Cancer Chemotherapy and Pharmacology

, Volume 71, Issue 6, pp 1417–1425

Preclinical evaluation of combined TKI-258 and RAD001 in hepatocellular carcinoma

  • Stephen L. Chan
  • Chi-Hang Wong
  • Cecilia P. Y. Lau
  • Qian Zhou
  • Connie W. C. Hui
  • Vivian W. Y. Lui
  • Brigette B. Y. Ma
  • Anthony T. C. Chan
  • Winnie Yeo
Original Article

Abstract

Purpose

RAD001 targets at the mammalian target of rapamycin (mTOR), while TKI-258 is a potent tyrosine kinase inhibitor targeting at fibroblast growth factor receptor, vascular endothelial growth factor receptor, platelet-derived growth factor receptor and c-kit. We aim to study the activity of combined RAD001 and TKI-258 in cell lines and xenograft model of hepatocellular carcinoma (HCC), with reference to the parallel and upstream pathways of Akt–mTOR axis.

Methods

A panel of 4 human HCC cell lines HepG2, Hep3B, PLC/PRF/5 and Huh7 and the Hep3B-derived xenograft were treated with TKI-258 or/and RAD001, respectively. Related mechanistic studies (including apoptosis and angiogenesis) were conducted.

Results

There was an enhanced increase in suppression of cell proliferation with combined TKI-258 and RAD001 compared with either drug alone. The combination could significantly suppress the phosphorylation of mTOR, MEK1/2 and p38 MAPK. Although the addition of the TKI258 only slightly suppressed the phosphorylation of AKT induced by RAD001, the pi-mTOR and its downstream signaling pathways including pi-p70S6K, pi-S6 and pi-4EBP1 were lowered in the combination. In Hep3B-derived xenograft, TKI-258 and RAD001 had shown an enhanced inhibition of tumor growth without impact on the weight of animals. There was a reduction in microvessel density in the xenograft with the combination, which indicated an enhanced inhibition on angiogenesis. Pro-caspases-3 and PARP cleavage were slightly detected at 48 h after treatment, suggesting that the combination mainly increased the cytostatic arrest ability.

Conclusions

The combination of RAD001 and TKI-258 was active in HCC via inhibition of both mTOR-mediated signaling and its parallel pathways.

Keywords

RAD001 TKI-258 Hepatocellular carcinoma AKT mTOR 

References

  1. 1.
    Parkin DM, Bray F, Ferlay J, Pisani P (2005) Global cancer statistics, 2002. CA Cancer J Clin 55(2):74–108PubMedCrossRefGoogle Scholar
  2. 2.
    Chan SL, Yeo W (2012) Targeted therapy of hepatocellular carcinoma: present and future. J Gastroenterol Hepatol 27(5):862–872. doi:10.1111/j.1440-1746.2012.07096.x PubMedCrossRefGoogle Scholar
  3. 3.
    Chan SL, Mo FK, Johnson PJ, Liem GS, Chan TC, Poon MC, Ma BB, Leung TW, Lai PB, Chan AT, Mok TS, Yeo W (2011) Prospective validation of the Chinese University Prognostic Index and comparison with other staging systems for hepatocellular carcinoma in an Asian population. J Gastroenterol Hepatol 26(2):340–347. doi:10.1111/j.1440-1746.2010.06329.x PubMedCrossRefGoogle Scholar
  4. 4.
    Llovet JM, Ricci S, Mazzaferro V, Hilgard P, Gane E, Blanc JF, de Oliveira AC, Santoro A, Raoul JL, Forner A, Schwartz M, Porta C, Zeuzem S, Bolondi L, Greten TF, Galle PR, Seitz JF, Borbath I, Haussinger D, Giannaris T, Shan M, Moscovici M, Voliotis D, Bruix J (2008) Sorafenib in advanced hepatocellular carcinoma. N Engl J Med 359(4):378–390PubMedCrossRefGoogle Scholar
  5. 5.
    Villanueva A, Chiang DY, Newell P, Peix J, Thung S, Alsinet C, Tovar V, Roayaie S, Minguez B, Sole M, Battiston C, Van Laarhoven S, Fiel MI, Di Feo A, Hoshida Y, Yea S, Toffanin S, Ramos A, Martignetti JA, Mazzaferro V, Bruix J, Waxman S, Schwartz M, Meyerson M, Friedman SL, Llovet JM (2008) Pivotal role of mTOR signaling in hepatocellular carcinoma. Gastroenterology 135(6):1972–1983, 1983 e1971–e1911. doi:10.1053/j.gastro.2008.08.008 Google Scholar
  6. 6.
    Sahin F, Kannangai R, Adegbola O, Wang J, Su G, Torbenson M (2004) mTOR and P70 S6 kinase expression in primary liver neoplasms. Clin Cancer Res 10(24):8421–8425. doi:10.1158/1078-0432.CCR-04-0941 PubMedCrossRefGoogle Scholar
  7. 7.
    Huynh H, Chow KH, Soo KC, Toh HC, Choo SP, Foo KF, Poon D, Ngo VC, Tran E (2009) RAD001 (everolimus) inhibits tumour growth in xenograft models of human hepatocellular carcinoma. J Cell Mol Med 13(7):1371–1380. doi:10.1111/j.1582-4934.2008.00364.x PubMedCrossRefGoogle Scholar
  8. 8.
    O’Reilly KE, Rojo F, She QB, Solit D, Mills GB, Smith D, Lane H, Hofmann F, Hicklin DJ, Ludwig DL, Baselga J, Rosen N (2006) mTOR inhibition induces upstream receptor tyrosine kinase signaling and activates Akt. Cancer Res 66(3):1500–1508PubMedCrossRefGoogle Scholar
  9. 9.
    Zhou Q, Lui VW, Lau CP, Cheng SH, Ng MH, Cai Y, Chan SL, Yeo W (2012) Sustained antitumor activity by co-targeting mTOR and the microtubule with temsirolimus/vinblastine combination in hepatocellular carcinoma. Biochem Pharmacol 83(9):1146–1158. doi:10.1016/j.bcp.2012.01.013 PubMedCrossRefGoogle Scholar
  10. 10.
    Wu X, Ge H, Lemon B, Vonderfecht S, Weiszmann J, Hecht R, Gupte J, Hager T, Wang Z, Lindberg R, Li Y (2010) FGF19-induced hepatocyte proliferation is mediated through FGFR4 activation. J Biol Chem 285(8):5165–5170. doi:10.1074/jbc.M109.068783 PubMedCrossRefGoogle Scholar
  11. 11.
    Lee SH, Lopes de Menezes D, Vora J, Harris A, Ye H, Nordahl L, Garrett E, Samara E, Aukerman SL, Gelb AB, Heise C (2005) In vivo target modulation and biological activity of CHIR-258, a multitargeted growth factor receptor kinase inhibitor, in colon cancer models. Clin Cancer Res 11(10):3633–3641. doi:10.1158/1078-0432.CCR-04-2129 PubMedCrossRefGoogle Scholar
  12. 12.
    Renhowe PA, Pecchi S, Shafer CM, Machajewski TD, Jazan EM, Taylor C, Antonios-McCrea W, McBride CM, Frazier K, Wiesmann M, Lapointe GR, Feucht PH, Warne RL, Heise CC, Menezes D, Aardalen K, Ye H, He M, Le V, Vora J, Jansen JM, Wernette-Hammond ME, Harris AL (2009) Design, structure-activity relationships and in vivo characterization of 4-amino-3-benzimidazol-2-ylhydroquinolin-2-ones: a novel class of receptor tyrosine kinase inhibitors. J Med Chem 52(2):278–292. doi:10.1021/jm800790t PubMedCrossRefGoogle Scholar
  13. 13.
    Trudel S, Li ZH, Wei E, Wiesmann M, Chang H, Chen C, Reece D, Heise C, Stewart AK (2005) CHIR-258, a novel, multitargeted tyrosine kinase inhibitor for the potential treatment of t(4;14) multiple myeloma. Blood 105(7):2941–2948. doi:10.1182/blood-2004-10-3913 PubMedCrossRefGoogle Scholar
  14. 14.
    Lopes de Menezes DE, Peng J, Garrett EN, Louie SG, Lee SH, Wiesmann M, Tang Y, Shephard L, Goldbeck C, Oei Y, Ye H, Aukerman SL, Heise C (2005) CHIR-258: a potent inhibitor of FLT3 kinase in experimental tumor xenograft models of human acute myelogenous leukemia. Clin Cancer Res 11(14):5281–5291. doi:10.1158/1078-0432.CCR-05-0358 PubMedCrossRefGoogle Scholar
  15. 15.
    Sarker D, Molife R, Evans TR, Hardie M, Marriott C, Butzberger-Zimmerli P, Morrison R, Fox JA, Heise C, Louie S, Aziz N, Garzon F, Michelson G, Judson IR, Jadayel D, Braendle E, de Bono JS (2008) A phase I pharmacokinetic and pharmacodynamic study of TKI258, an oral, multitargeted receptor tyrosine kinase inhibitor in patients with advanced solid tumors. Clin Cancer Res 14(7):2075–2081. doi:10.1158/1078-0432.CCR-07-1466 PubMedCrossRefGoogle Scholar
  16. 16.
    Huynh H, Chow PK, Tai WM, Choo SP, Chung AY, Ong HS, Soo KC, Ong R, Linnartz R, Shi MM (2012) Dovitinib demonstrates antitumor and antimetastatic activities in xenograft models of hepatocellular carcinoma. J Hepatol 56(3):595–601. doi:10.1016/j.jhep.2011.09.017 PubMedCrossRefGoogle Scholar
  17. 17.
    Tai WT, Cheng AL, Shiau CW, Liu CY, Ko CH, Lin MW, Chen PJ, Chen KF (2012) Dovitinib induces apoptosis and overcomes sorafenib resistance in hepatocellular carcinoma through SHP-1-mediated inhibition of STAT3. Mol Cancer Ther 11(2):452–463. doi:10.1158/1535-7163.MCT-11-0412 PubMedCrossRefGoogle Scholar
  18. 18.
    Abou-Alfa GK, Johnson P, Knox JJ, Capanu M, Davidenko I, Lacava J, Leung T, Gansukh B, Saltz LB (2010) Doxorubicin plus sorafenib vs doxorubicin alone in patients with advanced hepatocellular carcinoma: a randomized trial. JAMA 304(19):2154–2160. doi:10.1001/jama.2010.1672 PubMedCrossRefGoogle Scholar
  19. 19.
    Abou-Alfa GK, Chan SL, Lin CC, Chiorean EG, Holcombe RF, Mulcahy MF, Carter WD, Patel K, Wilson WR, Melink TJ, Gutheil JC, Tsao CJ (2011) PR-104 plus sorafenib in patients with advanced hepatocellular carcinoma. Cancer Chemother Pharmacol 68(2):539–545. doi:10.1007/s00280-011-1671-3 PubMedCrossRefGoogle Scholar
  20. 20.
    Chan SL, Mok T, Ma BB (2012) Management of hepatocellular carcinoma: beyond sorafenib. Curr Oncol Rep. doi:10.1007/s11912-012-0233-0
  21. 21.
    Cheng AL, Kang YK, Chen Z, Tsao CJ, Qin S, Kim JS, Luo R, Feng J, Ye S, Yang TS, Xu J, Sun Y, Liang H, Liu J, Wang J, Tak WY, Pan H, Burock K, Zou J, Voliotis D, Guan Z (2009) Efficacy and safety of sorafenib in patients in the Asia-Pacific region with advanced hepatocellular carcinoma: a phase III randomised, double-blind, placebo-controlled trial. Lancet Oncol 10(1):25–34PubMedCrossRefGoogle Scholar
  22. 22.
    El-Serag HB, Rudolph KL (2007) Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology 132(7):2557–2576PubMedCrossRefGoogle Scholar
  23. 23.
    Ma BB, Lui VW, Hui EP, Lau CP, Ho K, Ng MH, Cheng SH, Tsao SW, Chan AT (2010) The activity of mTOR inhibitor RAD001 (everolimus) in nasopharyngeal carcinoma and cisplatin-resistant cell lines. Invest New Drugs 28(4):413–420. doi:10.1007/s10637-009-9269-x PubMedCrossRefGoogle Scholar
  24. 24.
    Dey JH, Bianchi F, Voshol J, Bonenfant D, Oakeley EJ, Hynes NE (2010) Targeting fibroblast growth factor receptors blocks PI3K/AKT signaling, induces apoptosis, and impairs mammary tumor outgrowth and metastasis. Cancer Res 70(10):4151–4162. doi:10.1158/0008-5472.CAN-09-4479 PubMedCrossRefGoogle Scholar
  25. 25.
    Breuleux M, Klopfenstein M, Stephan C, Doughty CA, Barys L, Maira SM, Kwiatkowski D, Lane HA (2009) Increased AKT S473 phosphorylation after mTORC1 inhibition is rictor dependent and does not predict tumor cell response to PI3K/mTOR inhibition. Mol Cancer Ther 8(4):742–753. doi:10.1158/1535-7163.MCT-08-0668 PubMedCrossRefGoogle Scholar
  26. 26.
    Zhou Q, Wong CH, Lau CPY, Hui CWC, Lui VW, Chan SL, Yeo W (2013) Enhanced antitumor activity with combining effect of mTOR inhibition and in microtubule-stabilization in hepatocellular carcinoma. Int J Hepatol 2013:103830. doi:10.1155/2013/103830

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Stephen L. Chan
    • 1
  • Chi-Hang Wong
    • 1
  • Cecilia P. Y. Lau
    • 1
  • Qian Zhou
    • 1
  • Connie W. C. Hui
    • 1
  • Vivian W. Y. Lui
    • 2
  • Brigette B. Y. Ma
    • 1
  • Anthony T. C. Chan
    • 1
  • Winnie Yeo
    • 1
  1. 1.State Key Laboratory in Oncology in South China, Sir YK Pao Center for Cancer, Department of Clinical Oncology, Cancer Drug Testing Unit, Hong Kong Cancer Institute, Li Ka Shing Institute of Health Sciences and Prince of Wales HospitalThe Chinese University of Hong KongShatin, N.T.Hong Kong
  2. 2.Department of OtolaryngologyUniversity of Pittsburgh School of MedicinePittsburghUSA

Personalised recommendations