Andrographolide pre-treatment enhances the anti-tumor activity of topotecan on acute myeloid leukemia cells in vitro

Abstract

Background: Topotecan (TP) is an anticancer drug currently being used in the treatment of several cancer types including leukemia. However, due to its dose-limiting cytotoxicity, combination treatments have been investigated to achieve better outcome with less side effects. Andrographolide (AG) is a plant-derived compound that was reported to induce apoptosis and autophagy in acute myeloid leukemia cell lines. Objective: The aim of this study was to investigate the possible synergism between topotecan and andrographolide in acute myeloid cells in vitro, to enhance the effect of low doses of topotecan in controlling cancer progression. Materials and Methods: AML cells (U937) were treated with various concentrations of TP and AG, alone and in combination. Cell survival was assessed at different time intervals using cell-proliferation assays. Cell cycle analysis was performed by propidium iodide (PI) staining followed by flow cytometry to assess cell DNA content. Apoptosis induction was measured using Annexin V/PI staining. Results: When applied separately, topotecan and andrographolide both exhibited anti-proliferative effects in a dose-dependent manner. Cotreating the cells with both topotecan and andrographolide simultaneously at various concentrations did not result in any significant changes. However, pretreating the cells with andrographolide for 24 hours prior to topotecan addition, led to a significant decrease in cell proliferation (39%) as compared to treatment with the compounds separately (67%). Topotecan treatment caused a specific cell-cycle arrest at the S phase in addition to an increase in the percentage of cells in the preG phase. These alterations in cell cycle progression were further enhanced upon andrographolide pre-treatment: the percentage of cells in the S-phase reached 40.4%, compared to 19.3% upon treatment with topotecan alone. The pro-apoptotic effect upon combination of topotecan with andrographolide was further observed using annexin V/PI staining; even though andrographolide alone did not alter cell survival, however, when applied to the cells prior to topotecan treatment, the number of apoptotic cells increased from 16% to 57%. Conclusion: Pretreating acute myeloid leukemia cell lines with non-cytotoxic doses of andrographolide promotes the anti-tumor effect of topotecan in vitro.