Arsenic trioxide enhances the cytotoxic effect of thalidomide in a KG-1a human acute mylogenous leukemia cell line

Abstract

Studies have shown that thalidomide exerts modest activity as a single agent in the therapy of acute myeloid leukemia (AML). The present investigation was conducted to test the hypothesis that the cytotoxic effect of thalidomide is enhanced when properly combined with other chemotherapeutic agents. The human AML cell line KG-1a was used in this study. Cells were cultured for 48 h in the presence or absence of thalidomide, arsenic trioxide and a combination of the two substances. Results obtained indicate that thalidomide at concentrations of 1, 2 and 5 mg/l produced a dose-dependent cytotoxic effect and at 5 mg/ml resulted in late apoptosis in 49.39% of the total cell population (as compared to 5.35% in the control cells). When the cells were incubated with arsenic trioxide alone (4 µM), late apoptosis was detected in 16.97% of the total cell population. However, when cells were incubated with a combination of thalidomide (5 mg/l) and arsenic trioxide (4 µM), late apoptosis was noted to be 80.6% in the total cell population. This percentage of late apoptosis was statistically significant from that observed when cells were incubated with thalidomide alone. These findings clearly indicate that arsenic trioxide enhances the cytotoxic effects of thalidomide.

Figure 1

Flow cytometry analysis of KG-1a cells treated with arsenic trioxide (2 μM). The lower left quadrant shows live cells; the lower right, early apoptotic cells; the upper right, late apoptotic cells and the upper left quadrant shows necrotic cells.

Figure 2

Flow cytometry analysis of KG-1a cells treated with arsenic trioxide (2 μM) + ascorbic acid (100 μM). The lower left quadrant shows live cells; the lower right, early apoptotic cells; the upper right, late apoptotic cells and the upper left quadrant shows necrotic cells.

Figure 3

Flow cytometry of KG-1a cells treated with arsenic trioxide (4 μM). The lower left quadrant shows live cells; the lower right, early apoptotic cells; the upper right, late apoptotic cells and the upper left quadrant shows necrotic cells.

Figure 4

Flow cytometry of KG-1a cells treated with arsenic trioxide (4 μM) + ascorbic acid (100 μM). The lower left quadrant shows live cells; the lower right, early apoptotic cells; the upper right, late apoptotic cells and the upper left quadrant shows necrotic cells.

Figure 5

Necrosis of KG-1a cells incubated with various chemotherapeutic agents. Bars represent mean ± SE. Cells were incubated for 48 h with thalidomide (Th) (5 mg/ml), arsenic trioxide (As) (4 μM) and interleukin-2 (IL-2) (200 IU/ml) alone or in combination.

Figure 6

Early apoptosis of KG-1a cells incubated with various chemotherapeutic agents. Bars represent mean ± SE. Cells were incubated for 48 h with thalidomide (Th) (5 mg/ml), arsenic trioxide (As) (4 μM) and interleukin-2 (IL-2) (200 IU/ml) alone or in combination.

Figure 7

Late apoptosis of KG-1a cells incubated with various chemotherapeutic agents. Bars represent mean ± SE. Cells were incubated for 48 h with thalidomide (Th) (5 mg/ml), arsenic trioxide (As) (4 μM) and interleukin-2 (IL-2) (200 IU/ml) alone or in combination. ^*Significantly different from control (p<0.05); ^**significantly different from the control and thalidomide-treated cells (p<0.05).