Growth suppression of human ovarian cancer cell lines by the introduction of a p16 gene via a recombinant adenovirus

Objective. The cell cycle regulatory protein p16 (CDKN2/cyclin dependent ki nase 4 inhibitor/multiple tumor suppressor-1) causes cell cycle arrest at t he G1 checkpoint by inhibiting activity of cyclin D-CDK4 complexes. The pur pose of this study is to assess the effect of introduction of the p16 gene into two ovarian cancer cell lines via a recombinant adenoviral vector (Ad5 CMV-p16). Methods. Cells lines used were SKOV3, which has a p16 deletion, and OVCA420 , which has normal p16, Transduction efficiency was established by infectin g cells with an adenovirus containing the Escherichia coli beta-galactosida se gene (Ad5CMV-beta-gal) at multiplicity of infection from 0 to 1000 and s taining for X-gal. Cells were infected with Ad5CMV-p16 and cell growth was assessed by counting cells every other day for up to 7 days. Western blotti ng was done to assess for p16 expression after infection. Fluorescence-acti vated cell sorting after staining with propidium iodide was done to assess the effect of p16 on the cell cycle. Results. The SKOV3 cell line was transduced with the adenovirus at a slight ly lower MOI than the OVCA420 cell line. Growth of the AdSCMV-p16-infected cells was suppressed 75-80% by cell count in both cell lines and caused mor phologic changes of the cells consistent with apoptosis. The p16 protein ex pression was seen to increase within 24 h after introduction of the p16 gen e. G1 arrest of cells occurred beginning 24 h after introduction of the p16 gene. Conclusions. These results suggest that Ad5CMV-p16 may be further studied a s a potential therapeutic agent for ovarian cancer as introduction of the p 16 gene into ovarian cancer cell lines causes a G1 arrest and attenuation o f growth, regardless of the endogenous p16 status of the cells, (C) 1999 Ac ademic Press.