肝癌是全球最盛行的癌症之一，在台灣也是最常見之惡性腫瘤之一。肝細胞的惡性轉化通常發生於慢性肝細胞損傷及細胞再生後，伴隨著基因的突變以及生長因子的調控。目前已發現有一系列的生長因子對於肝細胞的分化與再生扮演著重要角色，脫序的生長因子調控可能是形成肝癌的重要原因。對於肝癌的治療主要有外科手術切除(包括肝臟移植)、經動脈栓塞、酒精注射等方式。然而治療效果並不理想，主要原因為大部分病患屬較末期個案，因此有必要發展新的治療方法。然而對於開發新穎的肝癌治療方法，設計原位性肝癌(orthotopic hepatoma)是一個重要的步驟。藉由超音波技術，可在免疫系統健全的 Sprague-Dawley (SD) 大白鼠肝葉中注射 Novikoff (N1-S1) 肝腫瘤細胞造成肝癌的表現。此外，超音波也可以作為監測腫瘤在動物?堛漸耵爣“峞C持續及非侵入性的原位肝癌檢測可運用於衡量肝癌的藥物治療。在第一章中，研究採用一種比較非侵犯性的方法在免疫健全的大鼠中建立原位肝癌模式。這是透過超音波引導下植入癌細胞而達成，並以此模式來評估短期和低劑量 epirubicin 之療效。超音波引導下植入 Novikoff 肝癌細胞可導致 60.4％ 的 SD 大鼠產生原位肝癌。除此之外，超音波測量腫瘤大小與犧牲動物後以卡鉗測量的結果有顯著相關(P < 0.00001)。超音波引導下植入所誘導出的腫瘤比率與外科手術相似(55/91, 60.4% vs. 39/52, 75%)，且兩種方法所引發的腫瘤大小並無顯著差異。接著，超音波和電腦斷層掃描分別測量出的腫瘤大小有顯著的相關性。在荷瘤大鼠以短期和低劑量之epirubicin 作化療可有效抑制腫瘤的生長，且證明與細胞凋亡和抑制細胞增生以及減少腫瘤微血管密度有關。在第二章中主要研究 celecoxib 對大鼠原位肝癌的化學預防效果及所參與之訊息傳導途徑。 Novikoff 肝癌的 COX–2 (Cyclooxygenase-2) 表現與人類肝癌是一致的。西方墨點和 PCR 檢測證明 N1-S1 肝癌細胞呈現低量 COX–2 &#37238;的表現。接著，體外實驗證明低劑量 celecoxib 能有效地抑制 N1-S1 細胞的增生和增加癌細胞凋亡，同時對正常的肝細胞是安全的。此外，celecoxib 的化學預防效果也呈現在抑制大鼠原位肝癌的生長上。腫瘤的發病率雖然未受 celecoxib 影響，但是，腫瘤重量的增加有顯著被抑制。Celecoxib 在動物模式中顯示的化學預防機制被認為與藥物的抗血管新生、抗細胞增生以及抗肝癌幹細胞(hCSC)的特性有關。在第三章中測試了合併低劑量 celecoxib 和 epirubicin 對肝癌細胞生長的聯合抑制效應。結果顯示此方法能有效地抑制肝癌幹細胞，腫瘤血管新生和腫瘤細胞增生，以及促進癌細胞凋亡。這些都與此兩種藥物個別在前面的兩章中提及對肝癌的治療效果吻合。一般情況下，比起個別藥物單獨使用來說，合併療法對肝癌顯示比較強的抑制效果，且有較低的骨髓抑制副作用。總結： 超音波引導下植入 Novikoff 肝癌細胞可有效地建立大鼠原位肝癌模式，且適合和方便應用在治療抗肝癌藥物上的試驗。在當前的研究中，我們在動物模式中證明單獨或合併使用低劑量的 epirubicin 和 celecoxib 兩個藥物對肝癌的有效性。

Hepatocellular carcinoma (HCC) is one of the most prevalent cancers in the world and Taiwan. The major factors involved in the molecular pathogenesis for the development of HCC had been explored in recent years. An extensive array of growth factors and their receptors had been identified and may act as positive and negative modulators in different stages of hepatocarcinogenesis. Current therapeutic approaches for HCC include surgical resection (include liver transplantation), trans-arterial embolization (TAE), alcohol injection, etc. However, the effect is limited due to most of the HCC patients present with advanced stages of the disease. Therefore, this underscores the need for the development of novel therapeutic strategies. It is pivotal to set up an orthotopic hepatoma model for the development of novel intervention strategies for HCC. Under the guidance of ultrasound, we are able to create hepatoma in the liver lobe of Sprague-Dawley (SD) rats by injection of Novikoff (N1-S1) hepatoma cells. In addition, sonographic technique was employed for the monitoring of tumor growth in this animal model in the following subprojects. The continuous, non-invasive measurement of orthotopic hepatoma development will be a valuable tool for the evaluation of effects of drugs for treatment of HCC. In Chapter 1, the study employed a relatively non-invasive approach to establish an orthotopic HCC model in immune-competent rats. This was done by ultrasound-guided implantation of cancer cells and the model was used to evaluate the therapeutic efficacy of short-term and low-dose epirubicin chemotherapy. Ultrasound-guided implantation of Novikoff hepatoma cells led to the formation of orthotopic HCC in 60.4% of the SD rats. Moreover, tumor sizes measured by ultrasound significantly correlated with those measured by calipers after sacrificing the animals (P < 0.00001). The rate of tumor induction by ultrasound-guided implantation was comparable to that of laparotomy (55/91, 60.4% vs. 39/52, 75%) and no significant difference in sizes of tumor was noted between the two groups. Moreover, there was a significant correlation in tumor size measurement by ultrasound and computerized tomography. In tumor-bearing rats, short-term and low-dose epirubicin chemotherapy caused a significant reduction in tumor growth, and was found to be associated with enhanced apoptosis and attenuated proliferation as well as a decrease in microvessel density in tumors. In chapter 2, we investigated the chemopreventive effects of celecoxib in the growth of orthotopic rat HCC and the possible signal pathways involved. The status of COX-2 expression in rat Novikoff HCC was consistent with that in human HCC. Both Western blot and PCR tests had proved that N1-S1 was a HCC model presenting with low COX-2 enzymes in tumor cells. Then, low doses of celecoxib was shown to effectively inhibited the proliferation and increased the apoptosis of N1-S1 cells in vitro, which were also safe to the normal hepatocytes. Moreover, chemoprevention by celecoxib inhibiting the HCC tumor growth was shown in rat orthotropic HCC model. Tumor incidence was not affected by the celecoxib prevention, but, tumor weight was found significantly suppressed by the drug. Possible mechanisms of chemoprevention by celecoxib seen in the animal model were thought to be related to the anti-angiogenic, anti-proliferative and anti-hCSC characters of the drug. In chapter 3, we tried to test the combined inhibitory effects of low doses of celecoxib and epirubicin on the growth of HCC. Combined low doses of epirubicin and celecoxib was effective in inhibiting the hepatic cancer stem cells, tumor angiogenesis, tumor cell proliferation, as well as promoting cancer apoptosis. These are compatible with the effects of the individual drugs on HCC growth shown in the previous two chapters. In general, combination therapy expressed more effectiveness in tumor suppression and less bone marrow suppression than the individual drugs used alone. Taken together, ultrasound-guided implantation of Novikoff hepatoma cells is an effective means of establishing orthotopic HCC in SD rats, which is suitable and convenient for therapeutic trial of anti-HCC treatment. In the current study, we had proved the efficacies of low doses of two drugs, epirubicin and celecoxib, acting individually, as well as the combined effects of them in treating HCC in this model.

致謝-------------------------------------------------------------------------------- 1
推薦序----------------------------------------------------------------------------- 3
INDEX----------------------------------------------------------------------------- 4
LIST OF FIGURES------------------------------------------------------------ 6
摘要--------------------------------------------------------------------------------9
ABSTRACT---------------------------------------------------------------------12
CHAPTER 1
Rapid induction of Orthotopic Hepatocellular Carcinoma in
Immune-competent Rats by Non-invasive Ultrasound-guided Cells
Implantation and the Subsequent Low-dose Epirubicin Chemotherapeutic
Trial in Rats Bearing the Established HCC
Background-----------------------------------------------------------------------17
Materials and methods----------------------------------------------------------21
Results-----------------------------------------------------------------------------27
Discussion------------------------------------------------------------------------32
Conclusions-----------------------------------------------------------------------36
CHAPTER 2
Chemopreventive Effects of Celecoxib on the Growth of Orthotopic Rat
Hepatocellular Carcinoma
Background-----------------------------------------------------------------------38
Materials and methods----------------------------------------------------------40
Results-----------------------------------------------------------------------------46
Discussion------------------------------------------------------------------------50
Conclusions-----------------------------------------------------------------------54
CHAPTER 3
Combined Celecoxib and Epirubicin Suppressed the Growth of Rat
Hepatocellular Carcinoma
Background-----------------------------------------------------------------------56
Materials and methods----------------------------------------------------------59
Results-----------------------------------------------------------------------------65
Discussion------------------------------------------------------------------------69
Conclusions-----------------------------------------------------------------------73
FUTURE PERSPECTIVE---------------------------------------------------74
REFERENCES-----------------------------------------------------------------76
FIGURES AND LEGENDS--------------------------------------------------93
PUBLICATIONS-------------------------------------------------------------118
CONFERENCES-------------------------------------------------------------123

1. El-Serag HB: Hepatocellular carcinoma: an epidemiologic view. J Clin Gastroenterol 2002, 35(5 Suppl 2):S72-78.
2. Trevisani F, D'Intino PE, Grazi GL, Caraceni P, Gasbarrini A, Colantoni A, Stefanini GF, Mazziotti A, Gozzetti G, Gasbarrini G et al: Clinical and pathologic features of hepatocellular carcinoma in young and older Italian patients. Cancer 1996, 77(11):2223-2232.
3. Befeler AS, Di Bisceglie AM: Hepatocellular carcinoma: diagnosis and treatment. Gastroenterology 2002, 122(6):1609-1619.
4. Bruix J, Llovet JM: HCC surveillance: who is the target population? Hepatology 2003, 37(3):507-509.
5. Poon D, Anderson BO, Chen LT, Tanaka K, Lau WY, Van Cutsem E, Singh H, Chow WC, Ooi LL, Chow P et al: Management of hepatocellular carcinoma in Asia: consensus statement from the Asian Oncology Summit 2009. The lancet oncology 2009, 10(11):1111-1118.
6. Di Maio M, De Maio E, Perrone F, Pignata S, Daniele B: Hepatocellular carcinoma: systemic treatments. J Clin Gastroenterol 2002, 35(5 Suppl 2):S109-114.
7. Armengol C, Tarafa G, Boix L, Sole M, Queralt R, Costa D, Bachs O, Bruix J, Capella G: Orthotopic implantation of human hepatocellular carcinoma in mice: analysis of tumor progression and establishment of the BCLC-9 cell line. Clin Cancer Res 2004, 10(6):2150-2157.
8. Schmitz V, Tirado-Ledo L, Tiemann K, Raskopf E, Heinicke T, Ziske C, Gonzalez-Carmona MA, Rabe C, Wernert N, Prieto J et al: Establishment of an orthotopic tumour model for hepatocellular carcinoma and non-invasive in vivo tumour imaging by high resolution ultrasound in mice. J Hepatol 2004, 40(5):787-791.
9. Meloni MF, Livraghi T, Filice C, Lazzaroni S, Calliada F, Perretti L: Radiofrequency ablation of liver tumors: the role of microbubble ultrasound contrast agents. Ultrasound Q 2006, 22(1):41-47.
10. Pohl J, Zuna I, Stremmel W, Rudi J: Systemic chemotherapy with epirubicin for treatment of advanced or multifocal hepatocellular carcinoma. Chemotherapy 2001, 47(5):359-365.
11. Zhu AX, Fuchs CS, Clark JW, Muzikansky A, Taylor K, Sheehan S, Tam K, Yung E, Kulke MH, Ryan DP: A phase II study of epirubicin and thalidomide in unresectable or metastatic hepatocellular carcinoma. Oncologist 2005, 10(6):392-398.
12. Hamada A, Yamakado K, Nakatsuka A, Takaki H, Akeboshi M, Takeda K: Hepatic arterial infusion chemotherapy with use of an implanted port system in patients with advanced hepatocellular carcinoma: prognostic factors. J Vasc Interv Radiol 2004, 15(8):835-841.
13. Plosker GL, Faulds D: Epirubicin. A review of its pharmacodynamic and pharmacokinetic properties, and therapeutic use in cancer chemotherapy. Drugs 1993, 45(5):788-856.
14. Browder T, Butterfield CE, Kraling BM, Shi B, Marshall B, O'Reilly MS, Folkman J: Antiangiogenic scheduling of chemotherapy improves efficacy against experimental drug-resistant cancer. Cancer Res 2000, 60(7):1878-1886.
15. Klement G, Baruchel S, Rak J, Man S, Clark K, Hicklin DJ, Bohlen P, Kerbel RS: Continuous low-dose therapy with vinblastine and VEGF receptor-2 antibody induces sustained tumor regression without overt toxicity. J Clin Invest 2000, 105(8):R15-24.
16. Lam T, Hetherington JW, Greenman J, Maraveyas A: From total empiricism to a rational design of metronomic chemotherapy phase I dosing trials. Anticancer Drugs 2006, 17(2):113-121.
17. Yu J, Qiao L, Zimmermann L, Ebert MP, Zhang H, Lin W, Rocken C, Malfertheiner P, Farrell GC: Troglitazone inhibits tumor growth in hepatocellular carcinoma in vitro and in vivo. Hepatology 2006, 43(1):134-143.
18. Kan Z, Phongkitkarun S, Kobayashi S, Tang Y, Ellis LM, Lee TY, Charnsangavej C: Functional CT for quantifying tumor perfusion in antiangiogenic therapy in a rat model. Radiology 2005, 237(1):151-158.
19. Bertazzoli C, Rovero C, Ballerini L, Lux B, Balconi F, Antongiovanni V, Magrini U: Experimental systemic toxicology of 4'-epidoxorubicin, a new, less cardiotoxic anthracycline antitumor agent. Toxicol Appl Pharmacol 1985, 79(3):412-422.
20. Launchbury AP, Habboubi N: Epirubicin and doxorubicin: a comparison of their characteristics, therapeutic activity and toxicity. Cancer Treat Rev 1993, 19(3):197-228.
21. Abou-Alfa GK, Huitzil-Melendez FD, O'Reilly EM, Saltz LB: Current management of advanced hepatocellular carcinoma. Gastrointest Cancer Res 2008, 2(2):64-70.
22. Hahnfeldt P, Folkman J, Hlatky L: Minimizing long-term tumor burden: the logic for metronomic chemotherapeutic dosing and its antiangiogenic basis. J Theor Biol 2003, 220(4):545-554.
23. Kerbel RS, Rak J, Kobayashi H, Man MS, St Croix B, Graham CH: Multicellular resistance: a new paradigm to explain aspects of acquired drug resistance of solid tumors. Cold Spring Harb Symp Quant Biol 1994, 59:661-672.
24. Jain RK: Delivery of novel therapeutic agents in tumors: physiological barriers and strategies. J Natl Cancer Inst 1989, 81(8):570-576.
25. Jain RK: Tumor angiogenesis and accessibility: role of vascular endothelial growth factor. Semin Oncol 2002, 29(6 Suppl 16):3-9.
26. Gorski DH, Beckett MA, Jaskowiak NT, Calvin DP, Mauceri HJ, Salloum RM, Seetharam S, Koons A, Hari DM, Kufe DW et al: Blockage of the vascular endothelial growth factor stress response increases the antitumor effects of ionizing radiation. Cancer Res 1999, 59(14):3374-3378.
27. Kamat AA, Kim TJ, Landen CN, Jr., Lu C, Han LY, Lin YG, Merritt WM, Thaker PH, Gershenson DM, Bischoff FZ et al: Metronomic chemotherapy enhances the efficacy of antivascular therapy in ovarian cancer. Cancer Res 2007, 67(1):281-288.
28. Colleoni M, Orlando L, Sanna G, Rocca A, Maisonneuve P, Peruzzotti G, Ghisini R, Sandri MT, Zorzino L, Nole F et al: Metronomic low-dose oral cyclophosphamide and methotrexate plus or minus thalidomide in metastatic breast cancer: antitumor activity and biological effects. Ann Oncol 2006, 17(2):232-238.
29. Watanuki A, Ohwada S, Fukusato T, Makita F, Yamada T, Kikuchi A, Morishita Y: Prognostic significance of DNA topoisomerase IIalpha expression in human hepatocellular carcinoma. Anticancer Res 2002, 22(2B):1113-1119.
30. Wong N, Yeo W, Wong WL, Wong NL, Chan KY, Mo FK, Koh J, Chan SL, Chan AT, Lai PB et al: TOP2A overexpression in hepatocellular carcinoma correlates with early age onset, shorter patients survival and chemoresistance. Int J Cancer 2009, 124(3):644-652.
31. Edeline J, Raoul JL, Vauleon E, Guillygomac'h A, Boudjema K, Boucher E: Systemic chemotherapy for hepatocellular carcinoma in non-cirrhotic liver: a retrospective study. World J Gastroenterol 2009, 15(6):713-716.
32. Takamatsu M, Matsuda T, Kawaguchi K, Ku Y: [A complete response to one-shot hepatic arterial infusion of epirubicin in a patient with highly advanced hepatocellular carcinoma]. Gan To Kagaku Ryoho 2007, 34(12):2102-2104.
33. Luo JJ, Yan ZP, Wang JH, Liu QX, Chen Y: [Transhepatic arterial chemoembolization by epirubicin mixed with microspheres for hepatocellular carcinoma]. Zhonghua Zhong Liu Za Zhi 2007, 29(8):619-622.
34. Ishikawa T, Imai M, Kamimura H, Tsuchiya A, Togashi T, Watanabe K, Seki K, Ohta H, Yoshida T, Kamimura T: Improved survival for hepatocellular carcinoma with portal vein tumor thrombosis treated by intra-arterial chemotherapy combining etoposide, carboplatin, epirubicin and pharmacokinetic modulating chemotherapy by 5-FU and enteric-coated tegafur/uracil: a pilot study. World J Gastroenterol 2007, 13(41):5465-5470.
35. Tanaka T, Ikeda M, Okusaka T, Ueno H, Morizane C, Ogura T, Hagihara A, Iwasa S: A phase II trial of transcatheter arterial infusion chemotherapy with an epirubicin-Lipiodol emulsion for advanced hepatocellular carcinoma refractory to transcatheter arterial embolization. Cancer Chemother Pharmacol 2008, 61(4):683-688.
36. Yamashita S, Niinobu T, Nakagawa S, Amano M, Nishikawa Y, Hayashida H, Higaki N, Fujita M, Sakon M: [Hepatic arterial infusion chemotherapy for advanced hepatocellular carcinoma providing a good QOL]. Gan To Kagaku Ryoho 2006, 33(12):1928-1930.
37. Sergio A, Cristofori C, Cardin R, Pivetta G, Ragazzi R, Baldan A, Girardi L, Cillo U, Burra P, Giacomin A et al: Transcatheter arterial chemoembolization (TACE) in hepatocellular carcinoma (HCC): the role of angiogenesis and invasiveness. Am J Gastroenterol 2008, 103(4):914-921.
38. Satti J: The emerging low-dose therapy for advanced cancers. Dose Response 2009, 7(3):208-220.
39. Tang TC, Man S, Lee CR, Xu P, Kerbel RS: Impact of metronomic UFT/cyclophosphamide chemotherapy and antiangiogenic drug assessed in a new preclinical model of locally advanced orthotopic hepatocellular carcinoma. Neoplasia 2010, 12(3):264-274.
40. Brown JR, DuBois RN: COX-2: a molecular target for colorectal cancer prevention. J Clin Oncol 2005, 23(12):2840-2855.
41. Kondo M, Yamamoto H, Nagano H, Okami J, Ito Y, Shimizu J, Eguchi H, Miyamoto A, Dono K, Umeshita K et al: Increased expression of COX-2 in nontumor liver tissue is associated with shorter disease-free survival in patients with hepatocellular carcinoma. Clin Cancer Res 1999, 5(12):4005-4012.
42. Giannitrapani L, Ingrao S, Soresi M, Florena AM, La Spada E, Sandonato L, D'Alessandro N, Cervello M, Montalto G: Cyclooxygenase-2 expression in chronic liver diseases and hepatocellular carcinoma: an immunohistochemical study. Annals of the New York Academy of Sciences 2009, 1155:293-299.
43. Leng J, Han C, Demetris AJ, Michalopoulos GK, Wu T: Cyclooxygenase-2 promotes hepatocellular carcinoma cell growth through Akt activation: evidence for Akt inhibition in celecoxib-induced apoptosis. Hepatology 2003, 38(3):756-768.
44. Behari J, Zeng G, Otruba W, Thompson MD, Muller P, Micsenyi A, Sekhon SS, Leoni L, Monga SP: R-Etodolac decreases beta-catenin levels along with survival and proliferation of hepatoma cells. J Hepatol 2007, 46(5):849-857.
45. Marquez-Rosado L, Trejo-Solis MC, Garcia-Cuellar CM, Villa-Trevino S: Celecoxib, a cyclooxygenase-2 inhibitor, prevents induction of liver preneoplastic lesions in rats. J Hepatol 2005, 43(4):653-660.
46. Chan HH, Chu TH, Chien HF, Sun CK, Wang EM, Pan HB, Kuo HM, Hu TH, Lai KH, Cheng JT et al: Rapid induction of orthotopic hepatocellular carcinoma in immune-competent rats by non-invasive ultrasound-guided cells implantation. BMC Gastroenterol, 10(1):83.
47. Ponthan F, Wickstrom M, Gleissman H, Fuskevag OM, Segerstrom L, Sveinbjornsson B, Redfern CP, Eksborg S, Kogner P, Johnsen JI: Celecoxib prevents neuroblastoma tumor development and potentiates the effect of chemotherapeutic drugs in vitro and in vivo. Clin Cancer Res 2007, 13(3):1036-1044.
48. Thun MJ, Henley SJ, Patrono C: Nonsteroidal anti-inflammatory drugs as anticancer agents: mechanistic, pharmacologic, and clinical issues. J Natl Cancer Inst 2002, 94(4):252-266.
49. Koga H, Sakisaka S, Ohishi M, Kawaguchi T, Taniguchi E, Sasatomi K, Harada M, Kusaba T, Tanaka M, Kimura R et al: Expression of cyclooxygenase-2 in human hepatocellular carcinoma: relevance to tumor dedifferentiation. Hepatology 1999, 29(3):688-696.
50. Fantappie O, Solazzo M, Lasagna N, Platini F, Tessitore L, Mazzanti R: P-glycoprotein mediates celecoxib-induced apoptosis in multiple drug-resistant cell lines. Cancer research 2007, 67(10):4915-4923.
51. Kern MA, Haugg AM, Koch AF, Schilling T, Breuhahn K, Walczak H, Fleischer B, Trautwein C, Michalski C, Schulze-Bergkamen H et al: Cyclooxygenase-2 inhibition induces apoptosis signaling via death receptors and mitochondria in hepatocellular carcinoma. Cancer research 2006, 66(14):7059-7066.
52. Cui W, Yu CH, Hu KQ: In vitro and in vivo effects and mechanisms of celecoxib-induced growth inhibition of human hepatocellular carcinoma cells. Clin Cancer Res 2005, 11(22):8213-8221.
53. Cui W, Hu SX, Tang ZY, Hu KQ: In-vivo effects and mechanisms of celecoxib-reduced growth of cyclooxygenase-2 (COX-2)-expressing versus COX-2-deleted human HCC xenografts in nude mice. Anticancer Drugs 2008, 19(9):891-897.
54. Song HJ, Kim YS, Han CH, Jang JY, Kim JH, Cheon YK, Kim YS, Moon JH, Cho YD, Shim CS et al: [A case of complete regression of hepatocellular carcinoma during administration of COX-2 inhibitor]. The Korean journal of hepatology 2006, 12(3):449-454.
55. Masferrer JL, Leahy KM, Koki AT, Zweifel BS, Settle SL, Woerner BM, Edwards DA, Flickinger AG, Moore RJ, Seibert K: Antiangiogenic and antitumor activities of cyclooxygenase-2 inhibitors. Cancer research 2000, 60(5):1306-1311.
56. Majima M, Isono M, Ikeda Y, Hayashi I, Hatanaka K, Harada Y, Katsumata O, Yamashina S, Katori M, Yamamoto S: Significant roles of inducible cyclooxygenase (COX)-2 in angiogenesis in rat sponge implants. Jpn J Pharmacol 1997, 75(2):105-114.
57. Tomozawa S, Nagawa H, Tsuno N, Hatano K, Osada T, Kitayama J, Sunami E, Nita ME, Ishihara S, Yano H et al: Inhibition of haematogenous metastasis of colon cancer in mice by a selective COX-2 inhibitor, JTE-522. Br J Cancer 1999, 81(8):1274-1279.
58. Leahy KM, Ornberg RL, Wang Y, Zweifel BS, Koki AT, Masferrer JL: Cyclooxygenase-2 inhibition by celecoxib reduces proliferation and induces apoptosis in angiogenic endothelial cells in vivo. Cancer research 2002, 62(3):625-631.
59. Ng IO, Poon RT, Lee JM, Fan ST, Ng M, Tso WK: Microvessel density, vascular endothelial growth factor and its receptors Flt-1 and Flk-1/KDR in hepatocellular carcinoma. Am J Clin Pathol 2001, 116(6):838-845.
60. Xie YH, Yuan MB: [A relationship between cyclooxygenase-2 expression and tumor angiogenesis in experimental rat liver carcinogenesis]. Zhonghua Gan Zang Bing Za Zhi 2006, 14(9):676-679.
61. Li W, Tan D, Zhang Z, Liang JJ, Brown RE: Activation of Akt-mTOR-p70S6K pathway in angiogenesis in hepatocellular carcinoma. Oncol Rep 2008, 20(4):713-719.
62. Dong ZZ, Yao DF, Wu W, Yao M, Yu HB, Shen JJ, Qiu LW, Yao NH, Sai WL, Yang JL: Delayed hepatocarcinogenesis through antiangiogenic intervention in the nuclear factor-kappa B activation pathway in rats. Hepatobiliary Pancreat Dis Int, 9(2):169-174.
63. Omar HA, Sargeant AM, Weng JR, Wang D, Kulp SK, Patel T, Chen CS: Targeting of the Akt-nuclear factor-kappa B signaling network by [1-(4-chloro-3-nitrobenzenesulfonyl)-1H-indol-3-yl]-methanol (OSU-A9), a novel indole-3-carbinol derivative, in a mouse model of hepatocellular carcinoma. Mol Pharmacol 2009, 76(5):957-968.
64. Whittaker S, Marais R, Zhu AX: The role of signaling pathways in the development and treatment of hepatocellular carcinoma. Oncogene, 29(36):4989-5005.
65. Cui J, Zhou X, Liu Y, Tang Z, Romeih M: Alterations of beta-catenin and Tcf-4 instead of GSK-3beta contribute to activation of Wnt pathway in hepatocellular carcinoma. Chinese medical journal 2003, 116(12):1885-1892.
66. Yang XR, Xu Y, Yu B, Zhou J, Li JC, Qiu SJ, Shi YH, Wang XY, Dai Z, Shi GM et al: CD24 is a novel predictor for poor prognosis of hepatocellular carcinoma after surgery. Clin Cancer Res 2009, 15(17):5518-5527.
67. Yao Z, Mishra L: Cancer stem cells and hepatocellular carcinoma. Cancer biology & therapy 2009, 8(18):1691-1698.
68. Zhu Z, Hao X, Yan M, Yao M, Ge C, Gu J, Li J: Cancer stem/progenitor cells are highly enriched in CD133+CD44+ population in hepatocellular carcinoma. Int J Cancer, 126(9):2067-2078.
69. Yip WM, Hung HG, Lok KH, Li KF, Li KK, Szeto ML: Outcome of inoperable hepatocellular carcinoma patients receiving transarterial chemoembolisation: a real-life retrospective analysis in a Hong Kong regional hospital. Hong Kong medical journal = Xianggang yi xue za zhi / Hong Kong Academy of Medicine 2009, 15(5):339-345.
70. Rampone B, Schiavone B, Martino A, Viviano C, Confuorto G: Current management strategy of hepatocellular carcinoma. World J Gastroenterol 2009, 15(26):3210-3216.
71. El-Serag HB, Mason AC: Rising incidence of hepatocellular carcinoma in the United States. The New England journal of medicine 1999, 340(10):745-750.
72. Ryberg M, Nielsen D, Cortese G, Nielsen G, Skovsgaard T, Andersen PK: New insight into epirubicin cardiac toxicity: competing risks analysis of 1097 breast cancer patients. J Natl Cancer Inst 2008, 100(15):1058-1067.
73. Gao J, Jia WD, Li JS, Wang W, Xu GL, Ma JL, Ge YS, Yu JH, Ren WH, Liu WB et al: Combined inhibitory effects of celecoxib and fluvastatin on the growth of human hepatocellular carcinoma xenografts in nude mice. The Journal of international medical research, 38(4):1413-1427.
74. A new prognostic system for hepatocellular carcinoma: a retrospective study of 435 patients: the Cancer of the Liver Italian Program (CLIP) investigators. Hepatology 1998, 28(3):751-755.
75. Czauderna P, Mackinlay G, Perilongo G, Brown J, Shafford E, Aronson D, Pritchard J, Chapchap P, Keeling J, Plaschkes J et al: Hepatocellular carcinoma in children: results of the first prospective study of the International Society of Pediatric Oncology group. J Clin Oncol 2002, 20(12):2798-2804.
76. Llovet JM, Ricci S, Mazzaferro V, Hilgard P, Gane E, Blanc JF, de Oliveira AC, Santoro A, Raoul JL, Forner A et al: Sorafenib in advanced hepatocellular carcinoma. The New England journal of medicine 2008, 359(4):378-390.
77. Cabrera R, Nelson DR: Review article: the management of hepatocellular carcinoma. Aliment Pharmacol Ther 2010, 31(4):461-476.
78. Cianchi F, Cortesini C, Bechi P, Fantappie O, Messerini L, Vannacci A, Sardi I, Baroni G, Boddi V, Mazzanti R et al: Up-regulation of cyclooxygenase 2 gene expression correlates with tumor angiogenesis in human colorectal cancer. Gastroenterology 2001, 121(6):1339-1347.
79. Bamba H, Ota S, Kato A, Kawamoto C, Fujiwara K: Prostaglandins up-regulate vascular endothelial growth factor production through distinct pathways in differentiated U937 cells. Biochem Biophys Res Commun 2000, 273(2):485-491.
80. Tsujii M, Kawano S, Tsuji S, Sawaoka H, Hori M, DuBois RN: Cyclooxygenase regulates angiogenesis induced by colon cancer cells. Cell 1998, 93(5):705-716.
81. Murono S, Inoue H, Tanabe T, Joab I, Yoshizaki T, Furukawa M, Pagano JS: Induction of cyclooxygenase-2 by Epstein-Barr virus latent membrane protein 1 is involved in vascular endothelial growth factor production in nasopharyngeal carcinoma cells. Proc Natl Acad Sci U S A 2001, 98(12):6905-6910.
82. Nzeako UC, Guicciardi ME, Yoon JH, Bronk SF, Gores GJ: COX-2 inhibits Fas-mediated apoptosis in cholangiocarcinoma cells. Hepatology 2002, 35(3):552-559.
83. Sheng H, Shao J, Dubois RN: K-Ras-mediated increase in cyclooxygenase 2 mRNA stability involves activation of the protein kinase B1. Cancer Res 2001, 61(6):2670-2675.
84. Sheng H, Shao J, Morrow JD, Beauchamp RD, DuBois RN: Modulation of apoptosis and Bcl-2 expression by prostaglandin E2 in human colon cancer cells. Cancer Res 1998, 58(2):362-366.
85. Bellacosa A, Kumar CC, Di Cristofano A, Testa JR: Activation of AKT kinases in cancer: implications for therapeutic targeting. Adv Cancer Res 2005, 94:29-86.
86. Meng Q, Xia C, Fang J, Rojanasakul Y, Jiang BH: Role of PI3K and AKT specific isoforms in ovarian cancer cell migration, invasion and proliferation through the p70S6K1 pathway. Cell Signal 2006, 18(12):2262-2271.
87. Gonzalez E, McGraw TE: The Akt kinases: isoform specificity in metabolism and cancer. Cell Cycle 2009, 8(16):2502-2508.
88. Chau NM, Ashcroft M: Akt2: a role in breast cancer metastasis. Breast Cancer Res 2004, 6(1):55-57.
89. Davies MA, Stemke-Hale K, Tellez C, Calderone TL, Deng W, Prieto VG, Lazar AJ, Gershenwald JE, Mills GB: A novel AKT3 mutation in melanoma tumours and cell lines. Br J Cancer 2008, 99(8):1265-1268.
90. Kim MS, Jeong EG, Yoo NJ, Lee SH: Mutational analysis of oncogenic AKT E17K mutation in common solid cancers and acute leukaemias. Br J Cancer 2008, 98(9):1533-1535.
91. Bellacosa A, Testa JR, Moore R, Larue L: A portrait of AKT kinases: human cancer and animal models depict a family with strong individualities. Cancer Biol Ther 2004, 3(3):268-275.