Constituents of Artemisia annua Dietary Supplements Induce ROS Elevation, ERK Activation, and Apoptosis in Treatment‐Resistant Triple Negative Human Breast Cancer Cells

Abstract

In recent years, Artemisia annua L. and especially artemisinin and its semisynthetic derivatives became subject of intense investigations for their potential anticancer activity. Yet, available evidence suggests that artemisinin might not be the most active ingredient of Artemisia annua. For this reason, we have analyzed different Artemisia annua extracts, which did not contain any detectable artemisinin marketed as dietary supplements for their activity against the triple negative human breast cancer cell line MDA‐MB‐231. A bioguided fractionation by semi‐preparative HPLC was conducted and the structures of individual compounds were identified by mass spectrometry and 1H‐ and 13C‐NMR spectroscopy. Extracts as well as the isolated compounds 6,7‐dimethoxy‐coumarin, chrysosplenol D, casticin, arteannuic acid and arteannuin B were further analyzed in vitro and in vivo. The extracts and the individual compounds inhibited the viability of several cancer cell lines. Interestingly, the viability of peripheral blood mononuclear cells was not inhibited at equivalent concentrations. Moreover, treatment either with extract or the individual compounds induced cell cycle arrest in the S‐phase or the G2/M‐phase. Apoptotic cell death was identified by DNA‐fragmentation, activation of caspase 3/7, and annexin V/propidium iodide staining. Moreover, the extract and individual compounds increased the phosphorylation of the extracellular regulated kinase ERK in the presence of sustained high ROS levels, which are required for the ERK‐mediated cell death. Pretreatment with the ERK ‐ inhibitor U‐0126 was able to inhibit chrysosplenol D induced toxicity towards MDA‐MB‐231 cells. Furthermore, antiproliferative efficacies of the extract and individual compounds were confirmed in vivo using 3D breast cancer xenografts grown on the chorioallantoic membrane of fertilized chick eggs, where the Artemisia annua extract and the isolated substances significantly reduced the MDA‐MB‐231 breast cancer tumor volume as well as the expression of the Ki‐67 proliferation marker. Importantly, no systemic toxic effects on the chick embryos could be detected. This study provides new insights into a potential rational use of Artemisia annua‐derived compounds in anticancer therapy and identifies compounds beside artemisinin with potent anticancer activity. Furthermore, these findings provide evidence for the implication of the MEK‐ERK‐pathway in apoptotic cell death induced by chrysosplenol D.Support or Funding InformationThis work was supported by the Academic Center for Complementary and Integrative Medicine (AZKIM), State Ministry of Baden‐Württemberg for Sciences, Research and Arts.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.