Inhibition of Wnt-1 in human prostate cancer cells blocks prostate cancer cell proliferation, invasion, growth and skeletal metastasis in vitro and in vivo

Proc Amer Assoc Cancer Res, Volume 47, 2006 3979 Prostate cancer is among the leading malignancies in men and is associated with a high incidence of skeletal metastases. Several studies have implicated the role of the Wnt/β-catenin pathway in a variety of malignancies including prostate cancer. In previous studies we have shown increased expression of Wnt-1 and β-catenin in highly invasive PC-3 human prostate cancer cells and in surgical biopsy specimens of prostate cancer patients with skeletal metastasis. In the current study, we evaluated the effects of inhibiting Wnt-1 in PC-3 cells using gene transfer technology. For these studies, full length cDNA encoding Wnt-1 was stably transfected into PC-3 cells in an antisense orientation. Three clones expressing the lowest levels of Wnt-1 (PC-3-Wnt-1-AS) were then selected These experimental cells exhibited a change in their morphology from spheroid to spindle shape and a decrease in cell adherence, proliferation and invasion as compared to the wild type cells and cells transfected with vector alone. These effects were associated with inhibition of E-cadherin, parathyroid hormone related peptide (PTHrP) and urokinase (uPA), genes which are involved in cell adhesion, proliferation and invasion respectively. The specificity of Wnt inhibition was confirmed by incubation of PC-3 cells with Wnt-1 antibody. In order to monitor the effect of Wnt-1 inhibition on tumor progression in vivo , male Balb c nu/nu mice were inoculated subcutaneously into the right flank with control and experimental cells. Animals inoculated with control cells developed primary tumors by 4 weeks post tumor cell inoculation and these continued to grow for the next 8 weeks. In contrast, animals inoculated with PC-3-Wnt-1-AS cells showed tumor development only at week 10 which exhibited a markedly slower rate of tumor growth. In order to evaluate the effect of Wnt-1 inhibition on experimental skeletal metastases, PC-3 and PC-3-Wnt-1-AS cells were inoculated into the tibia of male Fox Chase SCID mice. Radiological evidence of skeletal lesions was evident in tibias of 100% of animals inoculated with PC-3 cells. However only 10% of animals inoculated with PC-3-Wnt-1-AS cells showed any evidence of skeletal lesions as analyzed by X-ray, microCT and bone histomorphometric analyses. Furthermore immunohistochemical analysis of control and experimental primary tumors exhibited a marked inhibition of E-cadherin, PTHrP and uPA production in tumors from experimental animals inoculated with PC-3-Wnt-1-AS cells. The results obtained in these studies provide compelling evidence for the role of Wnt-1 and its signalling pathway in the establishment and progression of prostate cancer and its capacity to metastasize to the skeleton. Further elucidation of the role of this pathway will lead to the development of new strategies to block prostate cancer growth and metastasis in prostate cancer patients.