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TrkC signaling is activated in adenoid cystic carcinoma and requires NT-3 to stimulate invasive behavior

Abstract

Treatment options for adenoid cystic carcinoma (ACC) of the salivary gland, a slowly growing tumor with propensity for neuroinvasion and late recurrence, are limited to surgery and radiotherapy. Based on expression analysis performed on clinical specimens of salivary cancers, we identified in ACC expression of the neurotrophin-3 receptor TrkC/NTRK3, neural crest marker SOX10, and other neurologic genes. Here, we characterize TrkC as a novel ACC marker, which was highly expressed in 17 out of 18 ACC primary-tumor specimens, but not in mucoepidermoid salivary carcinomas or head and neck squamous cell carcinoma. Expression of the TrkC ligand NT-3 and Tyr-phosphorylation of TrkC detected in our study suggested the existence of an autocrine signaling loop in ACC with potential therapeutic significance. NT-3 stimulation of U2OS cells with ectopic TrkC expression triggered TrkC phosphorylation and resulted in Ras, Erk 1/2 and Akt activation, as well as VEGFR1 phosphorylation. Without NT-3, TrkC remained unphosphorylated, stimulated accumulation of phospho-p53 and had opposite effects on p-Akt and p-Erk 1/2. NT-3 promoted motility, migration, invasion, soft-agar colony growth and cytoskeleton restructuring in TrkC-expressing U2OS cells. Immunohistochemical analysis demonstrated that TrkC-positive ACC specimens also show high expression of Bcl2, a Trk target regulated via Erk 1/2, in agreement with activation of the TrkC pathway in real tumors. In normal salivary gland tissue, both TrkC and Bcl2 were expressed in myoepithelial cells, suggesting a principal role for this cell lineage in the ACC origin and progression. Sub-micromolar concentrations of a novel potent Trk inhibitor AZD7451 completely blocked TrkC activation and associated tumorigenic behaviors. Pre-clinical studies on ACC tumors engrafted in mice showed efficacy and low toxicity of AZD7451, validating our in vitro data and stimulating more research into its clinical application. In summary, we describe in ACC a previously unrecognized pro-survival neurotrophin signaling pathway and link it with cancer progression.

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References

  1. Fitzpatrick PJ, Theriault C . Malignant salivary gland tumors. Int J Radiat Oncol Biol Phys 1986; 12: 1743–1747.

    Article  CAS  Google Scholar 

  2. Chummun S, McLean NR, Kelly CG, Dawes PJ, Meikle D, Fellows S et al. Adenoid cystic carcinoma of the head and neck. Br J Plast Surg 2001; 54: 476–480.

    Article  CAS  Google Scholar 

  3. Phuchareon J, Ohta Y, Woo JM, Eisele DW, Tetsu O . Genetic profiling reveals cross-contamination and misidentification of 6 adenoid cystic carcinoma cell lines: ACC2, ACC3, ACCM, ACCNS, ACCS and CAC2. PLoS One 2009; 4: e6040.

    Article  Google Scholar 

  4. Harel L, Costa B, Fainzilber M . On the death Trk. Dev Neurobiol 2010; 70: 298–303.

    CAS  PubMed  Google Scholar 

  5. Denkins Y, Reiland J, Roy M, Sinnappah-Kang ND, Galjour J, Murry BP et al. Brain metastases in melanoma: roles of neurotrophins. Neuro Oncol 2004; 6: 154–165.

    Article  CAS  Google Scholar 

  6. Jin W, Kim GM, Kim MS, Lim MH, Yun C, Jeong J et al. TrkC plays an essential role in breast tumor growth and metastasis. Carcinogenesis 2010; 31: 1939–1947.

    Article  CAS  Google Scholar 

  7. Wang T, Yu D, Lamb ML . Trk kinase inhibitors as new treatments for cancer and pain. Expert Opin Ther Pat 2009; 19: 305–319.

    Article  CAS  Google Scholar 

  8. Kumar S, Kahn MA, Dinh L, de Vellis J . NT-3-mediated TrkC receptor activation promotes proliferation and cell survival of rodent progenitor oligodendrocyte cells in vitro and in vivo. J Neurosci Res 1998; 54: 754–765.

    Article  CAS  Google Scholar 

  9. Postigo A, Calella AM, Fritzsch B, Knipper M, Katz D, Eilers A et al. Distinct requirements for TrkB and TrkC signaling in target innervation by sensory neurons. Genes Dev 2002; 16: 633–645.

    Article  CAS  Google Scholar 

  10. Barnabe-Heider F, Miller FD . Endogenously produced neurotrophins regulate survival and differentiation of cortical progenitors via distinct signaling pathways. J Neurosci 2003; 23: 5149–5160.

    Article  CAS  Google Scholar 

  11. Bartkowska K, Paquin A, Gauthier AS, Kaplan DR, Miller FD . Trk signaling regulates neural precursor cell proliferation and differentiation during cortical development. Development 2007; 134: 4369–4380.

    Article  CAS  Google Scholar 

  12. Nikoletopoulou V, Lickert H, Frade JM, Rencurel C, Giallonardo P, Zhang L et al. Neurotrophin receptors TrkA and TrkC cause neuronal death whereas TrkB does not. Nature 2010; 467: 59–63.

    Article  CAS  Google Scholar 

  13. Fernandez RM, Sanchez-Mejias A, Mena MD, Ruiz-Ferrer M, Lopez-Alonso M, Antinolo G et al. A novel point variant in NTRK3, R645C, suggests a role of this gene in the pathogenesis of Hirschsprung disease. Ann Hum Genet 2009; 73: 19–25.

    Article  CAS  Google Scholar 

  14. Sanchez-Mejias A, Fernandez RM, Lopez-Alonso M, Antinolo G, Borrego S . Contribution of RET, NTRK3 and EDN3 to the expression of Hirschsprung disease in a multiplex family. J Med Genet 2009; 46: 862–864.

    Article  CAS  Google Scholar 

  15. Tauszig-Delamasure S, Bouzas-Rodriguez J . Targeting neurotrophin-3 and its dependence receptor tyrosine kinase receptor C: a new antitumoral strategy. Expert Opin Ther Targets 2011; 15: 847–858.

    Article  CAS  Google Scholar 

  16. Bouzas-Rodriguez J, Cabrera JR, Delloye-Bourgeois C, Ichim G, Delcros JG, Raquin MA et al. Neurotrophin-3 production promotes human neuroblastoma cell survival by inhibiting TrkC-induced apoptosis. J Clin Invest 2010; 120: 850–858.

    Article  CAS  Google Scholar 

  17. Wood LD, Calhoun ES, Silliman N, Ptak J, Szabo S, Powell SM et al. Somatic mutations of GUCY2F, EPHA3, and NTRK3 in human cancers. Hum Mutat 2006; 27: 1060–1061.

    Article  Google Scholar 

  18. Lannon CL, Sorensen PH . ETV6-NTRK3: a chimeric protein tyrosine kinase with transformation activity in multiple cell lineages. Semin Cancer Biol 2005; 15: 215–223.

    Article  CAS  Google Scholar 

  19. Bardelli A, Parsons DW, Silliman N, Ptak J, Szabo S, Saha S et al. Mutational analysis of the tyrosine kinome in colorectal cancers. Science 2003; 300: 949.

    Article  CAS  Google Scholar 

  20. Davies H, Hunter C, Smith R, Stephens P, Greenman C, Bignell G et al. Somatic mutations of the protein kinase gene family in human lung cancer. Cancer Res 2005; 65: 7591–7595.

    Article  CAS  Google Scholar 

  21. Stephens P, Edkins S, Davies H, Greenman C, Cox C, Hunter C et al. A screen of the complete protein kinase gene family identifies diverse patterns of somatic mutations in human breast cancer. Nat Genet 2005; 37: 590–592.

    Article  CAS  Google Scholar 

  22. Knezevich SR, McFadden DE, Tao W, Lim JF, Sorensen PH . A novel ETV6-NTRK3 gene fusion in congenital fibrosarcoma. Nat Genet 1998; 18: 184–187.

    Article  CAS  Google Scholar 

  23. Tognon C, Knezevich SR, Huntsman D, Roskelley CD, Melnyk N, Mathers JA et al. Expression of the ETV6-NTRK3 gene fusion as a primary event in human secretory breast carcinoma. Cancer Cell 2002; 2: 367–376.

    Article  CAS  Google Scholar 

  24. Skalova A, Vanecek T, Sima R, Laco J, Weinreb I, Perez-Ordonez B et al. Mammary analogue secretory carcinoma of salivary glands, containing the ETV6-NTRK3 fusion gene: a hitherto undescribed salivary gland tumor entity. Am J Surg Pathol 2010; 34: 599–608.

    Google Scholar 

  25. Kruttgen A, Schneider I, Weis J . The dark side of the NGF family: neurotrophins in neoplasias. Brain Pathol 2006; 16: 304–310.

    Article  Google Scholar 

  26. Sinnappah-Kang ND, Kaiser AJ, Blust BE, Mrak RE, Marchetti D . Heparanase TrkC and p75NTR: their functional involvement in human medulloblastoma cell invasion. Int J Oncol 2005; 27: 617–626.

    CAS  PubMed  Google Scholar 

  27. Marchetti D, Mrak RE, Paulsen DD, Sinnappah-Kang ND . Neurotrophin receptors and heparanase: a functional axis in human medulloblastoma invasion. J Exp Clin Cancer Res 2007; 26: 5–23.

    CAS  PubMed  Google Scholar 

  28. Jin W, Yun C, Jeong J, Park Y, Lee HD, Kim SJ . c-Src is required for tropomyosin receptor kinase C (TrkC)-induced activation of the phosphatidylinositol 3-kinase (PI3K)-AKT pathway. J Biol Chem 2008; 283: 1391–1400.

    Article  CAS  Google Scholar 

  29. Sakamoto Y, Kitajima Y, Edakuni G, Sasatomi E, Mori M, Kitahara K et al. Expression of Trk tyrosine kinase receptor is a biologic marker for cell proliferation and perineural invasion of human pancreatic ductal adenocarcinoma. Oncol Rep 2001; 8: 477–484.

    CAS  PubMed  Google Scholar 

  30. Wang T, Lamb ML, Scott DA, Wang H, Block MH, Lyne PD et al. Identification of 4-aminopyrazolylpyrimidines as potent inhibitors of Trk kinases. J Med Chem 2008; 51: 4672–4684.

    Article  CAS  Google Scholar 

  31. Buzanska L, Zychowicz M, Ruiz A, Ceriotti L, Coecke S, Rauscher H et al. Neural stem cells from human cord blood on bioengineered surfaces--novel approach to multiparameter bio-tests. Toxicology 2010; 270: 35–42.

    Article  CAS  Google Scholar 

  32. Marino S . Medulloblastoma: developmental mechanisms out of control. Trends Mol Med 2005; 11: 17–22.

    Article  CAS  Google Scholar 

  33. Katoh M . Network of WNT and other regulatory signaling cascades in pluripotent stem cells and cancer stem cells. Curr Pharm Biotechnol 2011; 12: 160–170.

    Article  CAS  Google Scholar 

  34. Kelsh RN . Sorting out Sox10 functions in neural crest development. Bioessays 2006; 28: 788–798.

    Article  Google Scholar 

  35. Wu SM, Choo AB, Yap MG, Chan KK . Role of Sonic hedgehog signaling and the expression of its components in human embryonic stem cells. Stem Cell Res 2010; 4: 38–49.

    Article  CAS  Google Scholar 

  36. Bohm J, Buck A, Borozdin W, Mannan AU, Matysiak-Scholze U, Adham I et al. Sall1, sall2, and sall4 are required for neural tube closure in mice. Am J Pathol 2008; 173: 1455–1463.

    Article  Google Scholar 

  37. Katayama K, Zine A, Ota M, Matsumoto Y, Inoue T, Fritzsch B et al. Disorganized innervation and neuronal loss in the inner ear of Slitrk6-deficient mice. PLoS One 2009; 4: e7786.

    Article  Google Scholar 

  38. Fagiani E, Giardina G, Luzi L, Cesaroni M, Quarto M, Capra M et al. RaLP, a new member of the Src homology and collagen family, regulates cell migration and tumor growth of metastatic melanomas. Cancer Res 2007; 67: 3064–3073.

    Article  CAS  Google Scholar 

  39. Sundram U, Harvell JD, Rouse RV, Natkunam Y . Expression of the B-cell proliferation marker MUM1 by melanocytic lesions and comparison with S100, gp100 (HMB45), and MelanA. Mod Pathol 2003; 16: 802–810.

    Article  Google Scholar 

  40. Moskaluk CA, Baras AS, Mancuso SA, Fan H, Davidson RJ, Dirks DC et al. Development and characterization of xenograft model systems for adenoid cystic carcinoma. Lab Invest 2011; 91: 1480–1490.

    Article  CAS  Google Scholar 

  41. Labouyrie E, Dubus P, Groppi A, Mahon FX, Ferrer J, Parrens M et al. Expression of neurotrophins and their receptors in human bone marrow. Am J Pathol 1999; 154: 405–415.

    Article  CAS  Google Scholar 

  42. Marchetti D, Denkins Y, Reiland J, Greiter-Wilke A, Galjour J, Murry B et al. Brain-metastatic melanoma: a neurotrophic perspective. Pathol Oncol Res 2003; 9: 147–158.

    Article  CAS  Google Scholar 

  43. Huang EJ, Reichardt LF . Trk receptors: roles in neuronal signal transduction. Annu Rev Biochem 2003; 72: 609–642.

    Article  CAS  Google Scholar 

  44. Pylayeva-Gupta Y, Grabocka E, Bar-Sagi D . RAS oncogenes: weaving a tumorigenic web. Nat Rev Cancer 2011; 11: 761–774.

    Article  CAS  Google Scholar 

  45. Goldberg L, Kloog YA . Ras inhibitor tilts the balance between Rac and Rho and blocks phosphatidylinositol 3-kinase-dependent glioblastoma cell migration. Cancer Res 2006; 66: 11709–11717.

    Article  CAS  Google Scholar 

  46. Edsjo A, Hallberg B, Fagerstrom S, Larsson C, Axelson H, Pahlman S . Differences in early and late responses between neurotrophin-stimulated trkA- and trkC-transfected SH-SY5Y neuroblastoma cells. Cell Growth Differ 2001; 12: 39–50.

    CAS  PubMed  Google Scholar 

  47. Jaglin XH, Poirier K, Saillour Y, Buhler E, Tian G, Bahi-Buisson N et al. Mutations in the beta-tubulin gene TUBB2B result in asymmetrical polymicrogyria. Nat Genet 2009; 41: 746–752.

    Article  CAS  Google Scholar 

  48. Medjkane S, Perez-Sanchez C, Gaggioli C, Sahai E, Treisman R . Myocardin-related transcription factors and SRF are required for cytoskeletal dynamics and experimental metastasis. Nat Cell Biol 2009; 11: 257–268.

    Article  CAS  Google Scholar 

  49. Rottner K, Hanisch J, Campellone KG . WASH, WHAMM and JMY: regulation of Arp2/3 complex and beyond. Trends Cell Biol 2010; 20: 650–661.

    Article  CAS  Google Scholar 

  50. Rajan N, Elliott R, Clewes O, Mackay A, Reis-Filho JS, Burn J et al. Dysregulated TRK signalling is a therapeutic target in CYLD defective tumours. Oncogene 2011; 30: 4243–4260.

    Article  CAS  Google Scholar 

  51. Dardick I, van Nostrand AW . Myoepithelial cells in salivary gland tumors--revisited. Head Neck Surg 1985; 7: 395–408.

    Article  CAS  Google Scholar 

  52. Zandi R, Larsen AB, Andersen P, Stockhausen MT, Poulsen HS . Mechanisms for oncogenic activation of the epidermal growth factor receptor. Cell Signal 2007; 19: 2013–2023.

    Article  CAS  Google Scholar 

  53. Mehlen P . Dependence receptors: the trophic theory revisited. Sci Signal 2010; 3: pe47.

    Article  CAS  Google Scholar 

  54. Montano X . P53 associates with trk tyrosine kinase. Oncogene 1997; 15: 245–256.

    Article  CAS  Google Scholar 

  55. Aloyz RS, Bamji SX, Pozniak CD, Toma JG, Atwal J, Kaplan DR et al. p53 is essential for developmental neuron death as regulated by the TrkA and p75 neurotrophin receptors. J Cell Biol 1998; 143: 1691–1703.

    Article  CAS  Google Scholar 

  56. Lavoie JF, Lesauteur L, Kohn J, Wong J, Furtoss O, Thiele CJ et al. TrkA induces apoptosis of neuroblastoma cells and does so via a p53-dependent mechanism. J Biol Chem 2005; 280: 29199–29207.

    Article  CAS  Google Scholar 

  57. Montano X . Repression of SHP-1 expression by p53 leads to trkA tyrosine phosphorylation and suppression of breast cancer cell proliferation. Oncogene 2009; 28: 3787–3800.

    Article  CAS  Google Scholar 

  58. Frenzel A, Grespi F, Chmelewskij W, Villunger A . Bcl2 family proteins in carcinogenesis and the treatment of cancer. Apoptosis 2009; 14: 584–596.

    Article  CAS  Google Scholar 

  59. Kummoona R, Mohammad Sami S, Al-Kapptan I, Al-Muala H . Study of antiapoptotic gene of oral carcinoma by using Bcl-2 oncogene. J Oral Pathol Med 2008; 37: 345–351.

    Article  Google Scholar 

  60. Jao W, Keh PC, Swerdlow MA . Ultrastructure of the basal cell adenoma of parotid gland. Cancer 1976; 37: 1322–1333.

    Article  CAS  Google Scholar 

  61. Regezi JA, Batsakis JG . Histogenesis of salivary gland neoplasms. Otolaryngol Clin North Am 1977; 10: 297–307.

    CAS  PubMed  Google Scholar 

  62. Azevedo RS, de Almeida OP, Kowalski LP, Pires FR . Comparative cytokeratin expression in the different cell types of salivary gland mucoepidermoid carcinoma. Head Neck Pathol 2008; 2: 257–264.

    Article  Google Scholar 

  63. Hubner G, Klein HJ, Kleinsasser O, Schiefer HG . Role of myoepithelial cells in the development of salivary gland tumors. Cancer 1971; 27: 1255–1261.

    Article  CAS  Google Scholar 

  64. Hardy G, Kramer B . The myoepithelium of human major salivary glands revisited. SADJ 1998; 53: 371–375.

    CAS  PubMed  Google Scholar 

  65. Slebos RJ, Yi Y, Ely K, Carter J, Evjen A, Zhang X et al. Gene expression differences associated with human papillomavirus status in head and neck squamous cell carcinoma. Clin Cancer Res 2006; 12: 3 Pt 1 701–709.

    Article  CAS  Google Scholar 

  66. Zhang B, Kirov S, Snoddy J . WebGestalt: an integrated system for exploring gene sets in various biological contexts. Nucleic Acids Res 2005; 33: (Web Server issue): W741–W748.

    Article  CAS  Google Scholar 

  67. Subramanian A, Tamayo P, Mootha VK, Mukherjee S, Ebert BL, Gillette MA et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc Natl Acad Sci USA 2005; 102: 15545–15550.

    Article  CAS  Google Scholar 

  68. Reimand J, Arak T, Vilo J . g:Profiler--a web server for functional interpretation of gene lists (2011 update). Nucleic Acids Res 2011; 39: (Web Server issue): W307–W315.

    Article  CAS  Google Scholar 

  69. Ivanova AV, Goparaju CM, Ivanov SV, Nonaka D, Cruz C, Beck A et al. Protumorigenic role of HAPLN1 and its IgV domain in malignant pleural mesothelioma. Clin Cancer Res 2009; 15: 2602–2611.

    Article  CAS  Google Scholar 

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Acknowledgements

This study was supported by funds from the Adenoid Cystic Carcinoma Research Foundation to SI and WGY and by the NIH Challenge Grant 5RC1DE020332-02 from the National Institute of Dental and Craniofacial Research to WGY. This work was also supported in part by the Vanderbilt Ingram Cancer Center, the Vanderbilt Bill Wilkerson Center for Otolaryngology and Communication Sciences, the Robert J Kleberg Jr and Helen C Kleberg Foundation, and by an endowment to the Barry Baker Laboratory for Head and Neck Oncology.

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Ivanov, S., Panaccione, A., Brown, B. et al. TrkC signaling is activated in adenoid cystic carcinoma and requires NT-3 to stimulate invasive behavior. Oncogene 32, 3698–3710 (2013). https://doi.org/10.1038/onc.2012.377

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