Purpose: In vivo studies have focused on the latter stages of the bone metastatic process (osteolysis), whereas little is known about earlier events, e.g., arrival, localization, and initial colonization. Defining these initial steps may potentially identify the critical points susceptible to therapeutic intervention. Experimental Design: MDA-MB-435 human breast cancer cells engineered with green fluorescent protein were injected into the cardiac left ventricle of athymic mice. Femurs were analyzed by fluorescence microscopy, immunohistochemistry, real-time PCR, flow cytometry, and histomorphometry at times ranging from 1hour to 6 weeks. Results: Single cells were found in distal metaphyses at 1 hour postinjection and remained as single cells up to 72 hours. Diaphyseal arrest occurred rarely and few cells remained there after 24 hours. At 1week, numerous foci (2-10 cells) were observed, mostly adjacent to osteoblast-like cells. By 2 weeks, fewer but larger foci (z50 cells) were seen. Most bones had a single large mass at 4 weeks (originating from a colony or coalescing foci) which extended into the diaphysis by 4 to 6 weeks. Little change (<20%) in osteoblast or osteoclast numbers was observed at 2 weeks, but at 4 to 6 weeks, osteoblasts were dramatically reduced (8% of control), whereas osteoclasts were reduced modestly (to f60% of control). Conclusions: Early arrest in metaphysis and minimal retention in diaphysis highlight the importance of the local milieu in determining metastatic potential.These results extend the Seed and Soil hypothesis by demonstrating both intertissue and intratissue differences governing metastatic location. Ours is the first in vivo evidence that tumor cells influence not only osteoclasts, as widely believed, but also eliminate functional osteoblasts, thereby restructuring the bone microenvironment to favor osteolysis.The data may also explain why patients receiving bisphosphonates fail to heal bone despite inhibiting resorption, implying that concurrent strategies that restore osteoblast function are needed to effectively treat osteolytic bone metastases.Breast cancer has a remarkable predilection to colonize bone, with an incidence between 70% and 85% in patients (1-3). At the time of death, metastatic bone disease accounts for the bulk of tumor burden (4). For women with bone metastases, the complications-severe, often intractable pain, pathologic fractures, and hypercalcemia-are catastrophic. Despite its obvious clinical importance, very little is understood about the fundamental mechanisms responsible for breast cancer metastasis to bone. Research progress has been hampered by the dearth of, and technical difficulties inherent in, the current models.Most models of metastasis poorly recapitulate the pathogenesis of breast cancer. The ideal model would involve dissemination from an orthotopic site (i.e., mammary fat pad), colonization, and osteolysis. None of the currently available human breast xenograft models spread to bone following orthotopic implantation and only on...

IC50 for S247 adhesion to alpha(v)beta3 or alpha(IIB)beta3a substrates was 0.2 nM vs. 244 nM, respectively. Likewise, S247 was not toxic at doses up to 1000 microM. However, osteoclast cultures treated with S247 exhibited marked morphological changes and impaired formation of the actin sealing zone. When S247 was administered prior to tumor cells, there was a significant, dose-dependent reduction (25-50% of vehicle-only-treated mice; P = 0.002) in osseous metastasis. Mice receiving S247 after tumor cell inoculation also developed fewer bone metastases, but the difference was not statistically significant. These data suggest that, in the MDA-MB-435 model, the alpha(v)beta3 integrin plays an important role in early events (e.g., arrest of tumor cells) in bone metastasis. Furthermore, the data suggest that alpha(v)beta3 inhibitors may be useful in the treatment and/or prevention of breast cancer metastases in bone.

The present study focused on CBM written language procedures by conducting an investigation of the developmental, gender, and practical considerations surrounding three categories of CBM written language scoring indices: production-dependent, production-independent, and accurateproduction. Students in first-through eighth-grade generated a three-minute writing sample in the fall and spring of the school year using standard CBM procedures. The writing samples were scored using all three types of scoring indices to assess the trends in scoring indices for students of varying ages and gender and of the time required to score writing samples using various scoring indices. With only one exception, older students outperformed younger students on all of the scoring indices. Although at the middle school level students' levels of writing fluency and writing accuracy were not closely associated, at the younger grade levels the CBM indices were significantly related. With regard to gender differences, girls outperformed boys on measures of writing fluency at all grade levels. The average scoring time per writing sample ranged from 1 1 2 _ to 2 1 2 _ minutes (depending on grade level).

Breast cancer cells exhibit a predilection for metastasis to bone. There, the metastases usually bring about bone loss with accompanying pain and loss of function. One way that breast cancer cells disrupt the normal pattern of bone remodeling is by activating osteoclasts, the bone degrading cells. Nevertheless, targeting the osteoclasts does not cure the disease or result in bone repair. These observations indicate that osteoblast function also may be compromised. The objective of this study was to investigate the interaction of metastatic breast cancer cells with osteoblasts. Human metastatic breast cancer cells, MDA-MB-435 or MDA-MB-231, or their conditioned media were co-cultured with a human osteoblast line hFOB1.19. The breast cancer cells caused an increase in the prevalence of apoptotic osteoblasts. Apoptotic osteoblasts detected by the TUNEL assay or by caspase activity increased approximately two to fivefold. This increase was not seen with non-metastatic MDA-MB-468 cells. In an investigation of the mechanism, it was determined that the hFOB1.19 cells expressed fas and that fas was functional. Likewise the hFOB1.19 cells were susceptible to TNF-alpha, but this cytokine was not detected in the conditioned medium of the breast cancer cells. This study indicates that osteoblasts are the target of breast cancer cell-induced apoptosis, but fas/fas-ligand and TNF-alpha, two common initiators of cell death, are probably not involved in this aspect of the metastases/bone cell axis. There are several mechanisms that remain to be explored in order to determine how breast cancer cells bring about osteoblast apoptosis. Even though the specific initiator of apoptosis remains to be identified, the results of this study suggest that the mechanism is likely to be novel.

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