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Nitric Oxide Synthase Type III Overexpression By Gene Therapy Exerts Antitumoral Activity In Mouse Hepatocellular
Raúl González1, Ángel J De la Rosa2, Santiago Romero-Brufau3
1Departament of Biochemistry and Molecular Biology, Universidad de Córdoba, Instituto Maimónides de Investigación Biomédica de Córdoba (IMIBIC), Spain.
Abstract:
Hepatocellular carcinoma develops in cirrhotic liver. The nitric oxide (NO) synthase type III (NOS-3) overexpression induces cell death in hepatoma cells. The study developed gene therapy designed to specifically overexpress NOS-3 in cultured hepatoma cells, and in tumors derived from orthotopically implanted tumor cells in fibrotic livers. Liver fibrosis was induced by CCl4 administration in mice. Hepa 1-6 cells were used for in vitro and in vivo experiments. The first generation adenovirus was designed to overexpress NOS-3 (or GFP) and luciferase cDNA under the regulation of murine alpha-fetoprotein (AFP) and Rous Sarcoma Virus (RSV) promoters, respectively. Both adenoviruses were administered through the tail vein two weeks after orthotopic tumor cell implantation. AFP-NOS-3/RSV-Luciferase increased oxidative-related DNA damage, p53, CD95/CD95L expression and caspase-8 activity in cultured Hepa 1-6 cells. The increased expression of CD95/CD95L and caspase-8 activity was abolished by l-NAME or p53 siRNA. The tail vein infusion of AFP-NOS- 3/RSV-Luciferase adenovirus increased cell death markers, and reduced cell proliferation of established tumors in fibrotic livers. The increase of oxidative/nitrosative stress induced by NOS-3 overexpression induced DNA damage, p53, CD95/CD95L expression and cell death in hepatocellular carcinoma cells. The effectiveness of the gene therapy has been demonstrated in vitro and in vivo.
Insights
Gene therapy overexpressing nitric oxide synthase type III (NOS-3) effectively induces cell death in hepatocellular carcinoma cells. This approach shows promise for treating liver cancer in fibrotic livers by increasing oxidative stress and DNA damage.
Area of Science:
- Hepatocellular Carcinoma Research
- Gene Therapy
- Molecular Oncology
Background:
- Hepatocellular carcinoma (HCC) commonly arises in cirrhotic livers.
- Overexpression of nitric oxide synthase type III (NOS-3) induces apoptosis in hepatoma cells.
Purpose of the Study:
- To develop and evaluate a gene therapy strategy for specifically overexpressing NOS-3 in HCC.
- To assess the efficacy of this gene therapy in vitro and in vivo models of HCC in fibrotic livers.
Main Methods:
- Development of a first-generation adenovirus vector (AFP-NOS-3/RSV-Luciferase) for NOS-3 overexpression.
- In vitro studies using Hepa 1-6 hepatoma cells.
- In vivo studies involving orthotopic implantation of Hepa 1-6 cells in CCl4-induced fibrotic mouse livers, followed by tail vein adenovirus administration.
Main Results:
- AFP-NOS-3/RSV-Luciferase enhanced oxidative DNA damage, p53, CD95/CD95L expression, and caspase-8 activity in cultured hepatoma cells.
- Adenovirus-mediated NOS-3 overexpression reduced tumor cell proliferation and increased cell death markers in established HCC tumors within fibrotic livers.
- The observed effects were linked to increased oxidative/nitrosative stress, DNA damage, and apoptosis-related protein expression.
Conclusions:
- Adenovirus-mediated gene therapy targeting NOS-3 overexpression is effective in inducing cell death in HCC.
- This gene therapy strategy demonstrates potential for treating HCC in the context of liver fibrosis.
- The mechanism involves enhancing oxidative stress, DNA damage, and apoptosis pathways within cancer cells.
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