Abstract 789: Adenovirus p53 enhances the antitumor effect of adenovirus thymidine kinase/ganciclovir on Hep3B cells by apoptosis.

Abstract

Abstract Background and Aims: Although Thymidine Kinase/Ganciclovir(TK/GCV) is a promising tumor-targeted chemotherapy by DNA damage, it is reported that tumor cells having wild-type p53 are more sensitive to chemotherapeutic agents. Furthermore some evidence have suggested approximately half of all cancers possess inactivated p53. Numerous studies have shown the key role of p53 as a tumor suppressor is blocking cell cycle progression and/or inducing apoptosis in response to high level of DNA damage. Given that TK/GCV results in DNA damage, exogenous wild-type p53 should enhance efficacy of adenovirus (adv)-mediated antitumor effect of TK/GCV on p53 −/- tumor cells. Thus, this article aims to determine whether p53 −/- cancer cells are sensitive at TK/GCV, whether coexpression of p53 and TK/GCV can actively kill p53−/- tumor cells and the main mechanisms responsible for TK/GCV tumor cell death. Methods:Hep3B cells (p53−/−) and HepG2(p53+/+) cells were infected with adv-p53 and adv-TK one day prior to GCV. The cellular growth inhibition was analyzed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and immunofluorescent staining (PI and Calcein AM). Apoptosis and cell cycle arrest were measured using annexin V-FITC /PI double staining and PI by flow cytometry. p53,PUMA and p21 mRNA expression were analyzed using real-time quantitative PCR . The protein levels of p53,PUMA and p21 were detected by Western blotting. Results:The MTT results showed that cell viability (%) of HepG2 cell (p53+/+) treated with adv-TK(107/ml)/GCV(10ug/ml) were lower than Hep3B (p53−/−)(p<0.01). Growth inhibition of Hep3B treated with adv-p53 in combination with adv-TK/GCV was higher than adv-p53 and adv-TK/GCV alone (p<0.01). Annexin V-FITC/PI double staining and propidium iodide (PI) staining demonstrates that Hep3B cell treated with adv-p53 and adv-TK/GCV had significantly increased apoptosis compared to adv-p53 (p<0.01) and adv-TK/GCV(p<0.05) alone. The expression level of p53,p21 and PUMA mRNA and protein was significantly higher in Hep3B cell treated with adv-p53 and adv-TK/GCV compared to adv-p53 and adv-TK/GCV alone in a time-dependent manner. Conclusions:These results suggest that the p53−/- cancer cell is not sensitive to TK/GCV. p53 −/- tumor cells were actively killed by a combination adv- p53 with TK/GCV. p53-mediated apoptosis is the main mechanisms responsible for TK/GCV kill tumor cells. Citation Format: Kai Liu, Xiuhong Liu, Tao Wen, Feng Ren, Jiming Yin, Daojie Liu, Huiguo Ding, Ning Li, Dexi Chen. Adenovirus p53 enhances the antitumor effect of adenovirus thymidine kinase/ganciclovir on Hep3B cells by apoptosis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 789. doi:10.1158/1538-7445.AM2013-789 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.