Adenovirus-mediated suicide-gene therapy using the herpes simplex virus thymidine kinase gene in cell and animal models of human prostate cancer: changes in tumour cell proliferative activity.
OBJECTIVE
To determine the feasibility and efficacy of suicide-gene therapy using adenovirus (Ad)-mediated herpes simplex virus thymidine kinase (HSV-TK) and the prodrug acyclovir, and to evaluate changes in the biological phenotype for tumour cell proliferative activity after suicide-gene therapy in animal models of human prostate cancer.


METHODS
Using a replication-defective adenoviral vector (cytomegalovirus, CMV) containing the beta-galactosidase gene (Ad-CMV-beta-gal) as a control and Ad-CMV-TK as the therapeutic vector under the transcriptional control of the CMV promoter, transduction efficiency was assessed in vitro by infecting LNCaP and PC-3 androgen-dependent and independent human prostate cancer cells with Ad-CMV-beta-gal, and using X-gal staining. The TK activity in prostate cancer cells infected with Ad-CMV-TK was determined by measuring TK-mediated [3H]-gancyclovir phosphorylation. The sensitivity of LNCaP and PC-3 cells to Ad-CMV-TK in vitro was determined after infection with the therapeutic vector with or without acyclovir. The inhibition of PC-3 tumour growth in vivo induced by the Ad-CMV-TK/acyclovir suicide-gene system was assessed in separate and controlled experiments using human prostate cancer mouse models. Ki-67 proliferative antigen and proliferating cell nuclear antigen (PCNA), both useful proliferative indices, were evaluated using immunohistochemical staining (MIB-1 monoclonal antibody and monoclonal anti-PCNA antibody) in formalin-fixed, paraffin-embedded tissues from gene therapy-treated and control animals.


RESULTS
The mean TK activity was significantly higher in LNCaP and PC-3 cells infected with Ad-CMV-TK than in cells infected with Ad-CMV-beta-gal, used as a control (P < 0.05). The growth of human prostate cancer cells with Ad-CMV-TK was significantly inhibited by adding acyclovir in vitro (P < 0.05). In the in vivo experiments using the PC-3 human prostate cancer mouse model, tumour volume and growth was lower in mice treated with Ad-CMV-TK/acyclovir than in those treated with Ad-CMV-TK only, acyclovir only or untreated (controls) (P < 0.05). Histochemical staining of tumour tissues showed that Ad-CMV-TK/acyclovir destroyed PC-3 tumours through tumour cell death and apoptosis, with local lymphatic infiltration. The mean PCNA labelling index in prostate cancer cells of mice treated with Ad-CMV-TK/acyclovir was significantly lower than that in untreated controls (P < 0.05, Mann-Whitney U-test). The Ki-67 labelling index in prostate cancer cells of mice treated with Ad-CMV-TK/acyclovir was also lower than that in untreated controls (P < 0.05, Student's t-test). Adenovirus-mediated suicide-gene therapy using the HSV-TK gene decreased the proliferative activity of PC-3 human prostatic cancer cells in vivo.


CONCLUSIONS
Adenovirus-mediated suicide-gene therapy using an HSV-TK/acyclovir system provided effective therapy in an experimental human prostate cancer mouse model, by significantly inhibiting tumour growth and decreasing the proliferative activity of human prostate cancer cells. Such therapy could be developed as a novel method for treating patients with androgen-independent prostate cancer.
Cheon
J
J
Department of Urology and Pathology, Korea University Hospital, Seoul, Korea. jcheon@ns.kumc.or.kr
Kim
H K
HK
Moon
D G
DG
Yoon
D K
DK
Cho
J H
JH
Koh
S K
SK
D000818:Animals; D000998:Antiviral Agents / Q000627:therapeutic use; D003587:Cytomegalovirus / Q000201:enzymology / Q000235:genetics*; D015774:Ganciclovir / Q000627:therapeutic use; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage; D006801:Humans; D008297:Male; D051379:Mice; D011471:Prostatic Neoplasms / Q000628:therapy*; D018139:Simplexvirus / Q000201:enzymology*; D018709:Statistics, Nonparametric; D013937:Thymidine Kinase / Q000235:genetics*; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000235:genetics
D000998:Antiviral Agents; D013937:Thymidine Kinase; D001616:beta-Galactosidase; D015774:Ganciclovir