Bone fractures and other skeletal-related events (SREs) are common, life-threatening complications of bone metastases and are a serious clinical challenge in the management of metastatic breast cancer. For breast cancer, bone is the predominant distant metastatic site. Breast cancer bone metastasis induces bone disease through accelerated bone loss, heightened fragility, and an increased risk of suffering an SRE. The current view of osteolytic tumor-induced bone disease (TIBD) is that tumor cells stimulate osteoblasts to increase osteoclast differentiation and resorption activity. Osteocytes are involved in bone metastasis as well, as they are specialized sensory bone cells that function as “managers” to sense and integrate physiochemical signals to appropriately coordinate bone tissue remodeling. Furthermore, mechanical loading, an anabolic stimulus to bone necessary for maintenance, has also been shown to be anti-tumorigenic. Despite this, the typical patient with bone metastasis is at risk for bone fracture. Therefore, little work exists exploring the role of osteocytes and mechanical loading within the context of bone metastatic breast cancer.

This thesis aims to characterize the role osteocytes play in the progression of bone metastatic cancer while also investigating how mechanical loading impacts how osteocytes respond to cancer. Presented in Chapter 2, we created a 3D hydroxyapatite scaffold to mimic the bone microenvironment to facilitate physiological analysis of osteocytes in vitro. Our scaffold proved suitable for osteocyte viability and was also able to sustain mechanical loading without causing widespread cell death. As expected, this loading resulted in an anabolic osteocyte response, validating our scaffold as a suitable model to measure these interactions. In Chapter 3, we cultured these osteocyte-seeded scaffolds in tumor-conditioned media (TCM), which contained tumor paracrine signals from primary and bone-homing cancer cell populations. Interestingly, despite mechanical loading, the bone-homing subpopulation of cancer cells was able to reverse the typical anabolic response to mechanical loading. This suggested that cancer paracrine signals were interfering with the osteocyte’s ability to sense mechanical signals, ultimately causing the osteocytes to become mechanically insensitive. Finally, in Chapter 4, we evaluated the effects of mechanical loading on mouse tibiae injected with tumor cells to determine if loading prior to inoculation could precondition bone to resist tumor engraftment and progression. While loading did not prevent engraftment, it did appear to protect trabecular bone from sclerotic lesion formation while also mitigating overall cortical bone osteolysis. Analysis of osteocytic sclerostin expression showed that cortical sclerostin was not reduced because of loading and that only highmagnitude loads were able to reduce trabecular sclerostin levels. The lack of sclerostin downregulation in response to mechanical loading supports the hypothesis that cancer may be causing osteocytes to become mechanically insensitive. Our results identify a potential mechanism by which cancer is affecting osteocytes and elucidate the roles osteocytes and mechanical loading play in the progression of bone metastatic breast cancer.