Acute myeloid leukemia (AML) is a heterogeneous disease that frequently relapses after standard chemotherapy. Therefore, there is a need for the development of novel chemotherapeutic agents that could treat AML effectively. Radotinib, an oral BCR-ABL tyrosine kinase inhibitor, was developed as a drug for the treatment of chronic myeloid leukemia. Previously, we reported that radotinib exerts increased cytotoxic effects towards AML cells. However, little is known about the effects of combining radotinib with Ara-C, a conventional chemotherapeutic agent for AML, with respect to cell death in AML cells. Therefore, we investigated combination effects of radotinib and Ara-C on AML in this study. Synergistic anti-cancer effects of radotinib and Ara-C in AML cells including HL60, HEL92.1.7, THP-1 and bone marrow cells from AML patients have been examined. Diverse cell biological assays such as cell viability assay, Annexin V-positive cells, caspase-3 activity, cell cycle distribution, and related signaling pathway have been performed. The combination of radotinib and Ara-C was found to induce AML cell apoptosis, which involved the mitochondrial pathway. In brief, combined radotinib and Ara-C significantly induced Annexin V-positive cells, cytosolic cytochrome C, and the pro-apoptotic protein Bax in AML cells including HL60, HEL92.1.7, and THP-1. In addition, mitochondrial membrane potential and Bcl-xl protein were markedly decreased by radotinib and Ara-C. Moreover, this combination induced caspase-3 activity. Cleaved caspase-3, 7, and 9 levels were also increased by combined radotinib and Ara-C. Additionally, radotinib and Ara-C co-treatment induced G0/G1 arrest via the induction of CDKIs such as p21 and p27 and the inhibition of CDK2 and cyclin E. Thus, radotinib/Ara-C induces mitochondrial-dependent apoptosis and G0/G1 arrest via the regulation of the CDKI–CDK–cyclin cascade in AML cells. In addition, our results showed that combined treatment with radotinib and Ara-C inhibits AML cell growth, including tumor volumes and weights in vivo. Also, the combination of radotinib and Ara-C can sensitize cells to chemotherapeutic agents such as daunorubicin or idarubicin in AML cells. Therefore, our results can be concluded that radotinib in combination with Ara-C possesses a strong anti-AML activity.

中文翻译：

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Radotinib增强阿糖胞苷（Ara-C）诱导的急性髓性白血病细胞死亡

急性髓细胞性白血病（AML）是一种异质性疾病，在标准化学疗法后经常复发。因此，需要开发可以有效治疗AML的新型化学治疗剂。Radotinib是一种口服BCR-ABL酪氨酸激酶抑制剂，被开发为用于治疗慢性粒细胞白血病的药物。以前，我们报道了拉多替尼对AML细胞具有增强的细胞毒性作用。但是，关于将拉多替尼与传统的AML化疗药物Ara-C联合使用对AML细胞死亡的影响知之甚少。因此，在这项研究中，我们研究了雷多替尼和Ara-C对AML的联合作用。已经检测了雷多替尼和Ara-C在AML细胞（包括来自AML患者的HL60，HEL92.1.7，THP-1和骨髓细胞）中的协同抗癌作用。已经进行了多种细胞生物学测定，例如细胞生存力测定，膜联蛋白V阳性细胞，caspase-3活性，细胞周期分布以及相关的信号通路。发现radotinib和Ara-C的组合可诱导AML细胞凋亡，涉及线粒体途径。简而言之，联合使用的拉多替尼和Ara-C可在包括HL60，HEL92.1.7和THP-1在内的AML细胞中显着诱导膜联蛋白V阳性细胞，胞质细胞色素C和促凋亡蛋白Bax。此外，雷多替尼和Ara-C显着降低了线粒体膜电位和Bcl-xl蛋白。而且，这种组合诱导了胱天蛋白酶3活性。radotinib和Ara-C的联合使用也可以使Caspase-3、7和9的水平降低。另外，radotinib和Ara-C共同处理通过诱导CDKI（例如p21和p27）以及抑制CDK2和cyclin E诱导G0 / G1阻滞。因此，radotinib / Ara-C诱导线粒体依赖性细胞凋亡和G0 / G1阻滞AML细胞中CDKI–CDK–细胞周期蛋白级联反应的调控。此外，我们的结果表明，雷多替尼和Ara-C的联合治疗可抑制AML细胞的生长，包括体内的肿瘤体积和重量。同样，雷多替尼和Ara-C的组合可以使细胞对AML细胞中的化疗药物（如柔红霉素或伊达比星）敏感。因此，我们的结果可以得出结论，拉多替尼联合Ara-C具有很强的抗AML活性。radotinib / Ara-C通过调节AML细胞中CDKI–CDK–cyclin级联反应诱导线粒体依赖性凋亡和G0 / G1阻滞。此外，我们的结果表明，雷多替尼和Ara-C联合治疗可抑制AML细胞的生长，包括体内的肿瘤体积和重量。同样，雷多替尼和Ara-C的组合可以使细胞对AML细胞中的化疗药物（如柔红霉素或伊达比星）敏感。因此，我们的结果可以得出结论，拉多替尼联合Ara-C具有很强的抗AML活性。radotinib / Ara-C通过调节AML细胞中CDKI–CDK–cyclin级联反应诱导线粒体依赖性凋亡和G0 / G1阻滞。此外，我们的结果表明，雷多替尼和Ara-C的联合治疗可抑制AML细胞的生长，包括体内的肿瘤体积和重量。同样，雷多替尼和Ara-C的组合可以使细胞对AML细胞中的化疗药物（如柔红霉素或伊达比星）敏感。因此，我们的结果可以得出结论，拉多替尼联合Ara-C具有很强的抗AML活性。雷多替尼和Ara-C的组合可以使细胞对AML细胞中的化疗药物（如柔红霉素或伊达比星）敏感。因此，我们的结果可以得出结论，拉多替尼联合Ara-C具有很强的抗AML活性。雷多替尼和Ara-C的组合可以使细胞对AML细胞中的化疗药物（如柔红霉素或伊达比星）敏感。因此，我们的结果可以得出结论，拉多替尼联合Ara-C具有很强的抗AML活性。