Sema4D expression and secretion are increased by HIF-1α and inhibit osteogenesis in bone metastases of lung cancer

  • Wu-gui Chen
  • Jing Sun
  • Wei-wei Shen
  • Si-zhen Yang
  • Ying Zhang
  • Xu Hu
  • Hao Qiu
  • Shang-cheng XuEmail author
  • Tong-wei ChuEmail author
Research Paper


Most lung cancer bone metastasis are characterized by osteolytic destruction and osteoblastic activity is significantly decreased, suggesting that hypoxia may play a critical role in the process, but the underlying mechanisms remain unknown. Semaphorin 4D (Sema4D) is a recently discovered osteogenic inhibitory factor that is expressed at high levels in lung cancers. Here, CoCl2-induced hypoxia significantly enhanced the inhibitory effect of lung cancer cell conditioned media on osteoblast differentiation by inducing the expression and secretion of Sema4D in a HIF-1α- but not HIF-2α-dependent manner. Moreover, HIF-1α directly regulated Sema4D expression by binding to bases 1171 to 798 in the Sema4D promoter. Furthermore, hypoxia increased Sema4D secretion by upregulating a disintegrin and metalloproteinase 17 (ADAM17) expression in lung cancer in a HIF-1α-dependent manner. In bone metastasis samples from 49 patients with lung cancer, Sema4D and ADAM17 expression significantly correlated with HIF-1α expression and strongly correlated with a poor differentiation status and osteolytic bone destruction. These results provide the first evidence that HIF-1α-induced Sema4D expression and secretion play important roles in lung cancer osteolytic bone metastasis by inhibiting osteoblast differentiation, thereby providing potential strategies for the treatment of bone metastasis via targeting osteoblasts.


Hypoxia-inducing factor Semaphorin 4D ADAM17 Bone metastasis Osteoblast Lung cancer 



Semaphorin 4D


Hypoxia-inducible factor-1α


Hypoxia-inducible factor-2α


A disintegrin and metalloproteinase-17


Disseminated tumor cells


Parathyroid hormone-related protein




Tumor necrosis factor-α


Receptor activator of NF-κB ligand


Conditioned media


Hypoxia response elements


Stromal cell-derived factor-1


Vascular endothelial growth factor


Monocyte chemotactic protein-1


Dickkopf 1 gene


Macrophage inflammatory protein-2


Transforming growth factor-β



We are grateful for the valuable assistance provided by Dr. Wei Xie (Department of central laboratory, Xinqiao Hospital, Army Medical University, Chongqing, China), Xiao-Jian Niu, MS, Wei Liu, MS, and Tao-song Li, MS (Department of Orthopedics, Xinqiao Hospital, Army Medical University, Chongqing, China) during the study. This research was supported by the National Natural Science Foundation of China (Grant Number 81570800) and the National Natural Youth Science Fund of China (Grant Number 81501853).

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

10585_2018_9951_MOESM1_ESM.docx (15 kb)
Supplementary material 1 (DOCX 15 KB)


  1. 1.
    Chen W, Zheng R, Baade PD, Zhang S, Zeng H, Bray F, Jemal A, Yu XQ, He J (2016) Cancer statistics in China, 2015. CA 66(2):115. CrossRefPubMedGoogle Scholar
  2. 2.
    Siegel RL, Miller KD, Jemal A (2017) Cancer statistics, 2017. CA 65(1):5–29. CrossRefGoogle Scholar
  3. 3.
    Coleman RE (1997) Skeletal complications of malignancy. Cancer 80 (8 Suppl):1588–1594CrossRefGoogle Scholar
  4. 4.
    Rove KO, Crawford ED (2009) Metastatic cancer in solid tumors and clinical outcome: skeletal-related events. Oncology 23(5):21–27PubMedGoogle Scholar
  5. 5.
    Yang Y, Ma Y, Sheng J, Huang Y, Zhao Y, Fang W, Hong S, Tian Y, Xue C, Zhang L (2016) A multicenter, retrospective epidemiologic survey of the clinical features and management of bone metastatic disease in China. Chin J Cancer 35(5):223–228. CrossRefGoogle Scholar
  6. 6.
    Sugiura H, Yamada K, Sugiura T, Hida T, Mitsudomi T (2008) Predictors of survival in patients with bone metastasis of lung cancer. Clin Orthop Relat Res 466(3):729–736. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Tang Y, Qu J, Wu J, Liu H, Chu T, Xiao J, Zhou Y (2016) Effect of surgery on quality of life of patients with spinal metastasis from non-small-cell lung cancer. J Bone Joint Surg Am 98(5):396–402. CrossRefPubMedGoogle Scholar
  8. 8.
    Gdowski AS, Ranjan A, Vishwanatha JK (2017) Current concepts in bone metastasis, contemporary therapeutic strategies and ongoing clinical trials. J Exp Clin Cancer Res 36(1):108. CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Suva LJ, Washam C, Nicholas RW, Griffin RJ (2011) Bone metastasis: mechanisms and therapeutic opportunities. Nat Rev Endocrinol 7(4):208. CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Clézardin P (2017) Pathophysiology of bone metastases from solid malignancies. Joint Bone Spine Revue Du Rhumatisme. CrossRefGoogle Scholar
  11. 11.
    Wu MY, Li CJ, Yiang GT, Cheng YL, Tsai AP, Hou YT, Ho YC, Hou MF, Chu PY (2018) Molecular regulation of bone metastasis pathogenesis. Cell Physiol Biochem 46(4):1423–1438. CrossRefPubMedGoogle Scholar
  12. 12.
    Spencer JA, Ferraro F, Roussakis E, Klein A, Wu J, Runnels JM, Zaher W, Mortensen LJ, Alt C, Turcotte R, Yusuf R, Cote D, Vinogradov SA, Scadden DT, Lin CP (2014) Direct measurement of local oxygen concentration in the bone marrow of live animals. Nature 508(7495):269–273. CrossRefGoogle Scholar
  13. 13.
    Johnson RW, Sowder ME, Giaccia AJ (2017) Hypoxia and bone metastatic disease. Curr Osteoporos Rep 7:1–8. CrossRefGoogle Scholar
  14. 14.
    Araos J, Sleeman JP, Garvalov BK (2018) The role of hypoxic signalling in metastasis: towards translating knowledge of basic biology into novel anti-tumour strategies. Clin Exp Metastasis 35(7):563–599. CrossRefPubMedGoogle Scholar
  15. 15.
    Hiraga T, Kizaka-Kondoh S, Hirota K, Hiraoka M, Yoneda T (2007) Hypoxia and hypoxia-inducible factor-1 expression enhance osteolytic bone metastases of breast cancer. Can Res 67(9):4157–4163. CrossRefGoogle Scholar
  16. 16.
    Storti P, Bolzoni M, Donofrio G, Airoldi I, Guasco D, Toscani D, Martella E, Lazzaretti M, Mancini C, Agnelli L (2013) Hypoxia-inducible factor (HIF)-1α suppression in myeloma cells blocks tumoral growth in vivo inhibiting angiogenesis and bone destruction. Leukemia 27(8):1697–1706. CrossRefPubMedGoogle Scholar
  17. 17.
    Shiozawa Y, Pedersen EA, Havens AM, Jung Y, Mishra A, Joseph J, Jin KK, Patel LR, Chi Y, Ziegler AM (2011) Human prostate cancer metastases target the hematopoietic stem cell niche to establish footholds in mouse bone marrow. J Clin Investig 121(4):1298–1312CrossRefGoogle Scholar
  18. 18.
    Jeong HM, Cho SW, Park SI (2016) Osteoblasts are the centerpiece of the metastatic bone microenvironment. Endocrinol Metab 31 4): 485 – 92. CrossRefGoogle Scholar
  19. 19.
    Han Y, You X, Xing W, Zhang Z, Zou W (2018) Paracrine and endocrine actions of bone-the functions of secretory proteins from osteoblasts, osteocytes, and osteoclasts.
  20. 20.
    Ruan SS, Li RC, Han Y, Liu J, Li GL, Song YQ, Wu G (2014) Expression and clinical significance of Semaphorin4D in non-small cell lung cancer and its impact on malignant behaviors of A549 lung cancer cells. J Huazhong Univ Sci Technol 34(4):491–496. CrossRefGoogle Scholar
  21. 21.
    Potiron VA, Roche J, Drabkin HA (2009) Semaphorins and their receptors in lung cancer. Cancer Lett 273(1):1. CrossRefPubMedGoogle Scholar
  22. 22.
    Jiang H, Chen C, Sun Q, Wu J, Qiu L, Gao C, Liu W, Yang J, Nie J, Dong J (2016) The role of semaphorin 4D in tumor development and angiogenesis in human breast cancer. Oncotargets Ther 9:5737–5750. CrossRefGoogle Scholar
  23. 23.
    Ch’Ng ES, Kumanogoh A (2010) Roles of Sema4D and Plexin-B1 in tumor progression. Mol Cancer 9(1):251. CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Zhou H, Yang YH, Binmadi NO, Proia P, Basile JR (2012) The hypoxia-inducible factor-responsive proteins semaphorin 4D and vascular endothelial growth factor promote tumor growth and angiogenesis in oral squamous cell carcinoma. Exp Cell Res 318(14):1685–1698. CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Sun Q, Zhou H, Binmadi NO, Basile JR (2011) Hypoxia-inducible factor-1-mediated regulation of semaphorin 4D affects tumor growth and vascularity. J Biol Chem 284(46):32066. CrossRefGoogle Scholar
  26. 26.
    Ying C, Lei Z, Yi P, Ren X, Quan H (2012) Over-expression of semaphorin4D, hypoxia-inducible factor-1α and vascular endothelial growth factor is related to poor prognosis in ovarian epithelial cancer. Int J Mol Sci 13(10):13264. CrossRefGoogle Scholar
  27. 27.
    T N-K MS, RH NKHBTK F, H T (2011) Suppression of bone formation by osteoclastic expression of semaphorin 4D. Nat Med 17(11):1473–1480. CrossRefGoogle Scholar
  28. 28.
    Zhang Y, Wei L, Miron RJ, Shi B, Bian Z (2016) Bone scaffolds loaded with siRNA-Semaphorin4d for the treatment of osteoporosis related bone defects. Sci Rep 6:26925CrossRefGoogle Scholar
  29. 29.
    Zhang Y, Wei L, Miron RJ, Shi B, Bian Z (2015) Anabolic bone formation via a site-specific bone-targeting delivery system by interfering with semaphorin 4D expression. J Bone Miner Res 30(2):286–296. CrossRefPubMedGoogle Scholar
  30. 30.
    Terpos E, Ntanasisstathopoulos I, Christoulas D, Bagratuni T, Bakogeorgos M, Gavriatopoulou M, Eleutherakispapaiakovou E, Kanellias N, Kastritis E, Dimopoulos MA (2018) Semaphorin 4D correlates with increased bone resorption, hypercalcemia, and disease stage in newly diagnosed patients with multiple myeloma. Blood Cancer J.
  31. 31.
    Yang YH, Buhamrah A, Schneider A, Lin YL, Zhou H, Bugshan A, Basile JR (2016) Semaphorin 4D promotes skeletal metastasis in breast cancer. Plos ONE 11(2):e0150151. CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Hu F, Deng X, Yang X, Jin H, Gu D, Lv X, Wang C, Zhang Y, Huo X, Shen Q (2015) Hypoxia upregulates Rab11-family interacting protein 4 through HIF-1α to promote the metastasis of hepatocellular carcinoma. Oncogene 34(49):6007–6017. CrossRefPubMedGoogle Scholar
  33. 33.
    Sun Q, Zhou H, Binmadi NO, Basile JR (2009) Hypoxia-inducible factor-1-mediated regulation of semaphorin 4D affects tumor growth and vascularity. J Biol Chem 284(46):32066–32074. CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Elhabazi A, Delaire S, Bensussan A, Boumsell L, Bismuth G (2001) Biological activity of soluble CD100. I. The extracellular region of CD100 is released from the surface of T lymphocytes by regulated proteolysis. J Immunol 166(7):4341–4347CrossRefGoogle Scholar
  35. 35.
    Zhu L, Bergmeier W, Wu J, Jiang H, Stalker TJ, Cieslak M, Fan R, Boumsell L, Kumanogoh A, Kikutani H (2007) Regulated surface expression and shedding support a dual role for semaphorin 4D in platelet responses to vascular injury. Proc Natl Acad Sci USA 104(5):1621–1626. CrossRefPubMedGoogle Scholar
  36. 36.
    Maleki KT, Cornillet M, Bjorkstrom NK (2016) Soluble SEMA4D/CD100: a novel immunoregulator in infectious and inflammatory diseases. Clin Immunol (Orlando Fla) 163:52–59. CrossRefGoogle Scholar
  37. 37.
    Chen T, Xu DZ, Li Q, Mou P, Zeng Z, Brass LF, Zhu L (2016) The regulation of Sema4D exodomain shedding by protein kinase A in platelets. Platelets 27(7):673. CrossRefPubMedGoogle Scholar
  38. 38.
    Mark E, Yibin K (2014) Targeting tumor-stromal interactions in bone metastasis. Pharmacol Ther 141(2):222–233. CrossRefGoogle Scholar
  39. 39.
    Rahim F, Hajizamani S, Mortaz E, Ahmadzadeh A, Shahjahani M, Shahrabi S, Saki N (2014) Molecular regulation of bone marrow metastasis in prostate and breast cancer. Bone Marrow Res 2014(4):405920. CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Liu F, Shen W, Qiu H, Hu X, Zhang C, Chu T (2015) Prostate cancer cells induce osteoblastic differentiation via semaphorin 3A. Prostate 75(4):370–380. CrossRefPubMedGoogle Scholar
  41. 41.
    Wang LM, Zhao N, Zhang J, Sun QF, Yang CZ, Yang PS (2018) Tumor necrosis factor-alpha inhibits osteogenic differentiation of pre-osteoblasts by downregulation of EphB4 signaling via activated nuclear factor-kappaB signaling pathway. 53(1): 66–72.
  42. 42.
    JS RLLUBGMCYS, CM C, JC D K (2015) High Glucose Up-regulates ADAM17 through HIF-1α in Mesangial Cells. J Biol Chem 290(35):21603–21614. CrossRefGoogle Scholar
  43. 43.
    Charbonneau M, Harper K, Grondin F, Pelmus M, Mcdonald PP, Dubois CM (2007) Hypoxia-inducible factor mediates hypoxic and tumor necrosis factor alpha-induced increases in tumor necrosis factor-alpha converting enzyme/ADAM17 expression by synovial cells. J Biol Chem 282(46):33714. CrossRefPubMedGoogle Scholar
  44. 44.
    Rzymski T, Petry A, Kračun D, Riess F, Pike L, Harris AL, Görlach A (2012) The unfolded protein response controls induction and activation of ADAM17/TACE by severe hypoxia and ER stress. Oncogene 31(31):3621. CrossRefPubMedGoogle Scholar
  45. 45.
    Lin Q, Cong X, Yun Z (2011) Differential hypoxic regulation of hypoxia-inducible factors 1alpha and 2alpha. Mol Cancer Res 9(6):757. CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Z PM, X ZQLXL, KM X, C W, LF RRL B, L Z (2013) Identification of a calmodulin-binding domain in Sema4D that regulates its exodomain shedding in platelets. Blood 121(20):4221–4230. CrossRefGoogle Scholar
  47. 47.
    Mou P, Zeng Z, Li Q, Liu X, Xin X, Wannemacher KM, Ruan C, Li R, Brass LF, Zhu L (2013) Identification of a calmodulin-binding domain in Sema4D that regulates its exodomain shedding in platelets. Blood 121(20):4221–4230. CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Black RA (1997) A metalloproteinase disintegrin that releases tumour-necrosisi factor-α from cells. Nature.
  49. 49.
    Zhou BBS, Peyton M, He B, Liu C, Girard L, Caudler E, Lo Y, Baribaud F, Mikami I, Reguart N (2006) Targeting ADAM-mediated ligand cleavage to inhibit HER3 and EGFR pathways in non-small cell lung cancer. Cancer Cell 10(1):39–50. CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Xu P, Derynck R (2010) Direct activation of TACE-mediated ectodomain shedding by p38 MAP kinase regulates EGF receptor-dependent cell proliferation. Mol Cell 37(4): 551–566.
  51. 51.
    Sharma A, Bender S, Zimmermann M, Riesterer O, Broggini-Tenzer A, Pruschy MN (2016) Secretome signature identifies ADAM17 as novel target for radiosensitization of non-small cell lung cancer. Clin Cancer Res 22(17):4428–4439. CrossRefPubMedGoogle Scholar
  52. 52.
    Ni SS, Zhang J, Zhao WL, Dong XC, Wang JL (2013) ADAM17 is overexpressed in non-small cell lung cancer and its expression correlates with poor patient survival. Tumour Biol 34(3):1813–1818. CrossRefPubMedGoogle Scholar
  53. 53.
    Sierra JR, Corso S, Caione L, Cepero V, Conrotto P, Cignetti A, Piacibello W, Kumanogoh A, Kikutani H, Comoglio PM, Tamagnone L, Giordano S (2008) Tumor angiogenesis and progression are enhanced by Sema4D produced by tumor-associated macrophages. J Exp Med 205(7):1673–1685. CrossRefPubMedPubMedCentralGoogle Scholar
  54. 54.
    Yoshida Y, Ogata A, Kang S, Ebina K, Shi K, Nojima S, Kimura T, Ito D, Morimoto K, Nishide M (2015) Semaphorin 4D contributes to rheumatoid arthritis by inducing inflammatory cytokine production: pathogenic and therapeutic implications. Arthritis Rheumatol 67(6):1481–1490. CrossRefPubMedPubMedCentralGoogle Scholar
  55. 55.
    Johnson RW, Schipani E, Giaccia AJ (2015) HIF targets in bone remodeling and metastatic disease. Pharmacol Ther 150:169–177.
  56. 56.
    Zhang Y, Wei L, Miron RJ, Zhang Q, Bian Z (2014) Prevention of alveolar bone loss in an osteoporotic animal model via interference of semaphorin 4d. J Dent Res 93(11):1095–1100. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Department of Orthopedics, Xinqiao HospitalArmy Medical UniversityChongqingChina
  2. 2.Department of OrthopedicsLanzhou Military Region General HospitalLanzhouChina
  3. 3.The Center of Laboratory MedicineThe Sixth People’s Hospital of ChongqingChongqingChina

Personalised recommendations