Adenovirus-mediated E2F-1 gene transfer efficiently induces apoptosis in melanoma cells

BACKGROUND, E2F-1 is a transcription factor that stimulates cellular prolif eration and cell cycle progression from G(1) to S-phase. Somewhat paradoxic ally, E2F-1 also has the properties of a tumor suppressor. Overexpression o f E2F-1 has been shown to induce apoptosis in some cancer cells. In the cur rent study, the effect of adenovirus-mediated E2F-1 gene transfer on human melanoma cell growth was investigated. METHODS. Two human melanoma cell lines, SK-MEL-28 (wild-type p53) and SK-ME L-2 (mutant p53), were treated by mock infection, infection with a control vector expressing the beta-galactosidase gene (Ad5CMV-LacZ), or infection w ith a vector expressing E2F-1 (Ad5CMV-E2F-1) at a multiplicity of infection of 100. Cell proliferation and viability were determined by WST-1 assay an d trypan blue exclusion, respectively. Apoptosis was assessed by cell flow cytometry and confirmed by cell morphology, in situ terminal deoxynucleotid yl nick end labeling assay, and poly(ADP-ribose) polymerase cleavage assay. RESULTS. Marked overexpression of E2F-1 was evident in both cell lines 24 h ours after infection with Ad5CMVE2F-1 by Western blot analysis. E2F-1 overe xpression resulted in growth inhibition and rapid loss of cell viability. O verexpression of E2F-1 also resulted in premature S-phase entry and G(2) ar rest at 24 hours followed by apoptotic cell death at 48 hours. After Ad5CMV E2F-1 infection, expression of Bar and Bak was unchanged, whereas Mcl-1 lev els decreased markedly. In SK-MEL-28 cells, Bcl-XL levels also declined aft er E2F-1 expression. Bcl-2 was undetectable in SK-MEL-28 cells but was incr eased in SK-MEL-2 cells in response to E2F-1 overexpression. CONCLUSIONS. Adenovirus-mediated E2F-1 gene transfer efficiently induces wi despread apoptosis in human melanoma cells. E2F-1 overexpression induced ap optosis in cell lines containing wild-type and mutant p53, suggesting that this effect does not require wild-type p53 function. Anti-apoptotic protein s of the Bcl-2 family, notably Mcl-1 and Bcl-XL, may be involved in mediati ng the response to E2F-1. These data suggest that adenovirus-mediated E2F-1 gene therapy may be effective in the treatment of melanoma. Cancer 1999;86 :2021-33. (C) 1999 American Cancer Society.