Presentation Authors: Udit Singhal*, Yugang Wang, Yuanyuan Qiao, Therese M. Roth, Ann Arbor, MI, Jae-Seung Chung, Busan, Korea, Republic of, Jacob A. Belardo, Hao Zhang, Russell S. Taichman, Alexander B. Zaslavsky, Ganesh S. Palapattu, Arul M. Chinnaiyan, Todd M. Morgan, Ann Arbor, MI
Introduction: Wnt signaling has been implicated as a driver of prostate cancer related osteoblast differentiation, and previous studies have linked modifications in Wnt function with the induction of tumor metastasis. A unique aspect of prostate cancer bone metastases is their relative predilection to the lower compared to upper extremity. We sought to better understand this phenomenon, focusing on the role of the Wnt signaling pathway as an important driver of prostate cancer bone metastasis.
Methods: Comparative gene expression profiling was performed within the humerus and femur of mice to assess for baseline differences in the microenvironments of these locations. Specifically, RNA was isolated from bone marrow cells from sites of prostate cancer metastases in the mouse humerus and femur, and a gene expression microarray was used to identify differentially expressed genes between these sites. In vitro studies using LNCaP and PC3 cell lines included shRNA knockdown targeting Wnt signaling and conditioned media experiments with and without Wnt inhibition.
Results: Relative overexpression of Wnt signaling inhibitors WIF1 and SOST was seen in the humerus compared to the femur, with increased WNT5a expression in femur bone marrow, suggesting a coordinated upregulation of Wnt signals within the femur relative to the humerus. Conditioned medium (CM) from bone marrow stromal cells (HS-5) was used to mimic the bone marrow microenvironment, which showed a strong ability to promote prostate cancer cell invasion (3.3 fold increase in PC3 cells, p < 0.05; 7 fold increase in LNCaP cells, p < 0.05). shRNA mediated WNT5a suppression within the CM significantly reduced PC3 (56%, p < 0.05) and LNCaP (60%, p < 0.05) cell invasion. Similarly, pre-incubation of CM with WIF1 significantly blocked LNCaP cell invasion (40%, p < 0.05). shRNA mediated knockdown of Wnt receptors FZD4 and FZD8 also strongly inhibited tumor cell invasion (60% inhibition for shFZD4, p < 0.05; 63% for shFZD8, p < 0.05), suggesting that Wnt signaling factors at least partially mediate prostate cancer spread within bone.
Conclusions: Comparative gene expression profiling from separate sites of prostate cancer bone metastases reveals a potential role for Wnt signaling as a driver of prostate cancer bone metastatic tropism. Further understanding the pattern of Wnt signaling within these sites may provide a basis to appreciate the larger interplay between tumor specific characteristics and host microenvironments. These data lend further support to the potential role for Wnt targeting in advanced prostate cancer.
Source of Funding: Prostate Cancer Foundation, Department of Defense