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Adenovirus-mediated p14(ARF) gene transfer in human mesothelioma cells
1Thoracic Oncology Laboratory, University of California, San Francisco Cancer Center, CA 94115, USA.
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 G(1)-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 G(1)-phase arrest and apoptotic cell death. These results suggest that this gene therapy-based approach may be of use in the treatment of mesothelioma.
Insights
Gene therapy using p14(ARF) shows promise for mesothelioma treatment. Introducing p14(ARF) via an adenoviral vector inhibits mesothelioma cell growth and induces cell death, highlighting its therapeutic potential.
Area of Science:
- Oncology
- Molecular Biology
- Gene Therapy
Background:
- Mesothelioma often involves homozygous deletion of the INK4a/ARF locus, leading to p14(ARF) loss and p53 inactivation.
- p14(ARF) protein normally promotes p53 activity by degrading MDM2, preventing MDM2-mediated p53 inhibition.
Purpose of the Study:
- To evaluate the biological and therapeutic potential of p14(ARF) expression in mesothelioma cells.
- To investigate the role of p14(ARF) in regulating cell cycle and apoptosis in mesothelioma.
Main Methods:
- Constructed Adp14, an adenoviral vector carrying human p14(ARF) cDNA.
- Transfected human mesothelioma cell lines (H28, H513, H2052, MSTO-211H) with Adp14.
- Assessed cell growth, cell cycle progression, apoptosis, and cytotoxicity in response to Adp14 and control vectors.
Main Results:
- Adp14 transfection led to overexpression of p14(ARF), increased p53 and p21(WAF) protein levels, and retinoblastoma protein dephosphorylation.
- Mesothelioma cell growth was significantly inhibited by Adp14, inducing G1-phase cell cycle arrest and apoptotic cell death.
- Adp14-mediated cytotoxicity was significantly greater in p53-proficient colon cancer cells than in p53-null cells, indicating functional p53 is critical.
Conclusions:
- p14(ARF) overexpression in mesothelioma cells induces G1-phase arrest and apoptosis.
- Gene therapy with Adp14 represents a potential therapeutic strategy for mesothelioma treatment.