Cell-specific expression of the diphtheria toxin A-chain coding sequence under the control of the upstream region of the human alpha-fetoprotein gene

✍ Scribed by Murayama, Yoshihiko; Tadakuma, Takushi; Kunitomi, Michito; Kumai, Koichiro; Tsutsui, Ken; Yasuda, Tatsuji; Kitajima, Masaki

• Publisher: John Wiley and Sons
• Year: 1999
• Tongue: English
• Weight: 297 KB
• Volume: 70
• Category: Article
• ISSN: 0022-4790
• DOI: 10.1002/(sici)1096-9098(199903)70:3<145::aid-jso1>3.0.co;2-o

✦ Synopsis

Background and Objectives: Development of the system to express a suicide gene selectively in tumor cells is essential for gene therapy. We constructed a plasmid containing the diphtheria toxin A (DTA) fragment linked to human alpha-fetoprotein (AFP) promoter and enhancer, and tested whether it can exert its cytocidal effect selectively on AFPproducing cells. Methods: The chloramphenical acetyltransferase (CAT) reporter gene or DTA gene was linked to the 5Ј upstream region of the AFP gene. The plasmids were transfected into AFP-producing or non-producing cells by the lipopolyamine-coated DNA method. Expression of CAT activity and effects on cell growth of transfected cells were assessed. Results: When the AFP-producing cells HuH-7 or HepG2 were cotransfected with CAT reporter plasmid and pAF5.1DTA plasmid, the CAT activity was greatly suppressed. In contrast, cotransfection with pAF5.1DTA-R, the inversely inserted DTA gene, did not inhibit CAT activity. Furthermore, cell growth of HuH-7 cells transfected with pAF5.1DTA plasmid was significantly inhibited compared with HuH-7 cells transfected with DTA-R plasmid. Conclusions: Our results indicate that selective killing of AFP-producing cells will be attained by introducing the DTA gene linked to the promoter and enhancer region of AFP.