Cancer-associated muscle weakness is poorly understood and there is no effective treatment. Here, we find that seven different mouse models of human osteolytic bone metastases, representing breast, lung and prostate cancers, as well as multiple myeloma exhibited impaired muscle function, implicating a role for the tumor-bone microenvironment in cancer-associated muscle weakness. We found that TGF-β, released from the bone surface as a result of metastasis-induced bone destruction upregulated NADPH oxidase 4 (Nox4), resulting in elevated oxidization of skeletal muscle proteins, including the ryanodine receptor/calcium (Ca2+) release channel (RyR1). The oxidized RyR1 channels leaked Ca2+, resulting in lower intracellular signaling required for proper muscle contraction. We found that inhibiting RyR1 leak, TGF-β signaling, TGF-β release from bone or Nox4 all improved muscle function in mice with MDA-MB-231 bone metastases. Humans with breast cancer- or lung cancer-associated bone metastases also had oxidized skeletal muscle RyR1 that is not seen in normal muscle. Similarly, skeletal muscle weakness, higher levels of Nox4 protein and Nox4 binding to RyR1, and oxidation of RyR1 were present in a mouse model of Camurati-Engelmann disease, a non-malignant metabolic bone disorder associated with increased TGF-β activity. Thus, metastasis-induced TGF-β release from bone contributes to muscle weakness by decreasing Ca2+-induced muscle force production.

Osteoporosis, a condition of skeletal decline that undermines quality of life, is treated with pharmacological interventions that are associated with poor adherence and adverse effects. Complicating efforts to improve clinical outcomes, the incidence of obesity is increasing, predisposing the population to a range of musculoskeletal complications and metabolic disorders. Pharmacological management of obesity has yet to deliver notable reductions in weight and debilitating complications are rarely avoided. By contrast, exercise shows promise as a non-invasive and non-pharmacological method of regulating both osteoporosis and obesity. The principal components of exercise — mechanical signals — promote bone and muscle anabolism while limiting formation and expansion of fat mass. Mechanical regulation of bone and marrow fat might be achieved by regulating functions of differentiated cells in the skeletal tissue while biasing lineage selection of their common progenitors — mesenchymal stem cells. An inverse relationship between adipocyte versus osteoblast fate selection from stem cells is implicated in clinical conditions such as childhood obesity and increased marrow adiposity in type 2 diabetes mellitus, as well as contributing to skeletal frailty. Understanding how exercise-induced mechanical signals can be used to improve bone quality while decreasing fat mass and metabolic dysfunction should lead to new strategies to treat chronic diseases such as osteoporosis and obesity.

Increased fracture risk is commonly reported in cancer patients receiving radiotherapy, particularly at sites within the field of treatment. The direct and systemic effects of ionizing radiation on bone at a therapeutic dose are not well characterized in clinically relevant animal models. Using twenty-week male C57Bl/6 mice, effects of irradiation (right hindlimb; 2 Gy) on bone volume and microarchitecture were evaluated prospectively by microcomputed tomography and histomorphometry and compared to contralateral-shielded bone (left hindlimb) and non-irradiated control bone. One-week post-irradiation, trabecular bone volume declined in irradiated tibiae (−22%; p<0.0001) and femora (−14%; p=0.0586) and microarchitectural parameters were compromised. Trabecular bone volume declined in contralateral tibiae (−17%; p=0.003), and no loss was detected at the femur. Osteoclast number, apoptotic osteocyte number and marrow adiposity were increased in irradiated bone relative to contralateral and non-irradiated bone, while osteoblast number was unchanged. Despite no change in osteoblast number one-week post-irradiation, dynamic bone formation indices revealed a reduction in mineralized bone surface and a concomitant increase in unmineralized osteoid surface area in irradiated bone relative to contralateral and non-irradiated control bone. Further, dose- and time-dependent calvarial culture and in vitro assays confirmed that calvarial osteoblasts and osteoblast-like MC3T3 cells were relatively radioresistant, while calvarial osteocyte and osteocyte-like MLO-Y4 cell apoptosis was induced as early as 48h post-irradiation (4 Gy). In osteoclastogenesis assays, radiation exposure (8 Gy) stimulated murine macrophage RAW264.7 cell differentiation and co-culture of irradiated RAW264.7 cells with MLO-Y4 or murine bone marrow cells enhanced this effect. These studies highlight the multi-faceted nature of radiation-induced bone loss by demonstrating direct and systemic effects on bone and its many cell types using clinically relevant doses and have important implications for bone health in patients treated with radiation therapy.

Bone metastases cause significant morbidity and mortality in late-stage breast cancer patients and are currently considered incurable. Investigators rely on translational models to better understand the pathogenesis of skeletal complications of malignancy in order to identify therapeutic targets that may ultimately prevent and treat solid tumor metastasis to bone. Many experimental models of breast cancer bone metastases are in use today, each with its own caveats. In this methods review, we characterize the bone phenotype of commonly utilized human- and murine-derived breast cell lines that elicit osteoblastic and/or osteolytic destruction of bone in mice and report methods for optimizing tumor-take in murine models of bone metastasis. We then provide protocols for four of the most common xenograft and syngeneic inoculation routes for modeling breast cancer metastasis to the skeleton in mice, including the intra-cardiac, intra-arterial, orthotopic and intra-tibial methods of tumor cell injection. Recommendations for and assessment of tumor progression and bone destruction are provided, followed by discussion of the strengths and limitations of the available tools and translational models that aid investigators in the study of breast cancer metastasis to bone.

While the chemotherapeutic effect of curcumin, one of three major curcuminoids derived from turmeric, has been reported, largely unexplored are the effects of complex turmeric extracts more analogous to traditional medicinal preparations, as well as the relative importance of the three curcuminoids and their metabolites as anti-cancer agents. These studies document the pharmacodynamic effects of chemically-complex turmeric extracts relative to curcuminoids on human breast cancer cell growth and tumor cell secretion of parathyroid hormone-related protein (PTHrP), an important driver of cancer bone metastasis. Finally, relative effects of structurally-related metabolites of curcuminoids were assessed on the same endpoints. We report that 3 curcuminoid-containing turmeric extracts differing with respect to the inclusion of additional naturally occurring chemicals (essential oils and/or polar compounds) were equipotent in inhibiting human breast cancer MDA-MB-231 cell growth (IC50=10–16μg/mL) and secretion of osteolytic PTHrP (IC50=2–3μg/mL) when concentrations were normalized to curcuminoid content. Moreover, these effects were curcuminoid-specific, as botanically-related gingerol containing extracts had no effect. While curcumin and bis-demethoxycurcumin were equipotent to each other and to the naturally occurring curcuminoid mixture (IC50=58 μM), demethoxycurcumin was without effect on cell growth. However, each of the individual curcuminoids inhibited PTHrP secretion (IC50=22–31μM) to the same degree as the curcuminoid mixture (IC50=16 μM). Degradative curcuminoid metabolites (vanillin and ferulic acid) did not inhibit cell growth or PTHrP, while reduced metabolites (tetrahydrocurcuminoids) had inhibitory effects on cell growth and PTHrP secretion but only at concentrations ≥10-fold higher than the curcuminoids. These studies emphasize the structural and biological importance of curcuminoids in the anti-breast cancer effects of turmeric and contradict recent assertions that certain of the curcuminoid metabolites studied here mediate these anti-cancer effects.