Basic & Clinical Medicine ›› 2010, Vol. 30 ›› Issue (8): 815-819.

• 研究论文 • Previous Articles     Next Articles

Construction of TK expression vector regulated by hypoxia and radiation and its radiosensitization effect on human breast cancer cells

Ling WEI, Xian-rang SONG, Xing-wu WANG, Ju-jie SUN, Bao SONG, Yan ZHENG   

  1. Cancer Research Center of Shandong Tumor Hospital Shandong Cancer Hospital
  • Received:2008-07-01 Revised:2009-02-19 Online:2010-08-05 Published:2010-08-05
  • Contact: Ling WEI,

Abstract: Objective To construct adenovirus vector Ad.HRE.CArG.HSV-TK regulated by hypoxia and radiation via AdEasy system and to observe its radiosensitization effect in human breast cancer cell lines Bcap37 and MDA-MB-435.Methods Using molecular clone technologies, Ad.HRE.CArG.HSV-TK inserted with HRE.CArG.HSV-TK fragment was constructed. The virus was purifed by CsCl gradient centrifuge. The functional titer of adenovirus was detected by GFP tag expression. Cell lines of Bcap37 and MDA-MB-435 were transducted with adenovirus in vitro, 48h later, expression rate of GFP was detected by flow cytometry.TK gene mRNA and protein expression were detected with real -time RT-PCR and Western blot respectively.The growth inhibition effect and sensitizer enhancement ratio( SER)of adenovirus(Ad)transduction combined with radiation(RT) were detected with MTT assay.Results The final titers of adenovirus was 2.1×109TU/ml. Bcap37 and MDA-MB-435 cells transducted by 50 MOI adenovirus in vitro for 48h, positive rate of GFP was 92% and 93%,respectively.TK expression increased dramatically(P<0.05). Growth inhibition rate in group of Ad+RT was significantly higher than that in Ad group and the same dose of RT group(P<0.05). SER were both 1.55.Conclusion The recombinant adenovirus expressing GFP and TK gene can be generated via AdEasy system.The adenovirus has radiosensitization effect on breast cancer cells. It may serve as a basis for later gene radiotherapy research in breast cancer.