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Effects of Hypericum perforatum hydroalcoholic extract, hypericin, and hyperforin on cytotoxicity and CYP3A4 mRNA expression in hepatic cell lines: a comparative study

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Abstract

Hypericum perforatum L. is one of the best-studied and consumed worldwide medicinal plants and several reports described various biological activities, including sedative, anti-inflammatory, anticancer, and antidepressant. H. perforatum, hypericin, and hyperforin can regulate the expression of CYP3A4 and, to our knowledge, studies are focused on in vivo and clinical evaluations and, there is no an in vitro study comparing CYP3A4 expression between hepatic cells. This study evaluated the effects of H. perforatum hydroalcoholic extract, hypericin, and hyperforin in cellular viability and CYP3A4 expression in HepG2, WRL-68, and HepaRG cells.

It was found that, in general, the cytotoxicity promoted by H. perforatum is similar to that promoted by hyperforin alone, time- and dose dependent and the most sensitive cell line is WRL-68 followed by HepG2 and HepaRG cells. Hypericin did not significantly compromise the cellular viability of cells. Regarding their effects in CYP3A4 expression, hyperforin strongly induced CYP3A4 mRNA expression in WRL-68 and HepG2, while in HepaRG it remains unaltered. Hypericin increased CYP3A4 levels in WRL-68 and HepaRG and diminished it in HepG2 cells. H. perforatum induced or decreased CYP3A4 mRNA expression cell- and time-dependent. Computational docking studies showed a lower binding energy between hypericin or hyperforin and its four major metabolites and the catalytic center of the CYP3A4 protein in comparison with ketoconazole.

Overall, depending on the cell line, concentration and time of incubation, H. perforatum, hypericin and hyperforin, can exert distinct effects both in cytotoxicity and in CYP3A4 mRNA expression. The results highlighted that at least the continuous and high-dose usage of H. perforatum should be carefully done and monitored, especially in combination with CYP3A4 metabolized drugs.

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Abbreviations

CYP:

cytochrome P450

CYP3A4:

CYP monooxygenase 3A4

CV :

cellular viability relative to controls

MTT assay:

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay

References

  • Barnes J, Anderson LA, Phillipson JD (2001) St John's wort (Hypericum perforatum L.): a review of its chemistry, pharmacology and clinical properties. J Pharm Pharmacol 53(5):583–600

    Article  CAS  PubMed  Google Scholar 

  • Bilia AR, Gallori S, Vincieri FF (2002) St. John's wort and depression: efficacy, safety and tolerability-an update. Life Sci 70(26):3077–3096

    Article  CAS  PubMed  Google Scholar 

  • Breidenbach T, Hoffmann MW, Becker T, Schlitt H, Klempnauer J (2000) Drug interaction of St John's wort with cyclosporin. Lancet 355(9218):1912

    Article  CAS  PubMed  Google Scholar 

  • Cantoni L, Rozio M, Mangolini A, Hauri L, Caccia S (2003) Hyperforin contributes to the hepatic CYP3A-inducing effect of Hypericum perforatum extract in the mouse. Toxicol Sci 75(1):25–30

    Article  CAS  PubMed  Google Scholar 

  • Chu V, Einolf HJ, Evers R, Kumar G, Moore D, Ripp S, Silva J, Sinha V, Sinz M, Skerjanec A (2009) In vitro and in vivo induction of cytochrome p450: a survey of the current practices and recommendations: a pharmaceutical research and manufacturers of america perspective. Drug Metab Dispos 37(7):1339–1354

    Article  CAS  PubMed  Google Scholar 

  • Cui Y, Ang CY, Beger RD, Heinze TM, Hu L, Leakey J (2004) In vitro metabolism of hyperforin in rat liver microsomal systems. Drug Metab Dispos 32(1):28–34

    Article  CAS  PubMed  Google Scholar 

  • Di Carlo G, Borrelli F, Ernst E, Izzo AA (2001) St John's wort: Prozac from the plant kingdom. Trends Pharmacol Sci 22(6):292–297

    Article  PubMed  Google Scholar 

  • Ding X, Kaminsky LS (2003) Human extrahepatic cytochromes P450: function in xenobiotic metabolism and tissue-selective chemical toxicity in the respiratory and gastrointestinal tracts. Annu Rev Pharmacol Toxicol 43:149–173

    Article  CAS  PubMed  Google Scholar 

  • Ernst E, Rand JI, Barnes J, Stevinson C (1998) Adverse effects profile of the herbal antidepressant St. John's wort (Hypericum perforatum L.). Eur J Clin Pharmacol 54(8):589–594

    Article  CAS  PubMed  Google Scholar 

  • Fasinu P, Bouic PJ, Rosenkranz B (2012) Liver-based in vitro technologies for drug biotransformation studies - a review. Curr Drug Metab 13(2):215–224

    Article  CAS  PubMed  Google Scholar 

  • Fox FE, Niu Z, Tobia A, Rook AH (1998) Photoactivated hypericin is an anti-proliferative agent that induces a high rate of apoptotic death of normal, transformed, and malignant T lymphocytes: implications for the treatment of cutaneous lymphoproliferative and inflammatory disorders. J Invest Dermatol 111(2):327–332

    Article  CAS  PubMed  Google Scholar 

  • Gerets HH, Tilmant K, Gerin B, Chanteux H, Depelchin BO, Dhalluin S, Atienzar FA (2012) Characterization of primary human hepatocytes, HepG2 cells, and HepaRG cells at the mRNA level and CYP activity in response to inducers and their predictivity for the detection of human hepatotoxins. Cell Biol Toxicol 28(2):69–87

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Greeson JM, Sanford B, Monti DA (2001) St. John's wort (Hypericum perforatum): a review of the current pharmacological, toxicological, and clinical literature. Psychopharmacology (Berl) 153(4):402–414

    Article  CAS  Google Scholar 

  • Guillouzo A, Corlu A, Aninat C, Glaise D, Morel F, Guguen-Guillouzo C (2007) The human hepatoma HepaRG cells: a highly differentiated model for studies of liver metabolism and toxicity of xenobiotics. Chem Biol Interact 168(1):66–73

    Article  CAS  PubMed  Google Scholar 

  • Gutierrez-Ruiz MC, Bucio L, Souza V, Gomez JJ, Campos C, Carabez A (1994) Expression of some hepatocyte-like functional properties of WRL-68 cells in culture. In Vitro Cell Dev Biol Anim 30A(6):366–371

    Article  CAS  PubMed  Google Scholar 

  • Hall SD, Wang Z, Huang SM, Hamman MA, Vasavada N, Adigun AQ, Hilligoss JK, Miller M, Gorski JC (2003) The interaction between St John's wort and an oral contraceptive. Clin Pharmacol Ther 74(6):525–535

    Article  CAS  PubMed  Google Scholar 

  • Hellum BH, Hu Z, Nilsen OG (2007) The induction of CYP1A2, CYP2D6 and CYP3A4 by six trade herbal products in cultured primary human hepatocytes. Basic Clin Pharmacol Toxicol 100(1):23–30

    Article  CAS  PubMed  Google Scholar 

  • Hellum BH, Nilsen OG (2008) In vitro inhibition of CYP3A4 metabolism and P-glycoprotein-mediated transport by trade herbal products. Basic Clin Pharmacol Toxicol 102(5):466–475

    Article  CAS  PubMed  Google Scholar 

  • Hokkanen J, Tolonen A, Mattila S, Turpeinen M (2011) Metabolism of hyperforin, the active constituent of St. John's wort, in human liver microsomes. Eur J Pharm Sci 42(3):273–284

    Article  CAS  PubMed  Google Scholar 

  • Hu ZP, Yang XX, Chen X, Cao J, Chan E, Duan W, Huang M, Yu XQ, Wen JY, Zhou SF (2007) A mechanistic study on altered pharmacokinetics of irinotecan by St. John's wort. Curr Drug Metab 8(2):157–171

    Article  CAS  PubMed  Google Scholar 

  • Ioannides C (1996) Cytochromes: metabolic and toxicological aspects. Taylor & Francis.

  • Ioannides C (2006) Topics in Xenobiochemistry - Pharmacokinetic interactions between herbal remedies and medicinal drugs. Xenebiotica 32(6):451–478

    Article  Google Scholar 

  • Jakovljevic V, Popovic M, Mimica-Dukic N, Sabo A, Gvozdenovic L (2000) Pharmacodynamic study of Hypericum perforatum L. Phytomedicine 7(6):449–453

    Article  CAS  PubMed  Google Scholar 

  • Jarema M (2008) Herbal drug treatment.. Neuro Endocrinol Lett 29(Suppl 1):93–104

    PubMed  Google Scholar 

  • Jennen DG, Magkoufopoulou C, Ketelslegers HB, van Herwijnen MH, Kleinjans JC, van Delft JH (2010) Comparison of HepG2 and HepaRG by whole-genome gene expression analysis for the purpose of chemical hazard identification. Toxicol Sci 115(1):66–79

    Article  CAS  PubMed  Google Scholar 

  • Kanebratt KP, Andersson TB (2008) Evaluation of HepaRG cells as an in vitro model for human drug metabolism studies. Drug Metab Dispos 36(7):1444–1452

    Article  CAS  PubMed  Google Scholar 

  • Karioti A, Bilia AR (2010) Hypericins as potential leads for new therapeutics. Int J Mol Sci 11(2):562–594

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Klemow KM, Bartlow A, Crawford J, Kocher N, Shah J and Ritsick M (2011) Medical attributes of St. John's Wort (hypericum perforatum). In: Benzie IFF & Wachtel-Galor S (eds) Herbal medicine: biomolecular and clinical aspects edited, 2nd edn. H Boca Raton (FL).

  • Komoroski BJ, Zhang S, Cai H, Hutzler JM, Frye R, Tracy TS, Strom SC, Lehmann T, Ang CY, Cui YY, Venkataramanan R (2004) Induction and inhibition of cytochromes P450 by the St. John's wort constituent hyperforin in human hepatocyte cultures. Drug Metab Dispos 32(5):512–518

    Article  CAS  PubMed  Google Scholar 

  • Lehmann JM, McKee DD, Watson MA, Willson TM, Moore JT, Kliewer SA (1998) The human orphan nuclear receptor PXR is activated by compounds that regulate CYP3A4 gene expression and cause drug interactions. J Clin Invest 102(5):1016–1023

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Linde K (2009) St. John's wort - an overview. Forsch Komplementmed 16(3):146–155. (2006)

    Article  PubMed  Google Scholar 

  • Madabushi R, Frank B, Drewelow B, Derendorf H, Butterweck V (2006) Hyperforin in St. John's wort drug interactions. Eur J Clin Pharmacol 62(3):225–233

    Article  CAS  PubMed  Google Scholar 

  • Mai I, Stormer E, Bauer S, Kruger H, Budde K, Roots I (2003) Impact of St John's wort treatment on the pharmacokinetics of tacrolimus and mycophenolic acid in renal transplant patients. Nephrol Dial Transplant 18(4):819–822

    Article  CAS  PubMed  Google Scholar 

  • Mannu J, Jenardhanan P, Mathur PP (2011) A computational study of CYP3A4 mediated drug interaction profiles for anti-HIV drugs. J Mol Model 17(8):1847–1854

    Article  CAS  PubMed  Google Scholar 

  • Markowitz JS, Donovan JL, DeVane CL, Taylor RM, Ruan Y, Wang JS, Chavin KD (2003) Effect of St John's wort on drug metabolism by induction of cytochrome P450 3A4 enzyme. JAMA 290(11):1500–1504

    Article  CAS  PubMed  Google Scholar 

  • Martinho A, Goncalves I, Costa M, Santos CR (2012) Stress and glucocorticoids increase transthyretin expression in rat choroid plexus via mineralocorticoid and glucocorticoid receptors. J Mol Neurosci 48(1):1–13

    Article  CAS  PubMed  Google Scholar 

  • Martinho A, Goncalves I, Santos CR (2013) Glucocorticoids regulate metallothionein-1/2 expression in rat choroid plexus: effects on apoptosis. Mol Cell Biochem 376(1–2):41–51

    Article  CAS  PubMed  Google Scholar 

  • Mathijssen RH, Verweij J, de Bruijn P, Loos WJ, Sparreboom A (2002) Effects of St. John's wort on irinotecan metabolism. J Natl Cancer Inst 94(16):1247–1249

    Article  CAS  PubMed  Google Scholar 

  • Morris GM, Goodsell DS, Halliday RS, Huey R, Hart WE, Belew RK, Olson AJ (1998) Automated docking using a Lamarckian genetic algorithm and an empirical binding free energy function. J Comput Chem 19(14):1639–1662

    Article  CAS  Google Scholar 

  • Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS, Olson AJ (2009) AutoDock4 and AutoDockTools4: automated docking with selective receptor flexibility. J Comput Chem 30(16):2785–2791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mueller SC, Majcher-Peszynska J, Uehleke B, Klammt S, Mundkowski RG, Miekisch W, Sievers H, Bauer S, Frank B, Kundt G, Drewelow B (2006) The extent of induction of CYP3A by St. John's wort varies among products and is linked to hyperforin dose. Eur J Clin Pharmacol 62(1):29–36

    Article  CAS  PubMed  Google Scholar 

  • Muller WE (2003) Current St John's wort research from mode of action to clinical efficacy. Pharmacol Res 47(2):101–109

    Article  CAS  PubMed  Google Scholar 

  • Nahrstedt A, Butterweck V (1997) Biologically active and other chemical constituents of the herb of Hypericum perforatum L. Pharmacopsychiatry 30(Suppl 2):129–134

    Article  CAS  PubMed  Google Scholar 

  • Nebert DW, Gonzalez FJ (1987) P450 genes: structure, evolution, and regulation. Annu Rev Biochem 56:945–993

    Article  CAS  PubMed  Google Scholar 

  • Novotna A, Krasulova K, Bartonkova I, Korhonova M, Bachleda P, Anzenbacher P, Dvorak Z (2014) Dual effects of ketoconazole cis-enantiomers on CYP3A4 in human hepatocytes and HepG2 Cells. PLoS One 9(10):e111286

    Article  PubMed  PubMed Central  Google Scholar 

  • Obach RS (2000) Inhibition of human cytochrome P450 enzymes by constituents of St. John's Wort, an herbal preparation used in the treatment of depression. J Pharmacol Exp Ther 294(1):88–95

    CAS  PubMed  Google Scholar 

  • Ohno Y, Hisaka A, Suzuki H (2007) General framework for the quantitative prediction of CYP3A4-mediated oral drug interactions based on the AUC increase by coadministration of standard drugs. Clin Pharmacokinet 46(8):681–696

    Article  CAS  PubMed  Google Scholar 

  • Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC, Ferrin TE (2004) UCSF Chimera--a visualization system for exploratory research and analysis. J Comput Chem 25(13):1605–1612

    Article  CAS  PubMed  Google Scholar 

  • Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29(9):e45

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Polli JE (2008) In vitro studies are sometimes better than conventional human pharmacokinetic in vivo studies in assessing bioequivalence of immediate-release solid oral dosage forms. AAPS J 10(2):289–299

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Portoles A, Terleira A, Calvo A, Martinez I, Resplandy G (2006) Effects of Hypericum perforatum on ivabradine pharmacokinetics in healthy volunteers: an open-label, pharmacokinetic interaction clinical trial. J Clin Pharmacol 46(10):1188–1194

    Article  CAS  PubMed  Google Scholar 

  • Rendic S, Di Carlo FJ (1997) Human cytochrome P450 enzymes: a status report summarizing their reactions, substrates, inducers, and inhibitors. Drug Metab Rev 29(1–2):413–580

    Article  CAS  PubMed  Google Scholar 

  • Rengelshausen J, Banfield M, Riedel KD, Burhenne J, Weiss J, Thomsen T, Walter-Sack I, Haefeli WE, Mikus G (2005) Opposite effects of short-term and long-term St John's wort intake on voriconazole pharmacokinetics. Clin Pharmacol Ther 78(1):25–33

    Article  CAS  PubMed  Google Scholar 

  • Roby CA, Anderson GD, Kantor E, Dryer DA, Burstein AH (2000) St John's Wort: effect on CYP3A4 activity. Clin Pharmacol Ther 67(5):451–457

    Article  CAS  PubMed  Google Scholar 

  • Russo E, Scicchitano F, Whalley BJ, Mazzitello C, Ciriaco M, Esposito S, Patane M, Upton R, Pugliese M, Chimirri S, Mammi M, Palleria C, De Sarro G (2014) Hypericum perforatum: pharmacokinetic, mechanism of action, tolerability, and clinical drug-drug interactions. Phytother Res 28(5):643–655

    Article  CAS  PubMed  Google Scholar 

  • Schulz HU, Schurer M, Bassler D, Weiser D (2006) Investigation of the effect on photosensitivity following multiple oral dosing of two different hypericum extracts in healthy men. Arzneimittelforschung 56(3):212–221

    CAS  PubMed  Google Scholar 

  • Shahrokh K, Cheatham 3rd TE, Yost GS (2012) Conformational dynamics of CYP3A4 demonstrate the important role of Arg212 coupled with the opening of ingress, egress and solvent channels to dehydrogenation of 4-hydroxy-tamoxifen. Biochim Biophys Acta 1820(10):1605–1617

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shelton RC, Keller MB, Gelenberg A, Dunner DL, Hirschfeld R, Thase ME, Russell J, Lydiard RB, Crits-Cristoph P, Gallop R, Todd L, Hellerstein D, Goodnick P, Keitner G, Stahl SM, Halbreich U (2001) Effectiveness of St John's wort in major depression: a randomized controlled trial. JAMA 285(15):1978–1986

    Article  CAS  PubMed  Google Scholar 

  • Sinz M, Wallace G, Sahi J (2008) Current industrial practices in assessing CYP450 enzyme induction: preclinical and clinical. AAPS J 10(2):391–400

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Smith PF, Bullock JM, Booker BM, Haas CE, Berenson CS, Jusko WJ (2004) Induction of imatinib metabolism by hypericum perforatum. Blood 104(4):1229–1230

    Article  CAS  PubMed  Google Scholar 

  • Tannergren C, Engman H, Knutson L, Hedeland M, Bondesson U, Lennernas H (2004) St John's wort decreases the bioavailability of R- and S-verapamil through induction of the first-pass metabolism. Clin Pharmacol Ther 75(4):298–309

    Article  CAS  PubMed  Google Scholar 

  • Venkatakrishnan K, von Moltke LL, Greenblatt DJ (1999) Nortriptyline E-10-hydroxylation in vitro is mediated by human CYP2D6 (high affinity) and CYP3A4 (low affinity): implications for interactions with enzyme-inducing drugs. J Clin Pharmacol 39(6):567–577

    Article  CAS  PubMed  Google Scholar 

  • Wang XD, Li JL, Lu Y, Chen X, Huang M, Chowbay B, Zhou SF (2007) Rapid and simultaneous determination of nifedipine and dehydronifedipine in human plasma by liquid chromatography-tandem mass spectrometry: application to a clinical herb-drug interaction study. J Chromatogr B Analyt Technol Biomed Life Sci 852(1–2):534–544

    Article  CAS  PubMed  Google Scholar 

  • Watkins PB, Wrighton SA, Schuetz EG, Molowa DT, Guzelian PS (1987) Identification of glucocorticoid-inducible cytochromes P-450 in the intestinal mucosa of rats and man. J Clin Invest 80(4):1029–1036

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xie HG, Kim RB (2005) St John's wort-associated drug interactions: short-term inhibition and long-term induction? Clin Pharmacol Ther 78(1):19–24

    Article  CAS  PubMed  Google Scholar 

  • Zanoli P (2004) Role of hyperforin in the pharmacological activities of St. John's Wort. CNS Drug Rev 10(3):203–218

    Article  CAS  PubMed  Google Scholar 

  • Zhou SF (2008) Drugs behave as substrates, inhibitors and inducers of human cytochrome P450 3A4. Curr Drug Metab 9(4):310–322

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

The authors acknowledge the Programa Operacional Regional do Centro 20072013 “Mais Centro”- Project “Therapeutic drug monitoring on age related diseases” (CENTRO-07-ST24-FEDER-002012), the Program COMPETE and the Portuguese Foundation for Science and Technology (PEst-OE/SAU/UI0709/2014).

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    1. CICS-UBI – Health Sciences Research Centre, University of Beira Interior, Av. Infante D. Henrique, Covilhã, 6200-506, Portugal

      Ana Martinho, Sara M. Silva, Sara Garcia, Ivo Moreno, Luiza Breitenfeld Granadeiro, Gilberto Alves, Ana Paula Duarte, Fernanda Domingues, Samuel Silvestre & Eugenia Gallardo

    2. CNC – Center for Neurosciences and Cell Biology, University of Coimbra, Coimbra, 3004-504, Portugal

      Gilberto Alves & Samuel Silvestre

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    Ana Martinho and Sara M. Silva contributed equally to this paper.

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    Martinho, A., Silva, S.M., Garcia, S. et al. Effects of Hypericum perforatum hydroalcoholic extract, hypericin, and hyperforin on cytotoxicity and CYP3A4 mRNA expression in hepatic cell lines: a comparative study. Med Chem Res 25, 2999–3010 (2016). https://doi.org/10.1007/s00044-016-1702-z

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