Additional gene therapy with Ad5CMV-p53 enhanced the efficacy of radiotherapy in human prostate cancer cells.
OBJECTIVE
The aim of this study was to investigate the efficacy of combination therapy of ionizing radiation (IR) and adenoviral p53 gene therapy and to evaluate its molecular mechanisms.


METHODS
Two human prostate cancer cell lines, DU145 and PC-3 cells, containing different types of p53 gene mutations, were investigated. The recombinant adenovirus vector containing the wild-type p53 gene (Ad5CMV-p53) was used for this study. Cells were irradiated (in 0, 2, 4, and 6 Gy, 300 cGy/min) and after 12 h of irradiation, the cells were infected with various doses of Ad5CMV-p53 (0-40 multiplicity of infection [MOI]). Cytotoxicity was determined by clonogenic assay. The molecular mechanisms were evaluated by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), apoptotic cell detection, and cell cycle analysis.


RESULTS
The cell growth inhibition in DU145 (p53-mutated) cells by IR was strongly enhanced by additional Ad5CMV-p53 infection in a viral dose-dependent manner. In DU145 cells, IR alone induced minimal p53 mRNA expression. However, IR combined with Ad5CMV-p53 infection stimulated significant increase in p53 mRNA expression supplemented with Bax and p21 mRNA expressions. In PC-3 (p53-null), IR induced Bax and p21 mRNA expression, while the combination effects were observed in p53, Bax, and p21 mRNA expression. Apoptotic cell deaths were rarely observed after IR alone (DU145: 3%, PC-3: 5%). However, after combination therapy, the proportion of apoptotic cells greatly increased (sevenfold in DU145 cells, and twice in PC-3 cells). G1 cell cycle arrest was observed after Ad5CMV-p53 infection and the combination in both cell lines.


CONCLUSIONS
In this study, we demonstrated that the combination of IR and Ad5CMV-p53 gene therapy resulted in remarkable synergistic effects in human prostate cancer cells. This combination therapy could be one of the optimal treatment strategies for radioresistant prostate cancer.
Sasaki
R
R
Department of Radiology, Kobe University School of Medicine, Hyogo, Kobe City, Japan.
Shirakawa
T
T
Zhang
Z J
ZJ
Tamekane
A
A
Matsumoto
A
A
Sugimura
K
K
Matsuo
M
M
Kamidono
S
S
Gotoh
A
A
D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis; D002453:Cell Cycle; D003131:Combined Modality Therapy; D016158:Genes, p53; D015316:Genetic Therapy; D006801:Humans; D008297:Male; D011471:Prostatic Neoplasms / Q000473:pathology / Q000628:therapy*; D011836:Radiation Tolerance; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured