Abstract:

Objective To investigate the effect of a novel BMI-1 inhibitor PTC-596 on the proliferation， cell cycle， and apoptosis of human breast cancer MCF-7 cells. Methods MCF-7 cells treated with 0 nmol/L PTC-596 were used as control group； MCF-7 cells treated with 25， and 50 nmol/L PTC-596 as experimental group. The CCK8 assay was used to evaluate the celular proliferation ability； DCFH-DA probe/flow cytometry （FCM） was performed to detect the change of cellular reactive oxygen species （ROS）. PI single-staining/FCM was used to analyze cell cycle； PI/FITC -Annexin V/FCM to analyze the proportion of apoptotic cells. JC-1/FCM was performed to detect the change of mitochondrial membrane potential（MMP）； western blot to analyze the expression of BMI-1， cell cycle related proteion（CyclinD1， CyclinB1， and P21） and apoptosis related proteion （Bax， Bcl-2， and c-PARP）. Results PTC-596 could significantly inhibit MCF-7 cell proliferation， with the IC₅₀ value of 49.33 ± 7.02 nmol/L（24 h）. BMI-1 expression was significantly decreased in the low- and high- drug concentration experimental group （P < 0.01）. The relative expression of CyclinB1 and P21 in the experimental groups were increased （P < 0.01）， while the expression of CyclinD1 was decreased （P < 0.01）， and cell mitosis was inhibited （P < 0.01）， resulting in G₂/M cycle arrest （P < 0.01）. At the same time， the accumulation of reactive oxygen species was increased （P < 0.01）； mitochondrial membrane potential was decreased （P < 0.01）. Bax expression was up-regulated （P < 0.01）， while Bcl-2 expression was down-regulated （P < 0.01）； c-PARP was increased （P < 0.01）， and the proportion of apoptotic cells was significantly increased from 2.04% to 10.56% and 26.74% （P < 0.01）. Conclusion PTC-596 can effectively inhibit the proliferation of human breast cancer MCF-7cells， and its mechanism may be closely related to the inhibition of BMI-1， the induction of cell cycle arrest， and the endogenous mitochondrial apoptosis.

Key words: B-cell-specificmoloney murine leukemia virus insection site 1, PTC-596S, human breast cancer MCF-7cells, cell proliferation, cell cycle, apoptosis