The bone is the third most common site of metastasis for a wide range of solid tumors including lung, breast, prostate, colorectal, thyroid, gynecologic, and melanoma, with 70% of metastatic prostate and breast cancer patients harboring bone metastasis. (1) Unfortunately, once cancer spreads to the bone, it is rarely cured and is associated with a wide range of morbidities including pain, increased risk of fracture, and hypercalcemia. This fact has driven experts in the fields of bone and cancer biology to study the bone, and has revealed that there is a great deal that each can teach the other. The complexity of the bone was first described in 1889 when Stephen Paget proposed that tumor cells have a proclivity for certain organs, where they "seed" into a friendly "soil" and eventually grow into metastatic lesions. Dr. Paget went on to argue that although many study the "seed" it would be paramount to understand the "soil." Since this original work, significant advances have been made not only in understanding the cell-autonomous mechanisms that drive metastasis, but also alterations which drive changes to the "soil" that allow a tumor cell to thrive. Indeed, it is now clear that the "soil" in different metastatic sites is unique, and thus the mechanisms that allow tumor cells to remain in a dormant or growing state are specific to the organ in question. In the bone, our knowledge of the components that contribute to this fertile "soil" continues to expand, but our understanding of how they impact tumor growth in the bone remains in its infancy. Indeed, we now appreciate that the endosteal niche likely contributes to tumor cell dormancy, and that osteoclasts, osteocytes, and adipocytes can impact tumor cell growth. Here, we discuss the bone microenvironment and how it impacts cancer cell seeding, dormancy, and growth. Fig. 1. Tumor cells closely associate with cells of the bone microenvironment in a mouse model of bone metastasis. Scale bar ¼ 50 mm. Ad ¼ adipocyte; EC ¼ endothelial cell; Oc ¼ osteocyte; Ocl ¼ osteoclast; Tu ¼ tumor.

Epithelial cell behavior is coordinated by the composition of the surrounding extracellular matrix (ECM); thus ECM protein identification is critical for understanding normal biology and disease states. Proteomic analyses of ECM proteins have been hindered by the insoluble and digestion-resistant nature of ECM. Here we explore the utility of combining rapid ultrasonication-and surfactant-assisted digestion for the detailed proteomics analysis of ECM samples. When compared with traditional overnight digestion, this optimized method dramatically improved the sequence coverage for collagen I, revealed the presence of hundreds of previously unidentified proteins in Matrigel, and identified a protein profile for ECM isolated from rat mammary glands that was substantially different from that found in Matrigel. In a three-dimensional culture assay to investigate epithelial cell-ECM interactions, mammary epithelial cells were found to undergo extensive branching morphogenesis when plated with mammary gland-derived matrix in comparison with Matrigel. Cumulatively these data highlight the tissue-specific nature of ECM composition and function and underscore the need for optimized techniques, such as those described here, for the proteomics characterization of ECM samples. Molecular & Cellular Proteomics 8:1648 -1657, 2009. Extracellular matrix (ECM)1 is a critical component of the tissue microenvironment. ECM plays a pivotal role in embryonic stem cell development and differentiation (1, 2) as well as many physiological (3) and pathological processes, including cancer progression (4, 5). Cell regulation by ECM has been studied with high frequency in recent years (7,8). However, our ability to globally characterize ECM composition both in vitro and in vivo has been severely limited because of several unique attributes of ECM proteins such as high molecular weight glycans and the presence of covalent protein crosslinks (6, 9, 10). Traditional proteomics approaches have proven to be ineffective for the identification of ECM proteins as demonstrated by the fact that collagens, despite being the most abundant protein in mammals, are significantly underrepresented in tissue-based proteomics data sets.Ultrasonication has long been used for the digestion of bioorganic materials to allow for maximal and reproducible extraction and hence the accurate identification of small molecule and inorganic analytes (11). More recently, Capelo et al. (12) have used ultrasonic energy to catalyze tryptic digestion of proteins for subsequent mass spectrometry-based identification. Here we sought to determine whether this method could be optimized to prepare ECM samples for mass spectrometry-based analysis. For method development, we used rat tail collagen as a representative ECM protein for which current proteomics approaches have proven relatively unsuccessful. Type I collagen is defined as a right-handed triple helix heterotrimer comprising two identical ␣1 chains and one ␣2 chain that form a fibrillar network (6). The physical properties of...

Bone destruction occurs in aging and numerous diseases, including osteoporosis and cancer. Many cancer patients have bone osteolysis that is refractory to state-of-the-art treatments, which block osteoclast activity with bisphosphonates or by inhibiting the RANKL pathway. We previously showed that macrophage-stimulating protein (MSP) signaling, which is elevated in approximately 40% of breast cancers, promotes osteolytic bone metastasis, but it was unknown whether this was due to activation of the MSP signaling pathway in the tumor cells or in the bone microenvironment. Here we show that MSP signals through host expression of its receptor, RON tyrosine kinase, to directly activate osteoclasts by a previously undescribed pathway that is complementary to RANKL signaling and converges on SRC. Genetic or pharmacologic inhibition of RON kinase blocked cancer-mediated bone destruction and osteoporosis in several mouse models. Furthermore, the RON kinase inhibitor BMS-777607/ASLAN002 altered markers of bone turnover in a first-in-human clinical cancer study, indicating potential for normalizing bone loss in patients. These findings uncover a new therapeutic target for pathogenic bone loss and provide a rationale for treatment of bone destruction in various diseases with RON inhibitors.