Everolimus combined with all-trans retinoid acid reverses drug resistance in acute promyelocytic leukemia NB4-R1 cells
10.3785/j.issn.1008-9292.2015.09.09
- VernacularTitle:依维莫司联合全反式维甲酸逆转急性早幼粒细胞白血病细胞耐药的研究
- Author:
Wei-Chao LIAO
1
;
Ying HE
;
Bin-Sheng WANG
;
He HUANG
Author Information
1. 浙江大学医学院附属第一医院骨髓移植中心
- Keywords:
Leukemia,promyelocytic,acute/pathology;
Everolimus;
Cell differentiation/drug effects;
Suppressor factors,immunologic;
Drug resistance,neoplasm
- From:
Journal of Zhejiang University. Medical sciences
2015;(5):525-531
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To investigate the effect of everolimus ( RAD001 ) combined with all-trans retinoid acid ( ATRA ) on drug resistance of ATRA-resistance acute promyelocytic leukemia ( APL ) cell line NB4-R1 and its molecular mechanism . Methods: APL NB4-R1 cells were treated with different concentrations of RAD 001 (1 nmol/L, 10 nmol/L and 100 nmol/L) with ATRA (1 μmol/L) for 24, 48 and 72 h, respectively .The differentiation of NB4-R1 cells was analyzed by flow cytometry with CD11b staining and nitro blue tetrozolium ( NBT) reduction test.Cell cycle was detected by cell cycle staining kit and apoptosis was detected by flow cytometry with Annexin V/PI staining.Protein expressions of LC-3II, PML-RARα, P-P70S6K and P-4E-BP1 were determined by Western blotting .Results: RAD001 combined with ATRA significantly induced NB 4-R1 cells differentiation , but RAD001 or ATRA alone did not enhance NB 4-R1 differentiation .The co-treatment induced accumulation of cells in G1 phase and decreased the proportion of cells in S phase .The combined treatment had no effect on cell apoptosis .The differentiation rate of NB 4-R1 cells in 100 nmol/L RAD001 , 1 μmol/L ATRA, RAD001 combined with ATRA and control groups was (2.29 ±0.57)%, (17.06 ±2.65)%, (54.47 ±4.91)% and (2.54 ±0.53)%, respectively;the proportion of cells in G 1 phase was ( 35 .20 ±11 .97 )%, ( 33 .54 ± 6.25)%, (53.70 ±8.73)% and (27.40 ±6.01)%, respectively; cells apoptosis rate was ( 2.30 ±0.14 )%, ( 2.25 ±0.21 )%, ( 2.40 ±0.28 )% and (1.95 ±0.07)%, respectively.The combination of RAD001 with ATRA significantly inhibited mTOR signaling downstream proteins P-P70S6K, P-4E-BP1 and enhanced autophagy-related protein LC3-II and Beclin 1. The co-treatment also induced degradation of fusion protein PML-RARα.Conclusion:RAD001 combined with ATRA can induce cell differentiation , inhibit cell cycle , resulting the reverse of drug resistance in NB4-R1 cells, which is associated with increase of autophagy level and degradation of PML-RARα.
