Objective: To generate an adenoviral vector that expressed the canine p53 gene and investigate its growth-inhibiting effect on canine osteosarcoma and mammary adenocarcinoma cell lines.
Sample population: 2 canine osteosarcoma cell lines (HOS, OOS) and 3 canine mammary adenocarcinoma cell lines (CHMp, CIPm, and CNMm).
Procedure: An adenoviral vector that expressed the canine p53 gene (AxCA-cp53) was generated. p53 gene expression was examined by use of reverse transcription (RT)-polymerase chain reaction (PCR) assay and immunohistochemistry. Susceptibility of cell lines to the adenoviral vector was determined by infection with an adenoviral vector that expresses beta-galactosidase (AxCA-LacZ) and 3-indolyl-beta-D-galactopyranoside staining. Growth inhibitory effects were examined by monitoring the numbers of cells after infection with mock (PBS) solution, AxCA-LacZ, or AxCA-cp53. The DNA contents per cell were measured by flow cytometry analysis. Apoptotic DNA fragmentation was detected by use of a terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay.
Results: AxCA-cp53-derived p53 gene mRNA and P53 protein were detected by RT-PCR analysis and immunohistochemistry, respectively. Multiplicity of infection at which 50% of cells had positive 3-indolyl-beta-D-galactopyranoside staining results ranged from 10 to 50. AxCA-cp53 induced growth inhibition in a dose-dependent manner. Arrest of the G1-phase population and apoptotic DNA fragmentation were observed in cells infected with AxCA-cp53.
Conclusions and clinical relevance: AxCA-cp53 inhibits cell growth via induction of cell cycle arrest and apoptosis in canine osteosarcoma and mammary adenocarcinoma cell lines that lack a functional p53 gene. AxCA-cp53 may be useful to target the p53 gene in the treatment of dogs with tumors.