The Mechanism of Artesunate Combined with Cytarabine and/or Daunorubicin on the Apoptosis of MV4-11 MLL-rearranged Acute Myeloid Leukemia Cell Line

Abstract

To investigate the effect and mechanism of artesunate (ARTS) combined with cytarabine(Ara-C) and/or daunorubicin (DNR) on the proliferation and apoptosis of MV4-11 human mixed-lineage leukemia rearranged（MLL-r） acute myeloid leukemia (AML) cell line. CCK-8 assay was used to detect the proliferation effect of individual or in combination of ARTS, DNR, Ara-C on MV4-11 cells. The IC50 of ARTS, DNR and Ara-C was calculated separately. The cell apoptosis and expression of receptors DR4 and DR5 were detected by flow cytometry. Western blot was used to detect the expression of Caspase-3 and Caspase-9 in each groups. The inhibition effect of ARTS, Ara-C and DNR on the proliferation of MV4-11 were all dose-dependently (r=0.99, 0.90 and 0.97, respectively). The IC50 of ARTS, Ara-C and DNR on MV4-11 for 48 hours were 0.31 μg/ml, 1.43 μmol/L and 22.47 nmol/L, respectively. At the dose of ARTS 0.3 μg/ml， Ara-C 1.0 μmol/L and DNR 15 nmol/L, the proliferation rate for 48 hours of the tri-combination treatment was significantly lower than that of the bi-combination treatment, while both were significantly lower than that of the individual treatment (all P＜0.05). In terms of bi-combination treatment, the cells proliferation rate for 48 hours of the ARTS+Ara-C group was significantly lower than that of the ARTS+DNR group, while both were significantly lower than that of the Ara-C+DNR group (all P＜0.05). The cooperativity index (CI) of bi- and tri-combination treatment were all less than 1. After 48 hours of drug action, the cell apoptosis rate of the ARTS+DNR+Ara-C group was significantly higher than that of the Ara-C+DNR group, while both were significantly higher than that of the ARTS+DNR group (all P＜0.05). Meanwhile, the was no statistical difference between the cells apoptotic rate of the ARTS+DNR+Ara-C group and the ARTS+Ara-C group (P>0.05). The expression of DR4 and DR5 also showed no difference between control group and drug group. Compared with the DNR+Ara-C group, the expressions of Caspase-3 were significantly down-regulated in both the ARTS+DNR+Ara-C group and the ARTS+Ara-C group (all P＜0.05). The down-regulation of Caspase-3 expression was the most significantly in the combination group of three drugs, while the Caspase-9 expressions in different groups showed no apparent change. The in vitro study showed that tri-combination of ARTS+Ara-C+DNR and bi-combination of ARTS+Ara-C could inhibit the proliferation and promote apoptosis of MV4-11 cell line. The inhibition effect of these two combinations were significantly superior to that of the traditional Ara-C+DNR treatment. The mechanism underlying this finding may be identified by the down regulation of Caspase-3, while no altered expression was observed of Caspase-9, DR4 and DR5.