Metabolic profiling study of shikonin's inhibitory effect on cancer cell lines

Shikonin, alkannin and their esters deriving mainly from the roots of the Boraginaceae plant family such as Alkanna tinctoria, Lithospermum erythrorhizon etc, have been shown to exhibit among others significant inhibitory effect on cancer cells and tumors [1 – 3]. Multiple mechanisms of action involved have been elucidated [1 – 4]. Metabolic profiling in cells is a promising tool for studying the underlying mechanisms. By this approach changes in the context of the global network of metabolic pathways in a cell can be revealed, providing information on cell's functions, phenotype and response to interventions.

In the current study the inhibitory effect of shikonin was examined in vitro, in human hepatocellular cell line (Huh-7). It was shown that shikonin exhibited an antiproliferative effect in a dose and time dependant manner with an IC₅₀ of 5µM. Shikonin-treated cell cultures exhibited gradual accumulation of non-viable cells that started 24h after treatment and reached the maximum value of over 50% at 96h. In an attempt to investigate the mechanisms by which shikonin acts against Huh-7 proliferation, a snapshot of the metabolic phenotype of the cells upon treatment (48h) with IC₅₀ was acquired by a LC-MS/MS targeted metabolic profiling method capable to determinate 105 primary metabolites. Our results show that the inhibitory effect of shikonin reflects perturbations both in the cell metabolome and in the extracellular medium. Differences of uptake of substrates or excretion of metabolites is been detected in the cell medium e.g. for aminoacids (arginine and proline) while more significant effects are revealed in the intracellular metabolites in response to shikonin intervention.

The study demonstrates the potential of metabolomics to improve the knowledge on the response of hepatocarcinoma cell line to shikonin exposure and could be further used as a roadmap for shikonin, alkannin and other cytotoxic compounds on several cancer cells and in vivo experiments.

Acknowledgements: We acknowledge A. Kyriazou for technical assistance on cell culturing.

Keywords : Metabolic profiling, in vitro, shikonin, cytotoxicity, alkannin

References:

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