Abstract
Aim:
To investigate the effects of BIIB021, an inhibitor of heat shock protein 90 (Hsp90) alone or in combination with triptolide (TPL) on T-cell acute lymphoblastic leukemia (T-ALL) and the mechanisms of action.
Methods:
Human T-ALL cells line Molt-4 was examined. The cell viability was measured using MTT assay. Apoptotic cells were studied with Hoechst 33258 staining. Cell apoptosis and cell cycle were analyzed using flow cytometry with Annexin V/PI staining and PI staining, respectively. The levels of multiple proteins, including Akt, p65, CDK4/6, p18, Bcl-2 family proteins, MDM2, and p53, were examined with Western blotting. The level of MDM2 mRNA was determined using RT-PCR.
Results:
Treatment of Molt-4 cells with BIIB021 (50–800 nmol/L) inhibited the cell growth in a dose-dependent manner (the IC50 value was 384.6 and 301.8 nmol/L, respectively, at 48 and 72 h). BIIB021 dose-dependently induced G0/G1 phase arrest, followed by apoptosis of Molt-4 cells. Furthermore, BIIB021 increased the expression of p18, decreased the expression of CDK4/6, and activated the caspase pathway in Molt-4 cells. Moreover, BIIB021 (50–400 nmol/L) dose-dependently decreased the phospho-MDM2 and total MDM2 protein levels, but slightly increased the phospho-p53 and total p53 protein levels, whereas TPL (5–40 nmol/L) dose-dependently enhanced p53 activation without affecting MDM2 levels. Co-treatment with BIIB021 and TPL showed synergic inhibition on Molt-4 cell growth. The co-treatment disrupted p53-MDM2 balance, thus markedly enhanced p53 activation. In addition, the co-treatment increased the expression of Bak and Bim, followed by increased activation of caspase-9.
Conclusion:
The combination of BIIB021 and TPL may provide a novel strategy for treating T-ALL by overcoming multiple mechanisms of apoptosis resistance.
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Acknowledgements
This work was supported by the Doctoral Fund of Ministry of Education of China (No 20120101110010), National Natural Science Foundation of China grants (No 81070419 and No 81200384), Zhejiang Provincial Natural Science Foundation of China (No R2090392), Funds of Science Technology Department of Zhejiang Province (No 2012C13021-2 and No 2012C37103), and Fund of Health Bureau of Zhejiang Province (No 2010SSA006).
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Li, M., Zhang, X., Zhou, Wj. et al. Hsp90 inhibitor BIIB021 enhances triptolide-induced apoptosis of human T-cell acute lymphoblastic leukemia cells in vitro mainly by disrupting p53-MDM2 balance. Acta Pharmacol Sin 34, 1545–1553 (2013). https://doi.org/10.1038/aps.2013.124
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DOI: https://doi.org/10.1038/aps.2013.124