Danusertib induces autophagy of leukemia cells via Aurora kinase B/ribosomal protein p70S6 kinase/ribosomal protein 15 signaling pathway
10.3760/cma.j.issn.2095-428X.2018.24.012
- VernacularTitle:达努塞替通过极光激酶B/核糖体p7OS6蛋白激酶/核糖体蛋白15信号通路调控白血病细胞自噬
- Author:
Sijia HE
1
,
2
;
Zhixu HE
;
Liping SHU
;
Zhiwei ZHOU
Author Information
1. 610091 成都市妇女儿童中心医院小儿呼吸内科
2. 550001贵阳,贵州医科大学附属医院儿科
- Keywords:
Leukemia;
Danusertib;
Autophagy;
Ribosomal protein 15
- From:
Chinese Journal of Applied Clinical Pediatrics
2018;33(24):1890-1894
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effects of Danusertib on the changes in Aurora kinase B (Aurora B)/ribosomal protein p70S6 kinase (p70S6K)/ribosomal protein 15 (RPL15) signaling pathway and autophagy in human leukemia cells and its mechanism.Methods Myeloid leukemia cell lines THP-1 and K562 were selected as the research subjects.The experiment was divided into 2 phases.Phase 1:each cell line was treated with the concentration of Danusertib in 0.1 p mol/L,1.0 p mol/L and 5.0 μ mol/L.In control group,2 mL/L dimethyl sulfoxide (DMSO)was given.All the treated cells were cultured for 24 hours.The viability of each cell line was examined by methyhhiazoletrazolium assay and the autophagy was assessed by flow cytometry.In addition,the protein levels of p70S6K,AuroraB,phosphatidylinositol 3-kinase (PI3K),AKT(phosphorylated protein kinase B),mammalian target of rapamycin (mTOR),microtubule-associated protein (LC3),Beclin1 and RPL15 were determined by using Western blot.Part 2:Aurora B and RPL15 were down-regulated in THP-1 and K562 cells,respectively.DMSO was used to dissolve Danusertib(5.0 μ mol/L).The grouping was designed as following:DMSO group (blank control group),Danusertib treated group,empty plasmid group,small interfering RNA(siRNA) group,empty plasmid + Danusertib-treated group and siRNA + Danusertib treated group.The protein levels of Aurora B,p70S6K,RPL15,Beclinl and LC3 were detected by using Western blot.Results (1) Danusertib decreased the viability of THP-1 and K562 cells and the half maximal inhibitory concentration values were 26.9 pmol/L and 30.2 μmol/L for THP-1 and K562 cells,respectively.(2)The protein levels of p-Aurora B/Aurora B,p-p70S6K/p70S6K,RPL15,p-mTOR/mTOR and p-AKT/AKT decreased compared with control cells after being treated with 0.1 μmol/L,1.0 μ mol/L and 5.0 pmol/L of Danusertib in THP-1 and K562 cells,and the differences were statistically significant (all P ＜ 0.05).(3) In THP-1 cells,compared with the empty plasmid group,the protein levels of p70S6K and RPL15 decreased by 22.1％,61.3％ (F =18.1,P =0.001) and 55.4％,56.1％ (F =19.4,P =0.001) in siRNA group and siRNA + Danusertib-treated group after knockdown of Aurora B.In contrast,the protein levels of LC3 increased by 13.6％ and 17.1％ (F =15.4,P =0.001)compared with the empty plasmid group.In addition,the protein levels of Beclin1 and LC3 increased by 39.5％,92.3％ (F=25.2,P=0.001) and 40.2％,58.3％ (F=23.9,P=0.001) in siRNA group and siRNA + Danusertib treated group,compared with the empty plasmid group after down-regulation of RPL15.In K562 cells,compared with the empty plasmid group,the protein levels of p70S6K and RPL15 decreased by 24.2％,62.7％ (F =20.4,P=0.001) and 57.2％,60.1％ (F =23.9,P =0.001) in siRNA group and siRNA + Danusertib treated group after downregulation of Aurora B.But the protein levels of LC3 increased by 17.9％ and 56.7％ (F =20.9,P =0.001)compared with the empty plasmid group.Moreover,the protein levels of Beclin1 and LC3 were increased by 20.6％,98.4％ (F=22.4,P =0.001) and 41.5％,70.1％ (F=26.2,P =0.001) in siRNA group and siRNA + Danusertib treated group,compared with the empty plasmid group after downregulation of RPL15.Conclusion Danusertib can induce autophagy via inhibition of the PI3K/AKT/mTOR signaling pathway and can negatively regulate Aurora B/p70S6K/RPL15 axis in THP-1 and K562 cells.In addition,RPL15 may be a key target of Aurora B/p70S6K/RPL15signaling pathway in the inhibition of tumor proliferation.
