Escin, a Natural Mixture of Triterpene Saponins, Exhibits Antitumor Activity Against Hepatocellular Carcinoma

Abstract

Escin, a mixture of triterpene saponins extracted from Aesculus wilsonii Rehd., was used to analyze the antitumor effect in hepatocellular carcinoma in vivo and in vitro. At a dose of 2.8 mg/kg, escin had a rather high inhibition ratio (43.5 %) on mice H22 tumor growth in vivo. The results of the SRB cell viability assay showed that escin could induce significant concentration- and time-dependent inhibition of HepG₂ cell viability. Disruption of the G₁/S phase of cell cycle progression accompanied by the induction of apoptosis were also observed in HepG₂ cells following escin treatment. The results of pulse-field gel electrophoresis and Western blot analysis show the induction of caspase-independent apoptosis by escin. This study provides evidence that escin induces cell cycle checkpoint arrest and caspase-independent cell death in HepG₂ cells, in support of its efficacious potential as a chemopreventive agent.

Key words

escin - antitumor activity - hepatocellular carcinoma - inhibition - cell cycle arrest - caspase‐ independent apoptosis

References

• 1 Llovet J M, Beaugrand M. Hepatocellular carcinoma: present status and future prospects.  J Hepatol. 2003;  38 S136-S149
• 2 German Drug Codex Alimentarius Commission .German drugs code. Aescin, A. 1997: 2030
• 3 Matsuda H, Li Y, Murakami T, Ninomiya K, Yamahara J, Yoshikawa M. Effects of escins Ia, Ib, IIa, and IIb from horse chestnut, the seeds of Aesculus hippocastanum L., on acute inflammation in animals.  Biol Pharm Bull. 1997;  20 1092-1095
• 4 Marhuenda E, Alarcon de la Lastra C, Martin M J. Antisecretory and gastroprotective effects of escine in rats.  Gen Pharmacol. 1994;  25 1213-1219
• 5 Fu F H, Hou Y Z, Jiang W L, Wang R H, Liu K. Escin: inhibiting inflammation and promoting gastrointestinal transit to attenuate formation of postoperative adhesions.  World J Surg. 2005;  29 1614-1620
• 6 Sun F, Fu F H, He J, Dong Q J, Li Z, Gou H T. Dose-and time-effect relationship on anti-inflammation of Aescine-Arginine and its effect on nitric oxide synthesis.  Chin Pharmacol Bull. 2008;  24 672-675
• 7 Konoshima T, Lee K H. Cytotoxic sapogenols from Aesculus hippocastanum.  J Nat Prod. 1986;  49 650-656
• 8 Guo W, Xu B, Yang X W, Liu Q, Cui J R. Preliminary studies of effect sodium aescinate against cancer in vitro and in vivo. .  Chin Pharmacol Bull. 2003;  19 351-352
• 9 Yang X W, Zhao J, Cui J R, Guo W. Studies on the biotransformation of escin Ia by human intestinal bacteria and the anti-tumor activities of desacylla I.  ASNUP. 2004;  36 31-35
• 10 Jagan M RP, Jayadev R, Malisetty V S, Chinthalapally V R. β-Escin inhibits colonic aberrant crypt foci formation in rats and regulates the cell cycle growth by inducing p 21 ^waf1/cip1 in colon cancer cells.  Mol Cancer Ther. 2006;  5 1459-1466
• 11 Zou M, Han Y, Ji X. Escin inhibits growth of HTC‐8 cells and enhances the cells' sensitivity to 5-FU.  Chin J Dig. 2001;  21 248-249
• 12 Skehan P, Storeng R, Scudiero D, Monks A, McMahon J, Vistica D. New colorimetric cytotoxicity assay for anticancer-drug screening.  J Natl Cancer Inst. 1990;  82 1107-1112
• 13 Kawabe Y, Ochi A. Programmed cell death and extrathymic reduction of Vb81 CD41 T cells in mice tolerant to Staphylococcus aureus enterotoxin B.  Nature (London). 1991;  349 245-248
• 14 Brown D G, Sun X M, Cohen G M. Dexamethasone-induced apoptosis involves cleavage of DNA to large fragments prior to internucleosomal fragmentation.  J Biol Chem. 1993;  268 3037-3039
• 15 Feng G P, Kaplowitz N. Mechanism of staurosporine-induced apoptosis in murine hepatocytes.  Am J Physiol Gastrointest Liver Physiol. 2002;  282 G825-G834
• 16 Abrams J M, White K, Fessler L I, Steller H. Programmed cell death during Drosophila embryogenesis.  Development. 1993;  117 29-43
• 17 Hsieh T C, Wu P, Park S, Wu J M. Induction of cell cycle changes and modulation of apoptogenic/anti-apoptotic and extracellular signaling regulatory protein expression by water extracts of I'm-Yunity (PSP).  BMC Complement Altern Med. 2006;  6 30-44
• 18 Watanabe H, Kanbe K, Shinozaki T, Hoshino H, Chigira M. Apoptosis of a fibrosarcoma induced by protein-free culture involves DNA cleavage to large fragments but not internucleosomal fragmentation.  Int J Cancer. 1995;  62 191-198
• 19 Liu X, Zou H, Slaughter C, Wang X. DFF, a heterodimeric protein that functions downstream of caspase 3 to trigger DNA fragmentation during apoptosis.  Cell. 1997;  89 175-184
• 20 Enari M, Sakahira H, Yokoyoma H, Okawa K, Iwamtsu A, Nagata S. A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD.  Nature. 1998;  391 43-50
• 21 Liu X S, Li P, Widlack P, Zou H, Luo X, Garrard W T, Wang X D. The 40-kDa subunit of DNA fragmentation factor induces DNA fragmentation and chromatin condensation during apoptosis.  Proc Natl Acad Sci USA. 1998;  95 8461-8466
• 22 Daugas E, Nochy D, Ravagnan L, Loeffler M, Susin S A, Zamzami N, Kroemer G. Apoptosis-inducing factor (AIF): a ubiquitous mitochondrial oxidoreductase involved in apoptosis.  FEBS Lett. 2000;  476 118-123
• 23 Lipton S A, Bossy-Wetzel E. Dueling activities of AIF in cell death versus survival: DNA binding and redox activity.  Cell. 2002;  111 147-150
• 24 Wang X C, Yang C L, Cai J J, Shi Y G, Xue D. Mechanisms of AIF-mediated apoptotic DNA degradation in Caenorhabditis elegans.  Science. 2002;  298 1587-1592
• 25 Cohen G M. Caspases: the executioners of apoptosis.  Biochem J. 1997;  326 (Pt 1) 1-16
• 26 Woo M, Hakem R, Soengas M S, Ducan G S, Shahinian A, Kägi D, Hakem A, McCurrach M, Khoo W, Kaufman S A, Senaldi G, Howard T, Lowe S W, Mak T W. Essential contribution of caspase 3/CPP32 to apoptosis and its associated nuclear changes.  Genes Dev. 1998;  12 806-819
• 27 Bröker L E, Kruyt F A, Giaccone G. Cell death independent of caspases: a review.  Clin Cancer Res. 2005;  11 3155-3162
• 28 Kunz K, Lorkowski G, Petersen G, Samcova E, Schaffler K, Wauschkuhn C H. Bioavailability of escin after administration of two oral formulations containing aesculus extract.  Arzneimittelforschung. 1998;  48 822-825
• 29 Loew D, Schrodter A, Schwankl W, Marz R W. Measurement of the bioavailability of aescin-containing extracts.  Methods Find Exp Clin Pharmacol. 2000;  22 537-542
• 30 Schrader E, Schwankl W, Sieder C, Christoffel V. Comparison of the bioavailability of beta-aescin after single oral administration of two different drug formulations containing an extract of horse-chestnut seeds.  Pharmazie. 1995;  50 623-627
• 31 Hu X M, Zeng F D. Pharmacokinetics of sodium β-aescinate induced by ischemia-reperfusion in cerebral damage rats.  Chin J Clin Pharmacol Ther. 2005;  10 828-831