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Chemosensitization of hepatocellular carcinoma cells to sorafenib by β-caryophyllene oxide-induced inhibition of ABC export pumps

Abstract

Several ATP-binding cassette (ABC) proteins reduce intracellular concentrations of antitumor drugs and hence weaken the response of cancer cells to chemotherapy. Accordingly, the inhibition of these export pumps constitutes a promising strategy to chemosensitize highly chemoresistant tumors, such as hepatocellular carcinoma (HCC). Here, we have investigated the ability of β-caryophyllene oxide (CRYO), a naturally occurring sesquiterpene component of many essential oils, to inhibit, at non-toxic doses, ABC pumps and improve the response of HCC cells to sorafenib. First, we have obtained a clonal subline (Alexander/R) derived from human hepatoma cells with enhanced multidrug resistance (MDR) associated to up-regulation (mRNA and protein) of MRP1 and MRP2. Analysis of fluorescent substrates export (flow cytometry) revealed that CRYO did not affect the efflux of fluorescein (MRP3, MRP4 and MRP5) but inhibited that of rhodamine 123 (MDR1) and calcein (MRP1 and MRP2). This ability was higher for CRYO than for other sesquiterpenes assayed. CRYO also inhibited sorafenib efflux, increased its intracellular accumulation (HPLC–MS/MS) and enhanced its cytotoxic response (MTT). For comparison, the effect of known ABC pumps inhibitors was also determined. They induced strong (diclofenac on MRPs), modest (verapamil on MDR1) or null (fumitremorgin C on BCRP) effect on sorafenib efflux and cytotoxicity. In the mouse xenograft model, the response to sorafenib treatment of subcutaneous tumors generated by mouse hepatoma Hepa 1–6/R cells, with marked MDR phenotype, was significantly enhanced by CRYO co-administration. In conclusion, at non-toxic dose, CRYO is able to chemosensitizating liver cancer cells to sorafenib by favoring its intracellular accumulation.

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Acknowledgements

This study was supported by the CIBERehd (EHD15PI05/2016) and Fondo de Investigaciones Sanitarias, Instituto de Salud Carlos III, Spain (PI16/00598, co-funded by European Regional Development Fund/European Social Fund, “Investing in your future”); Spanish Ministry of Economy, Industry and Competitiveness (SAF2016-75197-R); Junta de Castilla y Leon (SA063P17); Fundación Mutua Madrileña, Spain (Call 2015); Fundación Samuel Solórzano Barruso, Spain (FS/7-2016, FS/8-2017 and FS/13-2017); AECC Scientific Foundation (2017/2020), Spain. S. Di Giacomo and A. Di Sotto were supported by the “Enrico and Enrica Sovena” Foundation (Italy); S. Di Giacomo was also financed by the Italian Society of Pharmacology (SIF).

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Correspondence to Silvia Di Giacomo or Jose J. G. Marin.

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Di Giacomo, S., Briz, O., Monte, M.J. et al. Chemosensitization of hepatocellular carcinoma cells to sorafenib by β-caryophyllene oxide-induced inhibition of ABC export pumps. Arch Toxicol 93, 623–634 (2019). https://doi.org/10.1007/s00204-019-02395-9

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Keywords

  • Chemotherapy
  • Liver cancer
  • Multidrug resistance
  • Sesquiterpenes
  • Tyrosine kinase inhibitor