Adenovirus-Mediated E2F-1 Gene Transfer Augments Gemcitabine-Induced Apoptosis in Human Colon Cancer Cells

Abstract

Background: E2F-1 is a transcription factor that stimulates cellular proliferation and cell cycle progression. E2F-1 alone is sufficient to stimulate cells to initiate DNA synthesis, and this unscheduled entry into S phase is a potent trigger of apoptosis. Gemcitabine, a novel pyrimidine analogue with structural and metabolic similarities to cytarabine, also can efficiently induce apoptosis, especially for cancer cells that are already in S phase. Gemcitabine has established antitumor activity against solid tumors, including head and neck, ovarian, and non-small cell lung cancers. Therefore, we hypothesized that exogenous E2F-1 expression could accumulate cells in the S phase and thus sensitize them to gemcitabine.

Methods: We constructed an adenoviral vector (AdCMVE2F-1) to transduce the exogenous E2F-1 gene into human cancer cells. Infection of human colon cancer cells with AdCMVE2F-1 resulted in the overexpression of E2F-1 mRNA and protein in a dose-dependent manner and consequently induced accumulation in S phase as measured by FACS analysis. To assess the synergistic antitumor effect of AdCMVE2F-1 and gemcitabine, the human colon cancer cel lines SW620, DLD-1, and LoVo were infected with AdCMVE2F-1 at various multiplicities of infection and then exposed to various concentrations of gemcitabine 24 hours after infection.

Result: Isobologram analysis showed that E2F-1-transduced cancer cells exhibited higher sensitivity to gemcitabine treatment compared to control virus-infected cells. Treatment with AdCMVE2F-1 plus gemcitabine enhanced endogenous p53 expression in LoVo cells, which contain wild-type p53; however, the finding that the synergistic effect can also be observed in mutant p53-expressing SW620 and DLD-1 cells suggests that wild-type p53 function may not be necessary for the therapeutic effects of this drug combination. Conclusions: Our data demonstrate that overexpression of ectopic E2F-1 protein may render cels more sensitive to gemcitabine-mediated apoptosis, an outcome that has important general implications for the treatment of human cancer.