Cytotoxicity and modes of action of four Cameroonian dietary spices ethno-medically used to treat cancers: Echinops giganteus, Xylopia aethiopica, Imperata cylindrica and Piper capense

Abstract

Ethnopharmacological relevance: Echinops giganteus, Imperata cylindrica, Piper capense and Xylopia aethiopica are four medicinal spices used in Cameroon to treat cancers.

Aim of the study: The above plants previously displayed cytotoxicity against leukemia CCRF-CEM and CEM/ADR5000 cell lines as well as human pancreatic MiaPaCa-2 cells. The present study aims at emphasizing the study of the cytotoxicity and the modes of action of the above plants on a panel of ten cancer cell lines including various sensitive and drug-resistant phenotypes. The study has been extended to the isolation of the bioactive constituents from Echinops giganteus.

Materials and methods: The cytotoxicity of the extracts was determined using a resazurin reduction assay, whereas the caspase-Glo assay was used to detect the activation of caspases 3/7, caspase 8 and caspase 9 in cells treated with the four extracts. Flow cytometry was used for cell cycle analysis and detection of apoptotic cells, analysis of mitochondrial membrane potential (MMP) as well as measurement of reactive oxygen species (ROS).

Results: The four tested extracts inhibited the proliferation of all tested cancer cell lines including sensitive and drug-resistant phenotypes. Collateral sensitivity of cancer cells to the extract of Echinops giganteus was generally better than to doxorubicin. The recorded IC50 ranges were 3.29 µg/mL [against human knockout clones HCT116 (p53(-/-)) colon cancer cells] to 14.32 µg/mL (against human liver hepatocellular carcinoma HepG2 cells) for the crude extract from Echinops giganteus, 4.17 µg/mL (against breast cancer cells transduced with control vector MDA-MB231 cells) to 19.45 µg/mL (against MDA-MB-231 BCRP cells) for that of Piper capense, 4.11 µg/mL (against leukemia CCRF-CEM cells) to 30.60 µg/mL (against leukemia HL60AR cells) for Xylopia aethiopica, 3.28 µg/mL [against HCT116 (p53(-/-)) cells] to 33.43 µg/mL (against HepG2 cells) for Imperata cylindica and 0.11 µg/mL (against CCRF-CEM cells) to 132.47 µg/mL (against HL60AR cells) for doxorubicin. The four tested extracts induced apoptosis in CCRF-CEM cells via the alteration loss of MMP whilst that of Piper capense also enhanced the production of ROS.

Conclusion: The studied plants are potential cytotoxic drugs that deserve more detailed exploration in the future, to develop novel anticancer drugs against sensitive and otherwise drug-resistant phenotypes.

Keywords: 2′,7′-Dichlorodihydrofluorescein diacetate; 2′,7′-dichlorodihydrofluorescein; 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide; ATP; Apaf-1; CC; Cameroon; Cell cycle distribution; Cytotoxicity; DCF; DCM; DMSO; E. giganteus; Echinops giganteus; H(2)DCF; H(2)DCFH-DA; H(2)O(2); I. cylindrica; IC(50); Imperata cylindrica; JC-1; MDR; MMP; Mitochondrial membrane potential; P. capense; Piper capense; ROS; Spices; TLC; VCC; VIN; X. aethiopica; Xylopia aethiopica; adenosine triphospahte; apoptotic protease activating factor-1; column chromatography; ddH(2)O; dichlorofluorescein; dichloromethane; dimethylsufoxide; double distilled water; inhibitory concentrations 50%; melting points; mitochondrial membrane potential; mp; multidrug resistant; oxygenated water; reactive oxygen species; thin layer chromatography; vacuum column chromatography; vinblastine.