Abstract 3945: IKK-Epsilon, a novel factor identified by proteomics, contributes to prostate cancer cell-induced osteoclast activity in vitro.

Abstract

Abstract Prostate cancer preferentially metastasizes to skeleton resulting in osteoblastic lesions with underlying osteoclast activity. Novel strategies to inhibit the tumor skeletal metastasis involve targeting not only tumor cells but also bone microenvironment which is composed of osteoclasts, osteoblasts and stromal cells that include fibroblasts, immune and inflammatory cells, adipocytes, glial cells, smooth muscle cells, and resident and recruited vascular cells. The microenvironment can exert both stimulatory and inhibitory influences on proliferation and malignant properties of tumor cells. In this study, we intended to identify factors that may play key roles in this bidirectional and dynamic interaction between tumor cells and bone microenvironment. First, we fractionated the prostate cancer PC3 cell conditioned media by the molecular weight of proteins in media using centricon column. Then we applied the fractions to culturing osteoclast precursor RAW 264.7 cells in vitro. We repeated and scaled up the above steps to collect enough conditioned media, and concentrated the fractions of interest. The fractions that showed activity of osteoclast induction were determined by mass spectrometry. We found that the conditioned media contained a large amount of IKK-Epsilon, a subunit of PMA-inducible IKK (I-KappaB Kinase) complex. We further observed that recombinant IKK-Epsilon could induce osteoclast activity in vitro, but unexpectedly it had no effect on the activation of NF-kappa-B by TNF or IL-1 in these cells. Together with the findings that several identified factors in the conditioned media such as MCP-1 and IL-8 that induced osteoclast activity, in vitro validation of the identified factor IKK-Epsilon was confirmed by using neutralizing antibody on the functional assays. We have not yet identified unknown factors in the current study, however it is clear that unknown factors in the conditioned media may also play key roles in the tumor-induced osteoclast activation since blocking MCP-1, IL-8, and IKK-Epsilon did not completely block the tumor-induced osteoclast activity. We conclude that IKK-Epsilon, a novel factor identified by proteomics, may contribute to prostate cancer cell-induced osteoclast activity in vitro. Supported by NSF key project 81130046; NSF projects 81171993 and 81272415; Guangxi projects 201201ZD004 and 2012GXNSFCB53004. Citation Format: Xin Yang, Zhen Xiao, Jianhua Wang, Yi Lu, Yang Xu, Xin Huang, Atsushi Mizokami, Evan T. Keller, Jian Zhang. IKK-Epsilon, a novel factor identified by proteomics, contributes to prostate cancer cell-induced osteoclast activity in vitro. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3945. doi:10.1158/1538-7445.AM2013-3945