Archives of Toxicology

, Volume 93, Issue 3, pp 623–634 | Cite as

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

  • Silvia Di GiacomoEmail author
  • Oscar Briz
  • Maria J. Monte
  • Laura Sanchez-Vicente
  • Lorena Abete
  • Elisa Lozano
  • Gabriela Mazzanti
  • Antonella Di Sotto
  • Jose J. G. MarinEmail author
Molecular Toxicology


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.


Chemotherapy Liver cancer Multidrug resistance Sesquiterpenes Tyrosine kinase inhibitor 



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).

Compliance with ethical standards

Conflict of interest

The authors declare that there is no conflict of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted.

Supplementary material

204_2019_2395_MOESM1_ESM.docx (555 kb)
Supplementary material 1 (DOCX 555 KB)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Physiology and Pharmacology “V. Erspamer”Sapienza University of RomeRomeItaly
  2. 2.Laboratory of Experimental Hepatology and Drug Targeting (HEVEFARM), IBSALUniversity of SalamancaSalamancaSpain
  3. 3.Center for the Study of Liver and Gastrointestinal Diseases (CIBERehd)Carlos III National Institute of HealthMadridSpain
  4. 4.Department of Physiology and PharmacologyUniversity of SalamancaSalamancaSpain

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