抑制ATR协同增强阿糖胞苷杀伤急性髓系白血病细胞活性的机制研究
发布时间:2018-06-29 09:11
本文选题:急性髓系白血病(AML) + ATR ; 参考:《吉林大学》2017年硕士论文
【摘要】:急性髓系白血病(acute myeloid leukemia,AML)是最为常见的急性白血病,儿童患者的生存率为65%左右,而成人患者的生存率则仅为25%左右。阿糖胞苷在过去40年里一直是治疗AML最有效的临床一线药物之一。尽管对多数病人来说传统化疗能使病情达到完全缓解,但是这些病人的大多数会复发。AML细胞对阿糖胞苷的耐药是应用阿糖胞苷治疗AML失败的主要原因。因此急需一种新的、更加有效的治疗方案来对抗这种恶性疾病。导致阿糖胞苷产生耐药的一个主要机制就是化疗导致的DNA损伤激活了细胞DNA损伤应答(the DNA damage response,DDR)信号网络。ATR(ataxia telangiectasia and Rad3 relate)和ATM(ataxia telangiectasia mutated)是DDR信号网络中两个重要的组成蛋白。当细胞发生DNA双链断裂或者复制叉停滞时,细胞会对其进行修复并产生单链DNA,此时ATR就会被激活以应对单链DNA的产生。ATR能够调节多种重要的生物学功能,如调节DNA损伤修复、缓解DNA复制压力和激活细胞周期检验点等。ATR是促进DNA受损伤细胞存活的重要调节因子;此外大多数的癌细胞因G1细胞周期检验点缺失而更加依赖ATR调节的S和G2/M细胞周期检验点。因此抑制ATR能够增强DNA损伤类药物对AML细胞的杀伤活性。ATR抑制剂与吉西他滨、顺铂以及PARP(poly(ADP-ribose)polymerase)抑制剂等DNA损伤类药物联合使用在实体瘤的临床前模型中有着良好的抗肿瘤效果,预示着其良好的临床应用前景。ATR在多种重要的细胞活动中扮演重要的角色,探寻ATR抑制剂在联合用药中的具体作用机制有助于提升人们对此类药物的认识,并为合理设计用于治疗AML的药物联合方案提供重要的理论依据。在本研究中,我们用AML细胞株和AML患者临床样本细胞探究了ATR抑制剂AZ20单独或与阿糖胞苷联合使用抗AML的效果。结果表明,AZ20能够诱导细胞凋亡,部分解除G2/M细胞周期阻滞,引起DNA复制压力和DNA损伤,并导致非CDK1依赖的核苷酸还原酶(Ribonucleotide reductase,RR)M1亚基(RRM1)与M2亚基(RRM2)蛋白表达水平的下调。同时,AZ20能够协同增强阿糖胞苷诱导的AML细胞凋亡,解除阿糖胞苷诱导的S与G2/M细胞周期阻滞,增强阿糖胞苷诱导的DNA复制压力与DNA损伤。同时我们用另一种ATR的选择性抑制剂AZD6738验证了以上发现。因此,抑制ATR协同增强阿糖胞苷杀伤AML细胞活性的机制主要有两个:一是抑制ATR能够增强阿糖胞苷引起的DNA复制压力和DNA损伤;二是抑制ATR能够解除阿糖胞苷诱导的S和G2/M细胞周期阻滞。我们的研究阐明了ATR抑制剂AZ20和AZD6738与阿糖胞苷协同抗AML的分子机制,为ATR抑制剂与阿糖胞苷在AML治疗中的联合应用提供了重要的理论依据与实验基础。
[Abstract]:Acute myeloid leukemia (acute myeloid leukemiaAML) is the most common acute leukemia. The survival rate of children is about 65%, and that of adult patients is only 25%. Cytarabine has been one of the most effective first-line drugs in the treatment of AML for the past 40 years. Although traditional chemotherapy can lead to complete remission in most patients, the resistance of most AML cells to cytarabine is the main reason for the failure of cytosine arabinoside therapy. Therefore, a new and more effective treatment is urgently needed to combat this malignant disease. One of the main mechanisms leading to cytarabine resistance is that chemotherapy-induced DNA damage activates the damage response relate signaling network. ATR (ataxia telangiectasia and Rad3 relate and ATM (ataxia telangiectasia mutated) are two important proteins in the DDR signaling network. When a cell has a DNA double strand break or a replication fork stops, the cell will repair it and produce a single strand DNA, and ATR will be activated to respond to the production of single strand DNA. ATR can regulate several important biological functions, such as regulating DNA damage repair. ATR is an important regulatory factor to promote the survival of DNA damaged cells. In addition, most cancer cells are more dependent on ATR regulated S and G 2 / M cell cycle test points due to the absence of G1 cell cycle detection points. Therefore, inhibiting ATR can enhance the cytotoxicity of DNA-damaging drugs to AML cells. ATR inhibitors combined with gemcitabine, cisplatin and PARP (poly (ADP-ribose) polymerase) inhibitors have a good anti-tumor effect in the preclinical model of solid tumor. ATR plays an important role in many important cellular activities. Exploring the specific mechanism of ATR inhibitors in combination drugs will help to promote the understanding of such drugs. It also provides an important theoretical basis for the rational design of drug combination regimen for AML. In this study, we used AML cell lines and AML patient clinical sample cells to investigate the effects of ATR inhibitor AZ20 alone or in combination with cytarabine. The results showed that AZ20 could induce apoptosis, partially relieve G _ 2 / M cell cycle arrest, induce DNA replication pressure and DNA damage, and down-regulate the expression of non-CDK1-dependent nucleotide reductase-RR M1 subunit (RRM1) and M2 subunit (RRM2). AZ20 could enhance apoptosis of AML cells induced by cytarabine, relieve cycle arrest of S and G _ 2 / M cells induced by cytosine arabinoside, and enhance DNA replication pressure and DNA damage induced by cytosine arabinoside. At the same time, we verify the above findings with AZD 6738, another selective inhibitor of ATR. Therefore, inhibition of ATR synergistically enhances the cytotoxicity of cytarabine to AML cells through two mechanisms: first, inhibiting ATR can enhance the DNA replication pressure and DNA damage induced by cytarabine; Second, inhibition of ATR can relieve cytarabine induced cell cycle arrest in S and G 2 / M cells. The molecular mechanism of ATR inhibitor AZ20 and AZD6738 combined with cytarabine against AML has been elucidated, which provides an important theoretical and experimental basis for the combined application of ATR inhibitor and cytarabine in the treatment of AML.
【学位授予单位】:吉林大学
【学位级别】:硕士
【学位授予年份】:2017
【分类号】:R733.71
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