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Planta Med 2019; 85(11/12): 997-1007
DOI: 10.1055/a-0954-6704
Biological and Pharmacological Activities
Original Papers
Georg Thieme Verlag KG Stuttgart · New York

Silymarin Enriched Extract (Silybum marianum) Additive Effect on Doxorubicin-Mediated Cytotoxicity in PC-3 Prostate Cancer Cells[ 1 ]

Katerina Gioti
1   Department of Nutrition and Dietetics, School of Health Sciences and Education, Harokopio University, Kallithea, Greece
,
Anastasia Papachristodoulou
1   Department of Nutrition and Dietetics, School of Health Sciences and Education, Harokopio University, Kallithea, Greece
2   Faculty of Pharmacy, Department of Pharmaceutical Chemistry, University of Athens, Zografou, Greece
,
Dimitra Benaki
2   Faculty of Pharmacy, Department of Pharmaceutical Chemistry, University of Athens, Zografou, Greece
,
Sophia Havaki
3   Molecular Carcinogenesis Group, Department of Histology and Embryology, School of Medicine, National and Kapodistrian University of Athens, Athens, Greece
,
Apostolos Beloukas
4   Department of Biomedical Sciences, School of Health Sciences, University of West Attica, Greece
5   Department of Clinical Infection, Microbiology and Immunology, Institute of Infection and Global Health, University of Liverpool, Liverpool, United Kingdom
,
Argyro Vontzalidou
6   Faculty of Pharmacy, Department of Pharmacognosy and Natural Products Chemistry, University of Athens, Zografou, Greece
,
Nektarios Aligiannis
6   Faculty of Pharmacy, Department of Pharmacognosy and Natural Products Chemistry, University of Athens, Zografou, Greece
,
Alexios-Leandros Skaltsounis
6   Faculty of Pharmacy, Department of Pharmacognosy and Natural Products Chemistry, University of Athens, Zografou, Greece
,
Emmanuel Mikros
2   Faculty of Pharmacy, Department of Pharmaceutical Chemistry, University of Athens, Zografou, Greece
,
Roxane Tenta
1   Department of Nutrition and Dietetics, School of Health Sciences and Education, Harokopio University, Kallithea, Greece
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Publikationsverlauf

received 16. Februar 2019
revised 06. Juni 2019

accepted 10. Juni 2019

Publikationsdatum:
09. Juli 2019 (online)

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Abstract

Silymarin-enriched extract (SEE) is obtained from Silybum marianum (Asteraceae). Doxorubicin (DXR) is a widely used chemotherapeutical yet with severe side effects. The goal of the present study was to assess the pharmacologic effect of SEE and its bioactive components silibinin and silychristine when administrated alone or in combination with DXR in the human prostate cancer cells (PC-3). PC-3 cells were treated with SEE, silibinin (silybins A and B), silychristine, alone, and in combination with DXR, and cell proliferation was assessed by the MTT assay. Cell cycle, apoptosis, and autophagy rate were assessed by flow cytometry. Expression levels of autophagy-related genes were quantified by qRT-PCR, ELISA and western blot while transmission electron microscopy was performed to reveal autophagic structures. Finally, NMR spectrometry was used to identify specific metabolites related to autophagy. SEE inhibited PC-3 cell proliferation in a dose-dependent manner while the co-treatment (DXR-SEE) revealed an additive cytotoxic effect. Cell cycle, apoptosis, and autophagy variations were observed in addition to altered expression levels of autophagy related genes (LC3, p62, NBR1, Beclin1, ULK1, AMBRA1), while several modifications in autophagic structures were identified after DXR-SEE co-treatment. Furthermore, treated cells showed a different metabolic profile, with significant alterations in autophagy-related metabolites such as branched-chain amino acids. In conclusion, the DXR-SEE co-treatment provokes perturbations in the autophagic mechanism of prostate cancer cells (PC-3) compared to DXR treatment alone, causing an excessive cell death. These findings propose the putative use of SEE as an adjuvant cytotoxic agent.

Key words

Silybum marianum - Asteraceae - silymarin - doxorubicin - prostate cancer - autophagy

1 Dedicated to Professor Dr. Cosimo Pizza 70th birthday in recognition of his outstanding contribution to natural product research.


 

  • References

  • 1 Dean M, Lou H. Genetics and genomics of prostate cancer. Asian J Androl 2013; 15: 309-313
    CrossrefPubMedGoogle Scholar
  • 2 Minotti G, Menna P, Salvatorelli E, Cairo G, Gianni L. Anthracyclines: molecular advances and pharmacologic developments in antitumor activity and cardiotoxicity. Pharmacol Rev 2004; 56: 185-229
    CrossrefPubMedGoogle Scholar
  • 3 Gioti K, Tenta R. Bioactive natural products against prostate cancer: mechanism of action and autophagic/apoptotic molecular pathways. Planta Med 2015; 81: 543-562
    Artikel in Thieme ConnectPubMedGoogle Scholar
  • 4 Davis-Searles PR, Nakanishi Y, Kim NC, Graf TN, Oberlies NH, Wani MC, Wall ME, Agarwal R, Kroll DJ. Milk thistle and prostate cancer: differential effects of pure flavonolignans from Silybum marianum on antiproliferative end points in human prostate carcinoma cells. Cancer Res 2005; 65: 4448-4457
    CrossrefPubMedGoogle Scholar
  • 5 Elyasi S, Hosseini S, Niazi Moghadam MR, Aledavood SA, Karimi G. Effect of oral silymarin administration on prevention of radiotherapy induced mucositis: a randomized, double-blinded, placebo-controlled clinical trial. Phytother Res 2016; 30: 1879-1885
    CrossrefPubMedGoogle Scholar
  • 6 Razavi BM, Karimi G. Protective effect of silymarin against chemical-induced cardiotoxicity. Iran J Basic Med Sci 2016; 19: 916-923
    PubMedGoogle Scholar
  • 7 Ezzat A, Abdelhamid AO, El Awady MK, Abd El Azeem AS, Mohammed DM. The biochemical effects of nano tamoxifen and some bioactive components in experimental breast cancer. Biomed Pharmacother 2017; 95: 571-576
    CrossrefPubMedGoogle Scholar
  • 8 Khorsandi L, Saki G, Bavarsad N, Mombeini M. Silymarin induces a multi-targeted cell death process in the human colon cancer cell line HT-29. Biomed Pharmacother 2017; 94: 890-897
    CrossrefPubMedGoogle Scholar
  • 9 Won DH, Kim LH, Jang B, Yang IH, Kwon HJ, Jin B, Oh SH, Kang JH, Hong SD, Shin JA, Cho SD. In vitro and in vivo anti-cancer activity of silymarin on oral cancer. Tumour Biol 2018; 40: 1010428318776170
    Google Scholar
  • 10 Yurtcu E, Darcansoy Iseri O, Iffet Sahin F. Effects of silymarin and silymarin-doxorubicin applications on telomerase activity of human hepatocellular carcinoma cell line HepG2. J BUON 2015; 20: 555-561
    PubMedGoogle Scholar
  • 11 Singh RP, Agarwal R. Prostate cancer prevention by silibinin. Curr Cancer Drug Targets 2004; 4: 1-11
    CrossrefPubMedGoogle Scholar
  • 12 Tyagi AK, Singh RP, Agarwal C, Chan DC, Agarwal R. Silibinin strongly synergizes human prostate carcinoma DU145 cells to doxorubicin-induced growth Inhibition, G2-M arrest, and apoptosis. Clin Cancer Res 2002; 8: 3512-3519
    PubMedGoogle Scholar
  • 13 Shibutani ST, Yoshimori T. A current perspective of autophagosome biogenesis. Cell Res 2013; 24: 58-68
    PubMedGoogle Scholar
  • 14 Di Bartolomeo S, Corazzari M, Nazio F, Oliverio S, Lisi G, Antonioli M, Pagliarini V, Matteoni S, Fuoco C, Giunta L, DʼAmelio M, Nardacci R, Romagnoli A, Piacentini M, Cecconi F, Fimia GM. The dynamic interac-tion of AMBRA1 with the dynein motor complex regulates mammalian autophagy. J Cell Biol 2010; 191: 155-168
    CrossrefPubMedGoogle Scholar
  • 15 Koukourakis MI, Kalamida D, Giatromanolaki A, Zois CE, Sivridis E, Pouliliou S, Mitrakas A, Gatter KC, Harris AL. Autophagosome proteins LC3A, LC3B and LC3C have distinct subcellular distribution kinetics and expression in cancer cell lines. PLoS One 2015; 10: e0137675
    Google Scholar
  • 16 Mizushima N, Yoshimori T. How to interpret LC3 Immunoblotting. Autophagy 2007; 3: 542-545
    CrossrefPubMedGoogle Scholar
  • 17 Johansen T, Lamark T. Selective autophagy mediated by autophagic adapter proteins. Autophagy 2010; 7: 279-296
    PubMedGoogle Scholar
  • 18 Levine B, Kroemer G. Autophagy in the pathogenesis of disease. Cell 2008; 132: 27-42
    CrossrefPubMedGoogle Scholar
  • 19 Deep G, Singh RP, Agarwal C, Kroll DJ, Agarwal R. Silymarin and silibinin cause G1 and G2-M cell cycle arrest via distinct circuitries in human prostate cancer PC3 cells: a comparison of flavanone silibinin with flavanolignan mixture silymarin. Oncogene 2006; 25: 1053-1069
    CrossrefPubMedGoogle Scholar
  • 20 Mizushima N, Yoshimori T, Levine B. Methods in mammalian autophagy research. Cell 2010; 140: 313-326
    CrossrefPubMedGoogle Scholar
  • 21 Kuzu OF, Gowda R, Sharma A, Robertson GP. Leelamine mediates cancer cell death through inhibition of intracellular cholesterol transport. Mol Cancer Ther 2014; 13: 1690-1703
    CrossrefPubMedGoogle Scholar
  • 22 Kaini RR, Sillerud LO, Zhaorigetu S, Hu CA. Autophagy regulates lipolysis and cell survival through lipid droplet degradation in androgen-sensitive prostate cancer cells. Prostate 2012; 72: 1412-1422
    CrossrefPubMedGoogle Scholar
  • 23 Wang J, Tan X, Yang Q, Zeng X, Zhou Y, Luo W, Lin X, Song L, Cai J, Wang T, Wu X. Inhibition of autophagy promotes apoptosis and enhances anticancer efficacy of adriamycin via augmented ROS generation in prostate cancer cells. Int J Biochem Cell Biol 2016; 77: 80-90
    CrossrefPubMedGoogle Scholar
  • 24 Desideri E, Filomeni G, Ciriolo MR. Glutathione participates in the modulation of starvation-induced autophagy in carcinoma cells. Autophagy 2012; 8: 1769-1781
    CrossrefPubMedGoogle Scholar
  • 25 Duran RV, Oppliger W, Robitaille AM, Heiserich L, Skendaj R, Gottlieb E, Hall MN. Glutaminolysis activates Rag-mTORC1 signaling. Mol Cell 2012; 47: 349-358
    CrossrefPubMedGoogle Scholar
  • 26 Sengupta S, Peterson TR, Sabatini DM. Regulation of the mTOR complex 1 pathway by nutrients, growth factors, and stress. Mol Cell 2010; 40: 310-322
    CrossrefPubMedGoogle Scholar
  • 27 Sutter BM, Wu X, Laxman S, Tu BP. Methionine inhibits autophagy and promotes growth by inducing the SAM-responsive methylation of PP2A. Cell 2013; 154: 403-415
    CrossrefPubMedGoogle Scholar
  • 28 Wianowska D, Wisniewski M. Simplified procedure of silymarin extraction from Silybum marianum L. Gaertner. J Chromatogr Sci 2015; 53: 366-372
    PubMedGoogle Scholar
  • 29 Cheilari A, Sturm S, Intelmann D, Seger C, Stuppner H. Head-to-head comparison of ultra-high-performance liquid chromatography with diode array detection versus quantitative nuclear magnetic resonance for the quantitative analysis of the silymarin complex in Silybum marianum fruit extracts. J Agric Food Chem 2016; 64: 1618-1626
    Google Scholar
  • 30 Vistica DT, Skehan P, Scudiero D, Monks A, Pittman A, Boyd MR. Tetrazolium-based assays for cellular viability: a critical examination of selected parameters affecting formazan production. Cancer Res 1991; 51: 2515-2520
    PubMedGoogle Scholar
  • 31 Cristofani R, Montagnani Marelli M, Cicardi ME, Fontana F, Marzagalli M, Limonta P, Poletti A, Moretti RM. Dual role of autophagy on docetaxel-sensitivity in prostate cancer cells. Cell Death Dis 2018; 9: 889
    CrossrefPubMedGoogle Scholar
  • 32 Stankov MV, El Khatib M, Kumar Thakur B, Heitmann K, Panayotova-Dimitrova D, Schoening J, Bourquin JP, Schweitzer N, Leverkus M, Welte K, Reinhardt D, Li Z, Orkin SH, Behrens GM, Klusmann JH. Histone deacetylase inhibitors induce apoptosis in myeloid leukemia by suppressing autophagy. Leukemia 2013; 28: 577-588
    PubMedGoogle Scholar
  • 33 Kowalik MA, Perra A, Ledda-Columbano GM, Ippolito G, Piacentini M, Columbano A, Falasca L. Induction of autophagy promotes the growth of early preneoplastic rat liver nodules. Oncotarget 2016; 7: 5788-5799
    CrossrefPubMedGoogle Scholar
  • 34 Toepfer N, Childress C, Parikh A, Rukstalis D, Yang W. Atorvastatin induces autophagy in prostate cancer PC3 cells through activation of LC3 transcription. Cancer Biol Ther 2011; 12: 691-699
    CrossrefPubMedGoogle Scholar
  • 35 Le Belle JE, Harris NG, Williams SR, Bhakoo KK. A comparison of cell and tissue extraction techniques using high-resolution 1H-NMR spectroscopy. NMR Biomed 2002; 15: 37-44
    CrossrefPubMedGoogle Scholar