Antitumor effect of betulinic acid on human acute leukemia K562 cells in vitro

Abstract

The effects of betulinic acid (BA), a pentacyclic lupane-type triterpene, on the cell viability, cell cycle and apoptosis in human leukemia K562 cells were investigated. The effects of BA on the growth of K562 cells were studied by MTT assay. Apoptosis was assayed through Annexin V/propidium iodide (PI) double-labeled cytometry. The effects of BA on the cell cycle of K562 cells were studied by a PI method. The expression of Bax and capase-3 was detected by using Western blot. The results showed that BA was cytotoxic to K562 cells with an IC₅₀ of 21.26 μg/mL at 24 h. After treating K562 cells with 10 μg/mL BA for 72 h, the number of cells was reduced by 58%. BA induced apoptosis of K562 cells in a time- and dose-dependent manner. The proportion of cells in G₀/G₁ and G₂/M phases was decreased and that in S phase was increased after K562 cells were treated with BA for 24 h. BA treatment also increased the expression of the pro-apoptotic proteins Bax and caspase-3. It suggested that BA could inhibit the proliferation of K562 cells through the induction of cell cycle arrest and apoptosis. The antitumor effects of BA were related with up-regulation of the expression of Bax and caspase-3 proteins. BA may qualify for the development of new therapies for leukemia.

Keywords

betulinic acid / K562 cells / apoptosis / Bax / caspase-3

References

[1] EsfahaniM.K., MorrisE.L., DutcherJ.P., et al.. Blastic phase of chronic myelogenous leukemia. Curr Treat Options Oncol, 2006, 7(3): 189-199

[2] TheoA., MasebeT., SuzukiY., et al.. Peltophorum africanum, a traditional South African medicinal plant, contains an anti HIV-1 constituent, betulinic acid. Tohoku J Exp Med, 2009, 217(2): 93-99

[3] YaoD., LiH., GouY., et al.. Betulinic acid-mediated inhibitory effect on hepatitis B virus by suppression of manganese superoxide dismutase expression. FEBS J, 2009, 276(9): 2599-2614

[4] ZieglerH.L., FranzykH., SairafianpourM., et al.. Erythrocyte membrane modifying agents and the inhibiton of Plasmodium falciparum growth: structure-activity relationships for betulinic acid analogues. Bioorg Med Chem, 2004, 12(1): 119-127

[5] EiznhamerD.A., XuZ.Q.. Betulinic acid: a promising anticancer candidate. IDrugs, 2004, 7(4): 359-373

[6] KesslerJ.H., MullauerF.B., de RooG.M., et al.. Broad in vitro efficacy of plant-derived betulinic acid against cell lines derived from the most prevalent human cancer types. Cancer Lett, 2007, 251(1): 132-145

[7] JungG.R., KimK.J., ChoiC.H., et al.. Effect of betulinic acid on anticancer drug-resistant colon cancer cells. Basic Clin Pharmacol Toxicol, 2007, 101(4): 277-285

[8] EhrhardtH., FuldaS., FührerM., et al.. Betulinic acid-induced apoptosis in leukemia. Cells, 2004, 18(8): 1406-1012

[9] JungG.R., KimK.J., ChoiC.H., et al.. Effect of betulinic acid on anticancer drug-resistant colon cancer cells. Basic Clin Pharmacol Toxicol, 2007, 101(4): 277-285

[10] FuldaS., ScaffidiC., SusinS.A., et al.. Activation of mitochondria and release of mitochondrial apoptogenic factors by betulinic acid. J Biol Chem, 1998, 273(51): 33942-33948

[11] SelzerE., PimentelE., WacheckV., et al.. Effects of betulinic acid alone and in combination with irradiation in human melanoma cells. J Invest Dermatol, 2000, 114(5): 935-940

[12] SelzerE., ThallingerC., HoellerC., et al.. Betulinic acid-induced Mcl-1 expression in human melanoma-mode of action and functional significance. Mol Med, 2002, 8(12): 877-884

[13] ThurnherD., TurhaniD., PelzmannM., et al.. Betulinic acid: a new cytotoxic compound against malignant head and neck cancer cells. Head Neck, 2003, 25(9): 732-740

[14] FuldaS.. Betulinic acid for cancer treatment and prevention. Int J Mol Sci, 2008, 9(6): 1096-1107

[15] Raghuvar GopalD.V., NarkarA.A., BadrinathY., et al.. Betulinic acid induces apoptosis in human chronic myelogenous leukemia (CML) cell line K-562 without altering the levels of Bcr-Abl. Toxicol Lett, 2005, 155(3): 343-351

[16] PatelM.P., MasoodA., PatelP.S., et al.. Targeting the Bcl-2. Curr Opin Oncol, 2009, 21(6): 516-523

[17] DansonS., DeanE., DiveC., et al.. IAPs as a target for anticancer therapy. Curr Cancer Drug Targets, 2007, 7(8): 785-794

[18] FuldaS.. Therapeutic opportunities for counteracting apoptosis resistance in childhood leukaemia. Br J Haematol, 2009, 145(4): 441-454

[19] HailN.Jr, LotanR.. Cancer chemoprevention and mitochondria: targeting apoptosis in transformed cells via the disruption of mitochondrial bioenergetics/redox state. Mol Nutr Food Res, 2009, 53(1): 49-67

[20] VangestelC., Van de WieleC., MeesG., et al.. Forcing cancer cells to commit suicide. Cancer Biother Radiopharm, 2009, 24(4): 395-407

[21] KeeshanK., CotterT.G., McKennaS.L.. High Bcr-Abl expression prevents the translocation of Bax and Bad to the mitochondrion. Leukemia, 2002, 16(9): 1725-1734

[22] SkorskiT.. BCR/ABL regulates response to DNA damage: the role in resistance to genotoxic treatment and in genomic instability. Oncogene, 2002, 21(56): 8591-604

[23] FuldaS., DebatinK.M.. Betulinic acid induces apoptosis through a direct effect on mitochondria in neuroecto dermal tumors. Med Pediatr Oncol, 2000, 35(6): 616-618

[24] RzeskiW., StepulakA., SzymanskiM., et al.. Betulinic acid decreases expression of bcl-2 and cyclin D1, inhibits proliferation, migration and induces apoptosis in cancer cells. Naunyn Schmiedebergs Arch Pharmacol, 2006, 374(1): 11-20

[25] ZinkelS., GrossA., YangE.. BCL2 family in DNA damage and cell cycle control. Cell Death Differ, 2006, 13(8): 1351-1359