Despite the advances in the early detection of tumors and in the use of surgery, radiation and chemical therapies for disease management, cancer deaths are projected to more than double worldwide over the next two decades. New approaches to prevention and therapy for cancer include enhancement of Natural Killer Cells, antiangiogenic processes, antioxidant defense systems and direct cytotoxicity against cancer cells without harming normal cells are urgently needed. An assay system is described that addresses the ability of chemically defined natural products, such as Curcumin (derived from Curry), Genistein (derived from Soy), Resveratrol (derived from grape skins) and Artemisinin (derived from the Sweet Wormwood plant) to actually destroy cancer cells in vitro. These compounds are used as food supplements in many parts of the world and are important in the prevention of cancer, heart disease and immunological diseases such as arthritis. The purpose of this research is to develop an accurate, fast assay method to determine cell concentrations of viable cells in culture that are exposed to various compounds developed from natural sources. K562 Erythroleukemic cells were added to 48 well plates in 500ul portions [10(5) cells/ml], cultured 48 hr. and exposed to the various compounds for up to 72 hr. All compounds were dissolved in DMSO and diluted sufficiently to prevent solvent effects. A unique cell viability stain, which allowed the rapid staining of dead cells by membrane penetration using propidium iodide, was used to measure the cell viability of the surviving cells by gating on forward light scatter (size) and fluorescence flow cytometry. The control cultures were greater than 95% viable during and at the end of the experiments and were used as a baseline to measure the degree of killing. All samples were run as 3-6 replicates and SD determined. Both the Phytosome Curcumin derivative and pure Curcumin showed complete killing of as many as 10(6) cells/ml (zero detected cells) at a concentration of 25 µM as early as 20 hr. of culture. Genistein showed 94% killing at 65 hrs. but only 16% killing at 20 hrs. at a concentration of 100 µM. Artemisinin revealed 71% killing at 65 hrs. but only 2% killing at 20 hrs. for 100 µM. The delayed killing may reflect apoptotic events. Transferrin was used up to 800µg/ml without affecting K562 cell viability to potentially load the cells with iron. No enhancement in killing at 72 hr. was seen with Artemisinin, which requires iron for its cytotoxic peroxide activity, suggesting saturation with iron in the K562 cells had already occurred. Resveratrol at 50 µM completely destroyed the cancer cells at 67 hrs. N-acetylcysteine (NAC) showed minimal cytotoxicity of 13% for 200 µM at 65 hrs. This assay system will allow for the study of synergism of the compounds in a rapid, precise method to assess cancer cytotoxicity.
Citation Format: Jerry T. Thornthwaite, Brandon England, Spencer England, Michelle Clarke, Lee Roland, Hare Shah. The anticancer effects of chemically derived natural products. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3209. doi:10.1158/1538-7445.AM2014-3209