Investigational New Drugs

, Volume 30, Issue 2, pp 490–507 | Cite as

A new diaryl urea compound, D181, induces cell cycle arrest in the G1 and M phases by targeting receptor tyrosine kinases and the microtubule skeleton

  • Jin Zhang
  • Jing Zhou
  • Xiaomei Ren
  • Yanyan Diao
  • Honglin Li
  • Hualiang Jiang
  • Ke Ding
  • Duanqing Pei


Receptor tyrosine kinases (RTKs) modulate a variety of cellular events, including cell proliferation, differentiation, mobility and apoptosis. In addition, RTKs have been validated as targets for cancer therapies. Microtubules are another class of proven targets for many clinical anticancer drugs. Here, we report that 1-(4-chloro-3-(trifluoromethyl) phenyl)-3-(2-cyano-4-hydroxyphenyl)urea (D181) functions as both a receptor tyrosine kinase inhibitor and a tubulin polymerization enhancer. D181 displayed potent inhibitory activities against a panel of RTKs, including Flt3, VEGFR, cKit, FGFR1 and PDGFRβ. D181 also enhanced tubulin polymerization and modified the secondary structure of tubulin proteins to disrupt their dynamic instability. Because of synergistic cooperation, D181 strongly inhibited the proliferation of various cancer cell lines, induced LoVo cell cycle arrest in the G1 and M phases and suppressed tumor growth in nude mice bearing human LoVo and HT29 xenografts. Our studies have provided a new, promising lead compound and novel clues for multi-target anticancer drug design and development.


D181 Receptor tyrosine kinase G1/M arrest Tubulin polymerization 



This work was financially supported by the 100-talent program of CAS, CAS grant (KSCX2-YWR-27), the National Natural Science Foundation (Grant # 90813033) and the National High Technology Research and Development Program (Grant # 2008AA02Z420, 2009CB940904, 2010CB529706).


  1. 1.
    Schlessinger J (2000) Cell signaling by receptor tyrosine kinases. Cell 103(2):211–225PubMedCrossRefGoogle Scholar
  2. 2.
    Krause DS, Van Etten RA (2005) Tyrosine kinases as targets for cancer therapy. N Engl J Med 353(2):172–187PubMedCrossRefGoogle Scholar
  3. 3.
    Bild AH, Nevins JR et al (2006) Oncogenic pathway signatures in human cancers as a guide to targeted therapies. Nature 439(7074):353–357PubMedCrossRefGoogle Scholar
  4. 4.
    Xu AM, Huang PH (2010) Receptor tyrosine kinase coactivation networks in cancer. Cancer Res 70(10):3857–3860PubMedCrossRefGoogle Scholar
  5. 5.
    Lemmon MA, Schlessinger J (2010) Cell signaling by receptor tyrosine kinases. Cell 141(7):1117–1134PubMedCrossRefGoogle Scholar
  6. 6.
    Shih T, Lindley C (2006) Bevacizumab: an angiogenesis inhibitor for the treatment of solid malignancies. Clin Ther 28(11):1779–1802PubMedCrossRefGoogle Scholar
  7. 7.
    Di Costanzo F, Fet M et al (2008) Bevacizumab in non-small cell lung cancer. Drugs 68(6):737–746PubMedCrossRefGoogle Scholar
  8. 8.
    Vigneri P, Wang JY (2001) Induction of apoptosis in chronic myelogenous leukemia cells through nuclear entrapment of BCR-ABL tyrosine kinase. Nat Med 7(2):228–234PubMedCrossRefGoogle Scholar
  9. 9.
    Druker BJ, Guilhot F et al (2006) Five-year follow-up of patients receiving imatinib for chronic myeloid leukemia. N Engl J Med 355(23):2408–2417PubMedCrossRefGoogle Scholar
  10. 10.
    Kobayashi S, Halmos B et al (2005) EGFR mutation and resistance of non-small-cell lung cancer to gefitinib. N Engl J Med 352(8):786–792PubMedCrossRefGoogle Scholar
  11. 11.
    Toyooka S, Mitsudomi T et al (2005) EGFR mutation and response of lung cancer to gefitinib. N Engl J Med 352(20):2136PubMedCrossRefGoogle Scholar
  12. 12.
    Tsao MS, Shepherd FA (2005) Erlotinib in lung cancer-molecular and clinical predictors of outcome. N Engl J Med 353(2):133–144PubMedCrossRefGoogle Scholar
  13. 13.
    Weinstein IB, Joe AK (2006) Mechanisms of disease: oncogene addiction–a rationale for molecular targeting in cancer therapy. Nat Clin Pract Oncol 3(8):448–457PubMedCrossRefGoogle Scholar
  14. 14.
    Luo J, Elledge SJ et al (2009) Principles of cancer therapy: oncogene and non-oncogene addiction. Cell 136(5):823–837PubMedCrossRefGoogle Scholar
  15. 15.
    Wilhelm SM, Trail PA et al (2004) BAY 43-9006 exhibits broad spectrum oral antitumor activities and targets the RAF/MEK/ERK pathway and receptor tyrosine kinases involved in tumor progression and angiogenesis. Cancer Res 64(19):7099–7109PubMedCrossRefGoogle Scholar
  16. 16.
    Liu L, Cao Y et al (2006) Sorafenib blocks the RAF/MEK/ERK pathway, inhibits tumor angiogenesis, and induces tumor cell apoptosis in hepatocellular carcinoma model PLC/PRF/5. Cancer Res 66(24):1851–1858CrossRefGoogle Scholar
  17. 17.
    Schueneman AJ, Hallahan DE et al (2003) SU11248 maintenance therapy prevents tumor regrowth after fractionated irradiation of murine tumor models. Cancer Res 63(14):4009–4016PubMedGoogle Scholar
  18. 18.
    Xin H, Yu H et al (2009) Sunitinib inhibition of Stat3 induces renal cell carcinoma tumor cell apoptosis and reduces immunosuppressive cells. Cancer Res 69(6):2506–2513PubMedCrossRefGoogle Scholar
  19. 19.
    Cleveland DW (1982) Treadmilling of tubulin and actin. Cell 28(4):689–691PubMedCrossRefGoogle Scholar
  20. 20.
    Downing KH, Nogales E (1998) Tubulin and microtubule structure. Curr Opin Cell Biol 10(1):16–22PubMedCrossRefGoogle Scholar
  21. 21.
    Kueh HY, Mitchison TJ (2009) Structural plasticity in actin and tubulin polymer dynamics. Science 325(5943):960–963PubMedCrossRefGoogle Scholar
  22. 22.
    Nicolaou KC, Wrasidlo W et al (1993) Design, synthesis and biological activity of protaxols. Nature 364(6436):464–466PubMedCrossRefGoogle Scholar
  23. 23.
    Donoso JA, Samson FE et al (1977) Action of the vinca alkaloids vincristine, vinblastine, and desacetyl vinblastine amide on axonal fibrillar organelles in vitro. Cancer Res 37(5):1401–1407PubMedGoogle Scholar
  24. 24.
    Okouneva T, Hill BT (2003) The effects of vinflunine, vinorelbine, and vinblastine on centromere dynamics. Mol Cancer Ther 2(5):427–436PubMedGoogle Scholar
  25. 25.
    Shih C, Teicher BA (2001) Cryptophycins: a novel class of potent antimitotic antitumor depsipeptides. Curr Pharm Des 7(13):1259–1276PubMedCrossRefGoogle Scholar
  26. 26.
    Brossi A, Chignell CF et al (1983) Biological effects of modified colchicines. 2. Evaluation of catecholic colchicines, colchifolines, colchicide, and novel N-acyl- and N-aroyldeacetylcolchicines. J Med Chem 26(10):1365–1369PubMedCrossRefGoogle Scholar
  27. 27.
    Muhlradt PF, Sasse F (1997) Epothilone B stabilizes microtubuli of macrophases like taxol without showing taxol-like endotoxin activity. Cancer Res 57(16):3344–3346PubMedGoogle Scholar
  28. 28.
    Marx MA (2002) Small-molecule, tubulin-binding compounds as vascular targeting agents. Expert Opin Ther Pat 12(6):769–776CrossRefGoogle Scholar
  29. 29.
    Mani S, Colevas D et al (2004) The clinical development of new mitotic inhibitors that stabilize the microtubule. Anticancer Drugs 15(6):553–558PubMedCrossRefGoogle Scholar
  30. 30.
    Belleri M, Presta M et al (2005) Antiangiogenic and vascular-targeting activity of the microtubule-destabilizing trans-resveratrol derivative 3, 5, 4′-trimethoxystilbene. Mol Pharmacol 67(5):1451–1459PubMedCrossRefGoogle Scholar
  31. 31.
    Delmonte A, Sessa C (2009) AVE8062: a new combretastatin derivative vascular disrupting agent. Expert Opin Investig Drugs 18(10):1541–1548PubMedCrossRefGoogle Scholar
  32. 32.
    Halgren TA, Murphy RB et al (2004) Glide: a new approach for rapid, accurate docking and scoring enrichment factors in database screening. J Med Chem 47:1750–1759PubMedCrossRefGoogle Scholar
  33. 33.
    Sherman W, Farid R et al (2006) Novel procedure for modeling hLigand/Receptor induced fit effects. J Med Chem 49:534–553PubMedCrossRefGoogle Scholar
  34. 34.
    Merrill GF (1998) Cell synchronization. Meth Cell Biol 57:229–249CrossRefGoogle Scholar
  35. 35.
    Davis PK, Ho A, Dowdy SF (2001) Biological methods for cell-cycle synchronization of mammalian cells. Biotechniques 30(6):1322–1331PubMedGoogle Scholar
  36. 36.
    Fang L, Yang B et al (2008) MZ3 can induce G2/M-phase arrest and apoptosis in human leukemia cells. J Cancer Res Clin Oncol 134(12):1337–1345PubMedCrossRefGoogle Scholar
  37. 37.
    Avila J, Zabala JC et al (2008) Isolation of microtubules and microtubule proteins. Curr Protoc Cell Biol 2008 Jun; Chapter 3:Unit 3.9.Google Scholar
  38. 38.
    Whitmore L, Wallace BA (2008) Protein secondary structure analyses from circular dichroism spectroscopy: methods and reference databases. Biopolymers 89(5):392–400PubMedCrossRefGoogle Scholar
  39. 39.
    Ramos-Vara JA (2005) Technical aspects of immunohistochemistry. Vet Pathol 42(4):405–426PubMedCrossRefGoogle Scholar
  40. 40.
    Sanz M, Löwenberg B et al (2009) FLT3 inhibition as a targeted therapy for acute myeloid leukemia. Curr Opin Oncol 21(6):594–600PubMedCrossRefGoogle Scholar
  41. 41.
    Pratz KW, Levis MJ (2010) Bench to bedside targeting of FLT3 in acute leukemia. Curr Drug Targets 11(7):781–789PubMedCrossRefGoogle Scholar
  42. 42.
    Appelmann I, Liersch R, Kessler T (2010) Angiogenesis inhibition in cancer therapy: platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) and their receptors: biological functions and role in malignancy. Recent Results Cancer Res 180:51–81PubMedCrossRefGoogle Scholar
  43. 43.
    Hellberg C, Ostman A, Heldin CH (2010) PDGF and vessel maturation. Recent Results Cancer Res 180:103–114PubMedCrossRefGoogle Scholar
  44. 44.
    Niu G, Chen X (2010) Vascular endothelial growth factor as an anti-angiogenic target for cancer therapy. Curr Drug Targets 11(8):1000–1017PubMedCrossRefGoogle Scholar
  45. 45.
    Ciardiello F, Tortora G (2008) EGFR antagonists in cancer treatment. N Engl J Med 358(11):1160–1174PubMedCrossRefGoogle Scholar
  46. 46.
    O’Farrell AM, Abrams TJ et al (2003) SU11248 is a novel FLT3 tyrosine kinase inhibitor with potent activity in vitro and in vivo. Blood 101(9):3597–3605PubMedCrossRefGoogle Scholar
  47. 47.
    Zarrinkar PP, Gunawardane RN et al (2009) AC220 is a uniquely potent and selective inhibitor of FLT3 for the treatment of acute myeloid leukemia (AML). Blood 114(14):2984–2992PubMedCrossRefGoogle Scholar
  48. 48.
    Ma J, Ding J et al (2008) The marine-derived oligosaccharide sulfate (MdOS), a novel multiple tyrosine kinase inhibitor, combats tumor angiogenesis both in vitro and in vivo. PLoS ONE 3(11):e3774PubMedCrossRefGoogle Scholar
  49. 49.
    Matei D, Chang DD et al (2004) Imatinib mesylate (Gleevec) inhibits ovarian cancer cell growth through a mechanism dependent on platelet-derived growth factor receptor alpha and Akt inactivation. Clin Cancer Res 10(2):681–690PubMedCrossRefGoogle Scholar
  50. 50.
    Chen J, Higgins B et al (2007) Antitumor activity of HER1/EGFR tyrosine kinase inhibitor erlotinib, alone and in combination with CPT-11 (irinotecan) in human colorectal cancer xenograft models. Cancer Chemother Pharmacol 59(5):651–659PubMedCrossRefGoogle Scholar
  51. 51.
    Noble ME, Endicott JA, Johnson LN (2004) Protein kinase inhibitors: insights into drug design from structure. Science 303:1800–1805PubMedCrossRefGoogle Scholar
  52. 52.
    Mahboobi S, Dove S et al (2006) Novel bis(1 H-indol-2-yl)methanones as potent inhibitors of FLT3 and platelet-derived growth factor receptor tyrosine kinase. J Med Chem 49(11):3101–3115PubMedCrossRefGoogle Scholar
  53. 53.
    Wilhelm S, Carter C et al (2006) Discovery and development of Sorafenib: a multikinase inhibitor for treating cancer. Nat Rev Drug Disc 5:835–845CrossRefGoogle Scholar
  54. 54.
    Lukas J, Bartek J (1995) Retinoblastoma-protein-dependent cell-cycle inhibition by the tumour suppressor p16. Nature 375(6531):503–506PubMedCrossRefGoogle Scholar
  55. 55.
    Lundberg AS, Weinberg RA (1996) Convergence of mitogenic signalling cascades from diverse classes of receptors at the cyclinD-cyclin-dependent kinase-pRb-controlled G1 checkpoint. Mol Cell Biol 16(12):6917–6925Google Scholar
  56. 56.
    Connell-Crowley L, Harper JW, Goodrich DW (1997) Cyclin D1/Cdk4 regulates retinoblastoma protein-mediated cell cycle arrest by site-specific phosphorylation. Mol Biol Cell 8(2):287–301PubMedGoogle Scholar
  57. 57.
    Lauper N, Amati B et al (1998) Cyclin E2: a novel CDK2 partner in the late G1 and S phases of the mammalian cell cycle. Oncogene 17(20):2637–2643PubMedCrossRefGoogle Scholar
  58. 58.
    Ezhevsky SA, Dowdy SF et al (2001) Differential regulation of retinoblastoma tumor suppressor protein by G (1) cyclin-dependent kinase complexes in vivo. Mol Cell Biol 21(14):4773–4784PubMedCrossRefGoogle Scholar
  59. 59.
    Wells NJ, Hunter T et al (1999) The C-terminal domain of the Cdc2 inhibitory kinase Myt1 interacts with Cdc2 complexes and is required for inhibition of G(2)/M progression. J Cell Sci 112(Pt 19):3361–3371PubMedGoogle Scholar
  60. 60.
    Cheung P, Allis CD, Sassone-Corsi P (2000) Signaling to chromatin through histone modifications. Cell 103(2):263–271PubMedCrossRefGoogle Scholar
  61. 61.
    Jackman M, Pines J et al (2003) Active cyclin B1-Cdk1 first appears on centrosomes in prophase. Nat Cell Biol 5(2):143–148PubMedCrossRefGoogle Scholar
  62. 62.
    Peters JM (2002) The anaphase-promoting complex: proteolysis in mitosis and beyond. Mol Cell 9(5):931–943PubMedCrossRefGoogle Scholar
  63. 63.
    Murray AW (2004) Recycling the cell cycle: cyclins revisited. Cell 116(2):221–234PubMedCrossRefGoogle Scholar
  64. 64.
    Wang D, Siegal GP et al (2008) Immunohistochemistry in the evaluation of neovascularization in tumor xenografts. Biotech Histochem 83(3):179–189PubMedCrossRefGoogle Scholar
  65. 65.
    Nico B, Ribatti D et al (2008) Evaluation of microvascular density in tumors: pro and contra. Histol Histopathol 23(5):601–607PubMedGoogle Scholar
  66. 66.
    Petrelli A, Giordano S (2008) From single- to multi-target drugs in cancer therapy: when aspecificity becomes an advantage. Curr Med Chem 15(5):422–432PubMedCrossRefGoogle Scholar
  67. 67.
    Morphy R, Rankovic Z (2007) Fragments, network biology and designing multiple ligands. Drug Discov Today 12(3–4):156–160PubMedCrossRefGoogle Scholar
  68. 68.
    Hopkins AL (2008) Network pharmacology: the next paradigm in drug discovery. Nat Chem Biol 4(11):682–690PubMedCrossRefGoogle Scholar
  69. 69.
    Morphy R, Rankovic Z (2009) Designing multiple ligands—medicinal chemistry strategies and challenges. Curr Pharm Des 15(6):587–600PubMedCrossRefGoogle Scholar
  70. 70.
    MacKeigan JP, Collins TS, Ting JP (2000) MEK inhibition enhances paclitaxel-induced tumor apoptosis. J Biol Chem 275(50):38953–38956PubMedCrossRefGoogle Scholar
  71. 71.
    Okano J, Rustgi AK (2001) Paclitaxel induces prolonged activation of the Ras/MEK/ERK pathway independently of activating the programmed cell death machinery. J Biol Chem 276(22):19555–19564PubMedCrossRefGoogle Scholar
  72. 72.
    Ling X, Li F et al (2004) Induction of survivin expression by taxol (paclitaxel) is an early event, which is independent of taxol-mediated G2/M arrest. J Biol Chem 279(15):15196–15203PubMedCrossRefGoogle Scholar
  73. 73.
    Yang Y, Chen R et al (2007) p38 and JNK MAPK, but not ERK1/2 MAPK, play important role in colchicine-induced cortical neurons apoptosis. Eur J Pharmacol 576(1–3):26–33PubMedCrossRefGoogle Scholar
  74. 74.
    Abrams SL, McCubrey JA (2010) Enhancing therapeutic efficacy by targeting non-oncogene addicted cells with combinations of signal transduction inhibitors and chemotherapy. Cell Cycle 9(9):1839–1846PubMedCrossRefGoogle Scholar
  75. 75.
    Carlier MF, Chen Y et al (1987) Synchronous oscillations in microtubule polymerization. Proc Natl Acad Sci USA 84(15):5257–5261PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Jin Zhang
    • 1
  • Jing Zhou
    • 1
  • Xiaomei Ren
    • 1
  • Yanyan Diao
    • 2
  • Honglin Li
    • 2
  • Hualiang Jiang
    • 2
  • Ke Ding
    • 1
  • Duanqing Pei
    • 1
  1. 1.Key Laboratory of Regenerative Biology and Institute of Chemical Biology, Guangzhou Institute of Biomedicine and HealthChinese Academy of SciencesGuangzhouChina
  2. 2.School of PharmacyEast China University of Science and TechnologyShanghaiChina

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