition, we will examine the role of base excision repair in generation of these mutations. Results of our studies may lead to better treatment of BCR/ABL-positive leukemias.