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Adenovirus‐mediated E2F‐1 gene transfer efficiently induces apoptosis in melanoma cells
Author(s) -
Dong YanBin,
Yang HaiLiang,
Elliott Mary Jane,
Liu TaJen,
Stilwell Ariel,
Atienza Cesar,
McMasters Kelly M.
Publication year - 1999
Publication title -
cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.052
H-Index - 304
eISSN - 1097-0142
pISSN - 0008-543X
DOI - 10.1002/(sici)1097-0142(19991115)86:10<2021::aid-cncr20>3.0.co;2-1
Subject(s) - gene transfer , medicine , apoptosis , melanoma , cancer research , e2f , gene , genetic enhancement , microbiology and biotechnology , genetics , cancer , cell cycle , biology
BACKGROUND E2F‐1 is a transcription factor that stimulates cellular proliferation and cell cycle progression from G 1 to S‐phase. Somewhat paradoxically, E2F‐1 also has the properties of a tumor suppressor. Overexpression of E2F‐1 has been shown to induce apoptosis in some cancer cells. In the current 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‐MEL‐2 (mutant p53), were treated by mock infection, infection with a control vector expressing the β‐galactosidase gene (Ad5CMV‐LacZ), or infection with 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 and trypan blue exclusion, respectively. Apoptosis was assessed by cell flow cytometry and confirmed by cell morphology, in situ terminal deoxynucleotidyl nick end labeling assay, and poly(ADP‐ribose) polymerase cleavage assay. RESULTS Marked overexpression of E2F‐1 was evident in both cell lines 24 hours after infection with Ad5CMVE2F‐1 by Western blot analysis. E2F‐1 overexpression resulted in growth inhibition and rapid loss of cell viability. Overexpression of E2F‐1 also resulted in premature S‐phase entry and G 2 arrest at 24 hours followed by apoptotic cell death at 48 hours. After Ad5CMVE2F‐1 infection, expression of Bax and Bak was unchanged, whereas Mcl‐1 levels decreased markedly. In SK‐MEL‐28 cells, Bcl‐XL levels also declined after E2F‐1 expression. Bcl‐2 was undetectable in SK‐MEL‐28 cells but was increased in SK‐MEL‐2 cells in response to E2F‐1 overexpression. CONCLUSIONS Adenovirus‐mediated E2F‐1 gene transfer efficiently induces widespread apoptosis in human melanoma cells. E2F‐1 overexpression induced apoptosis in cell lines containing wild‐type and mutant p53, suggesting that this effect does not require wild‐type p53 function. Anti‐apoptotic proteins of the Bcl‐2 family, notably Mcl‐1 and Bcl‐XL, may be involved in mediating 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. © 1999 American Cancer Society.