Department of Clinical Laboratory, Guizhou Provincial People's Hospital, Guiyang, China.
Department of Clinical Laboratory, Guiyang Second People's Hospital, Guiyang, China.
Acta Haematol. 2022;145(5):484-498. doi: 10.1159/000524158. Epub 2022 Mar 21.
Chronic myeloid leukaemia (CML) is a myeloproliferative neoplasm characterized by constitutive activity of the tyrosine kinase BCR-ABL1. Drug resistance remains one of the major challenges in CML therapy. MicroRNA (miR)-199a-3p plays an important role in many tumours but has rarely been investigated in CML. We aimed to analyse the role and mechanism of miR-199a-3p in regulating imatinib resistance in CML.
The expression of miR-199a-3p and mammalian target of rapamycin (mTOR) in the serum of CML patients and CML cells was examined by quantitative real-time polymerase chain reaction. The levels of apoptosis-related proteins were determined using western blot. The relative cell survival rate and cell proliferation were determined using a CCK-8 assay and a bromodeoxyuridine (BrdU) assay, respectively. Cell cycle and apoptosis were analysed using flow cytometry. Moreover, a dual-luciferase reporter assay was performed to verify the correlation between miR-199a-3p and mTOR.
MiR-199a-3p was downregulated in the serum of CML patients and in CML cells, while mTOR was upregulated. Both miR-199a-3p overexpression and mTOR silencing inhibited CML cell proliferation, promoted CML cell apoptosis, and sensitized these cells to imatinib. mTOR silencing reversed the promoting effect of miR-199a-3p inhibition on the proliferation of CML cells and the inhibitory effects on cell apoptosis and sensitivity to imatinib. MiR-199a-3p directly targeted mTOR.
MiR-199a-3p suppressed cell propagation, facilitated apoptosis of CML cells, and sensitized CML cells to imatinib by downregulating mTOR signalling.
慢性髓性白血病(CML)是一种骨髓增生性肿瘤,其特征是酪氨酸激酶 BCR-ABL1 的组成性活性。耐药性仍然是 CML 治疗的主要挑战之一。微小 RNA(miR)-199a-3p 在许多肿瘤中发挥重要作用,但在 CML 中很少被研究。我们旨在分析 miR-199a-3p 在调节 CML 伊马替尼耐药中的作用和机制。
通过定量实时聚合酶链反应检测 CML 患者和 CML 细胞血清中 miR-199a-3p 和哺乳动物雷帕霉素靶蛋白(mTOR)的表达。使用 Western blot 测定凋亡相关蛋白水平。使用 CCK-8 测定和溴脱氧尿苷(BrdU)测定分别测定相对细胞存活率和细胞增殖。通过流式细胞术分析细胞周期和细胞凋亡。此外,进行双荧光素酶报告基因测定以验证 miR-199a-3p 和 mTOR 之间的相关性。
miR-199a-3p 在 CML 患者的血清和 CML 细胞中下调,而 mTOR 上调。miR-199a-3p 过表达和 mTOR 沉默均抑制 CML 细胞增殖,促进 CML 细胞凋亡,并使这些细胞对伊马替尼敏感。mTOR 沉默逆转了 miR-199a-3p 抑制对 CML 细胞增殖的促进作用以及对细胞凋亡和对伊马替尼敏感性的抑制作用。miR-199a-3p 直接靶向 mTOR。
miR-199a-3p 通过下调 mTOR 信号通路抑制 CML 细胞增殖,促进 CML 细胞凋亡,并使 CML 细胞对伊马替尼敏感。