Adenovirus-Mediated p14^ARF Gene Transfer in Human Mesothelioma Cells

Abstract

Background: The p14^ARF protein encoded by the INK4a/ARF locus promotes degradation of the MDM2 protein and thus prevents the MDM2-mediated inhibition of p53. Homozygous deletion of the INK4a/ARF locus is common in human mesothelioma and may result in the loss of p14^ARF and the inactivation of p53. We designed this study to evaluate the biologic and potential therapeutic roles of p14^ARF expression in mesothelioma cells. Methods and Results: We constructed Adp14, an adenoviral vector carrying human p14^ARF complementary DNA, and used it to transfect human mesothelioma cell lines H28, H513, H2052, and MSTO-211H. Overexpression of p14^ARF led to increased amounts of p53 and the p21^WAF proteins and dephosphorylation of the retinoblastoma protein. The growth rate of mesothelioma cells was inhibited markedly by infection with Adp14 compared with mock infection or infection with a control adenovirus vector, AdCtrl. Overexpression of p14^ARF induced G₁-phase cell cycle arrest and apoptotic cell death. Cytotoxicity assays showed that Adp14 had a statistically significantly (P = .002) greater effect on colon cancer (HCT116) cell lines containing two copies of the wild-type p53 gene than on p53-null cells, suggesting that functional p53 is a critical determinant of p14^ARF-mediated cytotoxicity. Conclusions: The transfection of p14^ARF into mesothelioma cells led to the overexpression of p14^ARF, which resulted in G₁-phase arrest and apoptotic cell death. These results suggest that this gene therapy-based approach may be of use in the treatment of mesothelioma.