Objective To construct a mammalian vector encoding angiostatin kringle 5 (K5) under the control of αfetoprotein （AFP） enhancer and albumin promoter, and to observe the expression of angiostatin by introducting angiostatin gene into hepatocellular carcinoma cells through gene transfection.
Methods Angiostatin cDNA was amplified from normal human eukaryotic cells by using RTPCR. Meanwhile, AFP enhancer and albumin promoter sequences were directed cloned and were inserted into vector pcDNA3.1. The recombinant vector of pcDNA3.1AFABangiostatin K5His was constructed, which contained the angiostatin K5 cDNA sequence that was under the control of the AFP enhancer and promoter. Angiostatin K5 cDNA was introduced into human AFP positive hepatocellular carcinoma cell lines with the transfected cultured cells that were mediated with Lipofectamine 2000. The expression of angiostatin K5 was analyzed by Western blot and the protein was dectected with antiHis antibody.
Results The 500base pair of angiostatin K5 was in accordance with the expected sequence and the recombinant vector of pcDNA3.1AFABangiostatin K5His was also confirmed as the anticipated sequence. The expression of angiostatin K5 in AFP positive hepatocellular carcinoma cells was detected both by SDSPAGE and Western blot.
Conclusion Efficient construction and expression of angiostatin K5 to AFP positive cells make it possible for antiangiogenesis therapy of human hepatocellular carcinomas, which may provide a promising approach.
Citation：
XU Hongyong,XU Li,GAO Jianhong,LI Kaizong,DOU Kefeng. Construction and Expression of Hepatocellular CarcinomaSpecific Expressing Eukaryotic Vector for AntiAngiogenesis Therapy. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2007, 14(1): 15-18. doi:
Copy
Copyright © the editorial department of CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY of West China Medical Publisher. All rights reserved
| 1. |
Farinati F, Marino D, De Giorgio M, et al. Diagnostic and prognostic role of alphafetoprotein in hepatocellular carcinoma: both or neither? [J]. Am J Gastroenterol, 2006; 101(3)∶524.
|
| 2. |
Perri SR, Nalbantoglu J, Annabi B, et al. Plasminogen kringle 5engineered glioma cells block migration of tumorassociated macrophages and suppress tumor vascularization and progression [J]. Cancer Res, 2005; 65(18)∶8359.
|
| 3. |
Kong N, Lim D, Lee K. Interacting partners for kringle domains of plasminogen: common binding with K1 and K5 domains [J]. Protein Pept Lett, 2004; 11(6)∶521.
|
| 4. |
Su H, Lu R, Chang JC, et al. Tissuespecific expression of herpes simplex virus thymidine kinase gene delivered by adenoassociated virus inhibits the growth of human hepatocellular carcinoma in athymic mice [J]. Proc Natl Acad Sci USA, 1997; 94(25)∶13891.
|
| 5. |
Cao YH, Andrew Chen, Seong SA, et al. Kringle5 of plasminogen is a novel inhibitor of endothelial cell growth [J]. J Biol Chem, 1997; 272(36)∶22924.
|
| 6. |
Su H, Chang JC, Xu SM, et al. Selective killing of AFPpositive hepatocellular carcinoma cells by adenoassociated virus transfer of the herpes simplex virus thymidine kinase gene [J]. Hum Gene Ther, 1996; 7(4)∶463.
|
| 7. |
Berent R, Auer J, Porodko M, et al. Influence of cardiopulmonary resuscitation on levels of tumour markers [J]. Eur J Cancer Care (Engl), 2006; 15(3)∶252.
|
- 1. Farinati F, Marino D, De Giorgio M, et al. Diagnostic and prognostic role of alphafetoprotein in hepatocellular carcinoma: both or neither? [J]. Am J Gastroenterol, 2006; 101(3)∶524.
- 2. Perri SR, Nalbantoglu J, Annabi B, et al. Plasminogen kringle 5engineered glioma cells block migration of tumorassociated macrophages and suppress tumor vascularization and progression [J]. Cancer Res, 2005; 65(18)∶8359.
- 3. Kong N, Lim D, Lee K. Interacting partners for kringle domains of plasminogen: common binding with K1 and K5 domains [J]. Protein Pept Lett, 2004; 11(6)∶521.
- 4. Su H, Lu R, Chang JC, et al. Tissuespecific expression of herpes simplex virus thymidine kinase gene delivered by adenoassociated virus inhibits the growth of human hepatocellular carcinoma in athymic mice [J]. Proc Natl Acad Sci USA, 1997; 94(25)∶13891.
- 5. Cao YH, Andrew Chen, Seong SA, et al. Kringle5 of plasminogen is a novel inhibitor of endothelial cell growth [J]. J Biol Chem, 1997; 272(36)∶22924.
- 6. Su H, Chang JC, Xu SM, et al. Selective killing of AFPpositive hepatocellular carcinoma cells by adenoassociated virus transfer of the herpes simplex virus thymidine kinase gene [J]. Hum Gene Ther, 1996; 7(4)∶463.
- 7. Berent R, Auer J, Porodko M, et al. Influence of cardiopulmonary resuscitation on levels of tumour markers [J]. Eur J Cancer Care (Engl), 2006; 15(3)∶252.
