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Excess TGF-β mediates muscle weakness associated with bone metastases in mice

Abstract

Cancer-associated muscle weakness is a poorly understood phenomenon, 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 transforming growth factor (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 and calcium (Ca2+) release channel (RyR1). The oxidized RyR1 channels leaked Ca2+, resulting in lower intracellular signaling, which is required for proper muscle contraction. We found that inhibiting RyR1 leakage, TGF-β signaling, TGF-β release from bone or Nox4 activity improved muscle function in mice with MDA-MB-231 bone metastases. Humans with breast- or lung cancer–associated bone metastases also had oxidized skeletal muscle RyR1 that is not seen in normal muscle. Similarly, skeletal muscle weakness, increased Nox4 binding to RyR1 and oxidation of RyR1 were present in a mouse model of Camurati-Engelmann disease, a nonmalignant metabolic bone disorder associated with increased TGF-β activity. Thus, pathological TGF-β release from bone contributes to muscle weakness by decreasing Ca2+-induced muscle force production.

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Figure 1: Skeletal muscle weakness is due to breast cancer bone metastases.
Figure 2: Treatment with Rycal (S107) improves muscle strength and lowers SR Ca2+ leakage.
Figure 3: Blocking TGF-β signaling or inhibiting bone resorption lowers SMAD3 phosphorylation and improves muscle function.
Figure 4: Blocking TGF-β ligand lowers SMAD3 phosphorylation and improves muscle function.
Figure 5: TGF-β activity leads to higher NADPH oxidase 4 (Nox4) expression and RyR1-Nox4 interaction.
Figure 6: NADPH oxidase 4 (Nox4) inhibition prevents RyR1 oxidation and improves muscle strength.

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Acknowledgements

This work was supported by the US National Institutes of Health (NIH) (grant U01CA143057 (T.A.G.) from the National Cancer Institute (NCI) Tumor Microenvironment Network, NCI-R01CA69158 (T.A.G.), NCI-R21CA179017 (G.D.R.), NHLBI-R01HL061503 (A.R.M.), NHLBI-R01HL102040 (A.R.M.), NIAMS-R01AR060037 (A.R.M.), NIH T32 HL120826 (A.R.M.), NINDS-R25NS076445 (A.R.M.), NIH-NINDS R25NS076445 (A.H.), NIH-NIAMS R01AR063943 (X.C.), and NIH-NHLBI T35 HL110854-01 (S.C. and P.K.)), the Susan G. Komen Foundation (grant SAC110013; T.A.G.), the Indiana Economic Development Grant (T.A.G.), the Jerry and Peggy Throgmartin Endowment of the Indiana University Simon Cancer Center (T.A.G.), the Indiana University Simon Cancer Center Breast Cancer Program (T.A.G.), the American Cancer Society and Indiana University Simon Cancer Center (grant IRG-84-002-28; D.L.W.), the Indiana University Health Strategic Research Initiative in Oncology (D.L.W.), the VA Merit Review Award (G.D.R.), the Fondation Leducq (A.R.M.), the Ellison Foundation (A.R.M.), the Swedish Heart Lung Foundation and Stockholm County Council (D.C.A.) and a generous donation from the Withycombe family (T.A.G.). Camurati-Engelmann disease (CED) mice were previously described and provided by X. Cao (Johns Hopkins, Baltimore, MD).

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All contributing authors have agreed to submission of this manuscript for publication. T.A.G. and A.R.M. conceived of the study. D.L.W., K.S.M., A.R.M. and T.A.G. designed and performed experiments, analyzed data and interpreted results. S.R., W.X., D.C.A., S.J., M.N., A.C., L.E.W., A.W., A.H., A.U., T.T., S.C., F.A.W. and P.K. performed experiments. M.S.B. analyzed data. G.D.R. and F.A.W. designed experiments and interpreted results. G.Z., X.W. and X.C. provided the CED mice and reviewed the results. D.L.W., K.S.M., A.R.M. and T.A.G. wrote the manuscript.

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Correspondence to Theresa A Guise.

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Competing interests

A.R.M. is a member of the board and consults for ARMGO Pharma Inc., a startup company developing RyR-targeted therapeutics; T.A.G. was a consultant and advisory board member for Novartis.

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Waning, D., Mohammad, K., Reiken, S. et al. Excess TGF-β mediates muscle weakness associated with bone metastases in mice. Nat Med 21, 1262–1271 (2015). https://doi.org/10.1038/nm.3961

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