Construction of Recombinant Adenovirus Containing tk Gene_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

CHEN Gengzhen , HAN Hui , XU Mingyan , DENG Xiaoling.

The Third Department of Surgery, The Second Affiliated Hospital, Shantou University Medical College, Shantou 515041, Guangdong Province, China;

Corresponding author：

Keywords：

Gene therapy; Suicide gene; Thymidine kinase; Adenovirus vector

Video：

Export PDF Favorites Scan Get Citation

Abstract Full text Figures/Tables Video References Cited by

Objective To construct replication-defective adenovirus containing tk gene (ADV-tk).
Methods Recombinant adenovirus of ADV-tk was constructed using homologous recombination in cells. After the interested tk gene fragment in the recombinant plasmid obtained was confirmed by PCR, the titre of purified recombinant adenovirus was detected. In vitro study, tk gene in SMMC7721 cells transfected by ADV-tk was investigated by RT-PCR. In vivo study, ADV-tk was injected intraperitoneally into BALB/c nude mice with liver cancer and apoptosis cells in tumor were observed.
Results Recombinant adenovirus containing ADV-tk was proved successfully. The titre of purified recombinant adenovirus was 1.4×1010 pfu/ml. In vitro study, tk was integrated and expressed by SMMC-7721 cells. In vivo study, with the injection of ADV-tk, apoptosis cells in tumor increased.
Conclusion A replication-defective adenovirus containing tk gene is successfully constructed, which may useful for further research on tumor suicide gene therapy with ADV-tk.

Citation： CHEN Gengzhen,HAN Hui,XU Mingyan,DENG Xiaoling.. Construction of Recombinant Adenovirus Containing tk Gene. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2010, 17(7): 703-706. doi: Copy

1.	Vassaux G, MartinDuque P. Use of suicide genes for cancer gene therapy: study of the different approaches [J]. Expert Opin Biol Ther, 2004; 4(4): 519530.
2.	Portsmouth D, Hlavaty J, Renner M. Suicide genes for cancer therapy [J]. Mol Aspects Med, 2007; 28(1): 441.
3.	Altaner C. Prodrug cancer gene therapy [J]. Cancer Lett, 2008; 270(2): 191201.
4.	Si S, Sun Y, Li Z, et al. Gene therapy by membraneexpressed superantigen for αfetoproteinproducing hepatocellular carcinoma [J]. Gene Ther, 2006; 13(22): 16031610.
5.	Stefani AL, Barzon L, Castagliuolo I, et al. Systemic efficacy of combined suicide/cytokine gene therapy in a murine model of hepatocellular carcinoma [J]. Hepatol, 2005; 42(5): 728735.
6.	Song TJ, Eisenberg DP, Adusumilli PS, et al. Oncolytic herpes viral therapy is effective in the treatment of hepatocellular carcinoma cell lines [J]. J Gastrointest Surg, 2006; 10(4): 532542.
7.	Cai X, Zhou J, Chang Y, et al. Targeting gene therapy for hepatocarcinoma cells with the E.coli purine nucleoside phosphorylase suicide gene system directed by a chimeric αfetoprotein promoter [J]. Cancer Lett, 2008; 264(1): 7182.
8.	Li YM, Song ST, Jiang ZF, et al. Telomerasespecific oncolytic virotherapy for human hepatocellular carcinoma [J]. World J Gastroenterol, 2008; 14(8): 12741279.
9.	Zhou J, Gao Q, Chen G, et al. Novel oncolytic adenovirus selectively targets tumorassociated pololike kinase 1 and tumor cell viability [J]. Clin Cancer Res, 2005; 11(23): 84318440.
10.	Majumdar AS, Hughes DE, Lichtsteiner SP, et al. The telomerase reverse transcriptase promoter drives efficacious tumor suicide gene therapy while preventing hepatotoxicity encountered with constitutive promoters [J]. Gene Ther, 2001; 8(7): 568578.
11.	Aguilar LK, The BS, Vaachaki MT, et al. AdVtk/valacyclovir gene therapy in combination with radiotherapy for prostate cancer: interim results of a phase Ⅰ/Ⅱ clinical trial [C]. Proc Am Soc Clin Oncol, 21: 2002 (abstr 25).
12.	Ling CQ, Li B, Zhang C, et al. Inhibitory effect of recombinant adenovirus carrying melittin gene on hepatocellular carcinoma [J]. Ann Oncol, 2005; 16(1): 109115.
13.	Gentry BG, Boucher PD, Shewach DS. Hydroxyurea induces bystander cytotoxicity in cocultures of herpes simplex virus thymidine kinaseexpressing and nonexpressing HeLa cells incubated with ganciclovir [J]. Cancer Res, 2006; 66(7): 38453851.
14.	Li N, Zhou J, Weng D, et al. Adjuvant adenovirusmediated delivery of herpes simplex virus thymidine kinase administration improves outcome of liver transplantation in patients with advanced hepatocellular carcinoma [J]. Clin Cancer Res, 2007; 13(19): 58475854.
15.	Palmer DH, Mautner V, Mirza D, et al. Virusdirected enzyme prodrug therapy: intratumoral administration of a replicationdeficient adenovirus encoding nitroreductase to patients with resectable liver cancer [J]. J Clin Oncol, 2004; 22(9): 15461552.
16.	van Dillen IJ, Mulder NH, Vaalburg W, et al. Influence of the bystander effect on HSVtk/GCV gene therapy. A review [J]. Curr Gene Ther, 2002; 2(3): 307322.
17.	Unger MM, Wahl J, Ushmorov A, et al. Enriching suicide gene bearing tumor cells for an increased bystander effect [J]. Cancer Gene Ther, 2007; 14(1): 3038.
18.	Zhang Q, Chen G, Peng L, et al. Increased safety with preserved antitumoral efficacy on hepatocellular carcinoma with dualregulated oncolytic adenovirus [J]. Clin Cancer Res, 2006; 12(21): 65236531.
19.	Hsieh JL, Lee CH, Teo ML, et al. Transthyretindriven oncolytic adenovirus suppresses tumor growth in orthotopic and ascites models of hepatocellular carcinoma [J]. Cancer Sci, 2009; 100(3): 537545.
20.	Li G, Sham J, Yang J, et al. Potent antitumor efficacy of an E1B 55kDadeficient adenovirus carrying murine endostatin in hepatocellular carcinoma [J]. Int J Cancer, 2005; 113(4): 640648.
21.	He LF, Gu JF, Tang WH, et al. Significant antitumor activity of oncolytic adenovirus expressing human interferonβ for hepatocellular carcinoma [J]. J Gene Med, 2008; 10(9): 983992.
22.	Grimm D, Kay MA. From virus evolution to vector revolution: use of naturally occurring serotypes of adenoassociated virus (AAV) as novel vectors for human gene therapy [J]. Curr Gene Ther, 2003; 3(4): 281304.
23.	Coura Rdos S, Nardi NB. The state of the art of adenoassociated virusbased vectors in gene therapy [J]. Virol J, 2007; 4: 99105.
24.	Liu H, Peng CH, Liu YB, et al. Inhibitory effect of adenoassociated virusmediated gene transfer of human endostatin on hepatocellular carcinoma [J]. World J Gastroenterol, 2005; 11(22): 33313334.
25.	Blechacz B, Splinter PL, Greiner S, et al. Engineered measles virus as a novel oncolytic viral therapy system for hepatocellular carcinoma [J]. Hepatology, 2006; 44(6): 14651477.
26.	刘自明, 严律南, 李馨筱. 包载携有HSVTK基因腺病毒PLG微球的制备 [J]. 中国普外基础与临床杂志, 2007; 14(4): 399402.
27.	罗涛, 张强, 张建, 等. 腺病毒介导胸苷激酶基因对静脉平滑肌细胞增殖及内膜增生抑制作用的研究 [J]. 中国普外基础与临床杂志, 2006; 13(3): 291294.
28.	李梅生, 梁力建, 黄洁夫, 等. HSVtk/CD融合基因杀伤人胆管癌细胞QBC939的体内实验 [J]. 中国普外基础与临床杂志, 2005; 12(6): 581584.

1. Vassaux G, MartinDuque P. Use of suicide genes for cancer gene therapy: study of the different approaches [J]. Expert Opin Biol Ther, 2004; 4(4): 519530.
2. Portsmouth D, Hlavaty J, Renner M. Suicide genes for cancer therapy [J]. Mol Aspects Med, 2007; 28(1): 441.
3. Altaner C. Prodrug cancer gene therapy [J]. Cancer Lett, 2008; 270(2): 191201.
4. Si S, Sun Y, Li Z, et al. Gene therapy by membraneexpressed superantigen for αfetoproteinproducing hepatocellular carcinoma [J]. Gene Ther, 2006; 13(22): 16031610.
5. Stefani AL, Barzon L, Castagliuolo I, et al. Systemic efficacy of combined suicide/cytokine gene therapy in a murine model of hepatocellular carcinoma [J]. Hepatol, 2005; 42(5): 728735.
6. Song TJ, Eisenberg DP, Adusumilli PS, et al. Oncolytic herpes viral therapy is effective in the treatment of hepatocellular carcinoma cell lines [J]. J Gastrointest Surg, 2006; 10(4): 532542.
7. Cai X, Zhou J, Chang Y, et al. Targeting gene therapy for hepatocarcinoma cells with the E.coli purine nucleoside phosphorylase suicide gene system directed by a chimeric αfetoprotein promoter [J]. Cancer Lett, 2008; 264(1): 7182.
8. Li YM, Song ST, Jiang ZF, et al. Telomerasespecific oncolytic virotherapy for human hepatocellular carcinoma [J]. World J Gastroenterol, 2008; 14(8): 12741279.
9. Zhou J, Gao Q, Chen G, et al. Novel oncolytic adenovirus selectively targets tumorassociated pololike kinase 1 and tumor cell viability [J]. Clin Cancer Res, 2005; 11(23): 84318440.
10. Majumdar AS, Hughes DE, Lichtsteiner SP, et al. The telomerase reverse transcriptase promoter drives efficacious tumor suicide gene therapy while preventing hepatotoxicity encountered with constitutive promoters [J]. Gene Ther, 2001; 8(7): 568578.
11. Aguilar LK, The BS, Vaachaki MT, et al. AdVtk/valacyclovir gene therapy in combination with radiotherapy for prostate cancer: interim results of a phase Ⅰ/Ⅱ clinical trial [C]. Proc Am Soc Clin Oncol, 21: 2002 (abstr 25).
12. Ling CQ, Li B, Zhang C, et al. Inhibitory effect of recombinant adenovirus carrying melittin gene on hepatocellular carcinoma [J]. Ann Oncol, 2005; 16(1): 109115.
13. Gentry BG, Boucher PD, Shewach DS. Hydroxyurea induces bystander cytotoxicity in cocultures of herpes simplex virus thymidine kinaseexpressing and nonexpressing HeLa cells incubated with ganciclovir [J]. Cancer Res, 2006; 66(7): 38453851.
14. Li N, Zhou J, Weng D, et al. Adjuvant adenovirusmediated delivery of herpes simplex virus thymidine kinase administration improves outcome of liver transplantation in patients with advanced hepatocellular carcinoma [J]. Clin Cancer Res, 2007; 13(19): 58475854.
15. Palmer DH, Mautner V, Mirza D, et al. Virusdirected enzyme prodrug therapy: intratumoral administration of a replicationdeficient adenovirus encoding nitroreductase to patients with resectable liver cancer [J]. J Clin Oncol, 2004; 22(9): 15461552.
16. van Dillen IJ, Mulder NH, Vaalburg W, et al. Influence of the bystander effect on HSVtk/GCV gene therapy. A review [J]. Curr Gene Ther, 2002; 2(3): 307322.
17. Unger MM, Wahl J, Ushmorov A, et al. Enriching suicide gene bearing tumor cells for an increased bystander effect [J]. Cancer Gene Ther, 2007; 14(1): 3038.
18. Zhang Q, Chen G, Peng L, et al. Increased safety with preserved antitumoral efficacy on hepatocellular carcinoma with dualregulated oncolytic adenovirus [J]. Clin Cancer Res, 2006; 12(21): 65236531.
19. Hsieh JL, Lee CH, Teo ML, et al. Transthyretindriven oncolytic adenovirus suppresses tumor growth in orthotopic and ascites models of hepatocellular carcinoma [J]. Cancer Sci, 2009; 100(3): 537545.
20. Li G, Sham J, Yang J, et al. Potent antitumor efficacy of an E1B 55kDadeficient adenovirus carrying murine endostatin in hepatocellular carcinoma [J]. Int J Cancer, 2005; 113(4): 640648.
21. He LF, Gu JF, Tang WH, et al. Significant antitumor activity of oncolytic adenovirus expressing human interferonβ for hepatocellular carcinoma [J]. J Gene Med, 2008; 10(9): 983992.
22. Grimm D, Kay MA. From virus evolution to vector revolution: use of naturally occurring serotypes of adenoassociated virus (AAV) as novel vectors for human gene therapy [J]. Curr Gene Ther, 2003; 3(4): 281304.
23. Coura Rdos S, Nardi NB. The state of the art of adenoassociated virusbased vectors in gene therapy [J]. Virol J, 2007; 4: 99105.
24. Liu H, Peng CH, Liu YB, et al. Inhibitory effect of adenoassociated virusmediated gene transfer of human endostatin on hepatocellular carcinoma [J]. World J Gastroenterol, 2005; 11(22): 33313334.
25. Blechacz B, Splinter PL, Greiner S, et al. Engineered measles virus as a novel oncolytic viral therapy system for hepatocellular carcinoma [J]. Hepatology, 2006; 44(6): 14651477.
26. 刘自明, 严律南, 李馨筱. 包载携有HSVTK基因腺病毒PLG微球的制备 [J]. 中国普外基础与临床杂志, 2007; 14(4): 399402.
27. 罗涛, 张强, 张建, 等. 腺病毒介导胸苷激酶基因对静脉平滑肌细胞增殖及内膜增生抑制作用的研究 [J]. 中国普外基础与临床杂志, 2006; 13(3): 291294.
28. 李梅生, 梁力建, 黄洁夫, 等. HSVtk/CD融合基因杀伤人胆管癌细胞QBC939的体内实验 [J]. 中国普外基础与临床杂志, 2005; 12(6): 581584.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Construction of Recombinant Adenovirus Containing tk Gene

Abstract Full text Figures/Tables Video References Cited by

Format

Content