The peculiar behavior of dihydroartemisinin in hypoxia: effect on endothelial cells

Artemisinin derivatives are the most effective and safe antimalarial drugs presently available. They also exert antiangiogenic effects on human endothelium and tumours. Moreover, artemisinins are specifically cytotoxic against cancer cells and cause reduction of tumour grow in animal models. Therefore these compounds are promising drugs as adjuvants in anticancer chemotherapy. Hypoxia plays a crucial role both in malaria and tumours. In severe malaria, the cytoadherence of infected erythrocytes to endothelial cells (ECs) generates microvascular occlusion and subsequent tissue hypoxia. In cancer, the growth of the tumour mass causes the failure of oxygen and nutrients supply. However, the role of oxygen deprivation in the mechanism of action of artemisinins has not been studied yet. To this purpose, we assessed the activity of dihydroartemisinin (DHA), the active metabolite of most artemisinins, on microvascular ECs (HMEC-1) both in normoxic or hypoxic (1% oxygen) conditions. The doses of DHA were chosen on the base of their pharmacological use: low doses (1μM or 0.5μM) are those reached in the plasma of artemisinins-treated malaria patients and high doses (50μM or 12.5μM) are similar to those used in literature as antitumor or antiangiogenic. By measuring cell growth (MTT assay), we observed that DHA at high dose was more toxic in normoxia, whereas at low dose was more effective in hypoxia. Only the high dose of DHA induced significant apoptosis (measured as phosphatidylserine exposure and caspase activation) and cell cycle arrest in G2. These effects were more evident in normoxia than hypoxia. The role of oxidative stress was also investigated. Again, only DHA 50μM induced ROS production and lipid peroxidation (measured as TBARS production). Doses higher than 10μM, induced ECs death through oxidative stress. We do not know at present whether the toxicity exerted by low doses of DHA under hypoxia could be exploited to reach pathogens or tumour cells in hypoxic niches. Taken together our results suggest that, depending on the pathologic situation, different doses of artemisinins could achieve different effects. This should be taken into account, for example, in the treatment of pregnant women with artemisinins and in the studies on the possible use of these drugs as antitumor agents. Acknowledgments The financial support of the FP6-IP18834 ANTIMAL and Università di Milano (PUR 2009) is acknowledged.