International Journal of Hematology

, Volume 76, Issue 5, pp 427–435 | Cite as

Constitutive Activation of C-kit by the Juxtamembrane but Not the Catalytic Domain Mutations Is Inhibited Selectively by Tyrosine Kinase Inhibitors STI571 and AG1296

  • Shuji Ueda
  • Hirokazu Ikeda
  • Masao Mizuki
  • Jun Ishiko
  • Itaru Matsumura
  • Hirokazu Tanaka
  • Hirohiko Shibayama
  • Hiroyuki Sugahara
  • Emi Takai
  • Xian Zhang
  • Takashi Machii
  • Yuzuru Kanakura
Progress in hematology


The c-kit receptor tyrosine kinase (KIT) is constitutively activated by 2 types of naturally occurring mutations, the Val559→Gly (G559) mutation in the juxtamembrane domain and the Asp814→Val (V814) mutation in the catalytic domain. We evaluated the effects of the tyrosine kinase inhibitors STI571 and AG1296 on BaF3 cells expressing wild-type KIT (KITWT) or activating mutants of KIT (KITG559 and KITV814) in the presence or absence of the KIT ligand, stem cell factor (SCF). Both STI571 and AG1296 inhibited SCF-dependent activation of KITWT and SCF-independent activation of KITG559 more efficiently, whereas SCF-independent activation of KITV814 was scarcely affected. Furthermore, both inhibitors inhibited SCF-dependent growth of BaF3-KITWT cells and, with higher potencies, SCF-independent growth of BaF3-KITG559 cells through the induction of apoptosis. In contrast, the inhibitors had little or no effect on SCF-independent growth of BaF3-KITV814 cells or on IL-3-dependent growth of BaF3-Mock cells. These results suggested that both inhibitors may be effective therapeutic agents for oncogenic KIT with the juxtamembrane domain mutation, but not with the catalytic domain mutation, and that the activation mechanism of the catalytic domain mutant KIT is complex and entirely different from that of the wild-type KIT or the juxtamembrane domain mutant KIT.Int J Hematol. 2002; 76: 427-435.

Key words

KIT Activating mutation Juxtamembrane domain Catalytic domain Tyrosine kinase inhibitor 


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  1. 1.
    Qiu FH, Ray P, Brown K, et al. Primary structure of c-kit: relationship with the CSF-1/PDGF receptor kinase family—oncogenic activation of v-kit involves deletion of extracellular domain and C terminus.EMBO J. 1988; 7: 1003–1011.PubMedCentralCrossRefGoogle Scholar
  2. 2.
    Yarden Y, Kuang WJ, Yang-Feng T, et al. Human proto-oncogene c-kit: a new cell surface receptor tyrosine kinase for an unidentified ligand.EMBO J. 1987; 6: 3341–3351.PubMedCentralCrossRefGoogle Scholar
  3. 3.
    Galli SJ, Kitamura Y. Genetically masT-cell-deficient W/Wv and Sl/Sld mice: their value for the analysis of the roles of mast cells in biologic responses in vivo.Am J Pathol. 1987; 127: 191–198.PubMedCentralPubMedGoogle Scholar
  4. 4.
    Isozaki K, Hirota S, Nakama A, et al. Disturbed intestinal movement, bile reflux to the stomach, and deficiency of c-kit-expressing cells in Ws/Ws mutant rats.Gastroenterology. 1995; 109: 456–464.CrossRefPubMedGoogle Scholar
  5. 5.
    Kitamura Y. Heterogeneity of mast cells and phenotypic change between subpopulations.Annu Rev Immunol. 1989; 7: 59–76.CrossRefPubMedGoogle Scholar
  6. 6.
    Russell ES. Hereditary anemias of the mouse: a review for geneticists.Adv Genet. 1979; 20: 357–459.PubMedGoogle Scholar
  7. 7.
    Ueda S, Mizuki M, Ikeda H, et al. Critical roles of c-Kit tyrosine residues 567 and 719 in stem cell factor-induced chemotaxis: contribution of src family kinase and PI3-kinase on calcium mobilization and cell migration.Blood. 2002; 99: 3342–3349.CrossRefPubMedGoogle Scholar
  8. 8.
    Hirota S, Isozaki K, Moriyama Y, et al. Gain-of-function mutations of c-kit in human gastrointestinal stromal tumors.Science. 1998; 279: 577–580.CrossRefPubMedGoogle Scholar
  9. 9.
    Kitayama H, Kanakura Y, Furitsu T, et al. Constitutively activating mutations of c-kit receptor tyrosine kinase confer factor-independent growth and tumorigenicity of factor-dependent hematopoietic cell lines.Blood. 1995; 85: 790–798.PubMedGoogle Scholar
  10. 10.
    Kitayama H, Tsujimura T, Matsumura I, et al. Neoplastic transformation of normal hematopoietic cells by constitutively activating mutations of c-kit receptor tyrosine kinase.Blood. 1996; 88: 995–1004.PubMedGoogle Scholar
  11. 11.
    Furitsu T, Tsujimura T, Tono T, et al. Identification of mutations in the coding sequence of the proto-oncogene c-kit in a human mast cell leukemia cell line causing ligand-independent activation of c-kit product.J Clin Invest. 1993; 92: 1736–1744.CrossRefPubMedCentralPubMedGoogle Scholar
  12. 12.
    Dror Y, Leaker M, Caruana G, Bernstein A, Freedman MH. Mastocytosis cells bearing a c-kit activating point mutation are characterized by hypersensitivity to stem cell factor and increased apoptosis.Br J Haematol. 2000; 108: 729–736.CrossRefPubMedGoogle Scholar
  13. 13.
    Fritsche-Polanz R, Jordan JH, Feix A, et al. Mutation analysis of C-KIT in patients with myelodysplastic syndromes without mastocytosis and cases of systemic mastocytosis.Br J Haematol. 2001; 113: 357–364.CrossRefPubMedGoogle Scholar
  14. 14.
    Longley BJ Jr, Metcalfe DD, Tharp M, et al. Activating and dominant inactivating c-KIT catalytic domain mutations in distinct clinical forms of human mastocytosis.Proc Natl Acad Sci U S A. 1999; 96: 1609–1614.CrossRefPubMedCentralPubMedGoogle Scholar
  15. 15.
    Nagata H, Worobec AS, Oh CK, et al. Identification of a point mutation in the catalytic domain of the protooncogene c-kit in peripheral blood mononuclear cells of patients who have mastocytosis with an associated hematologic disorder.Proc Natl Acad Sci U S A. 1995; 92: 10560–10564.CrossRefPubMedCentralPubMedGoogle Scholar
  16. 16.
    Reinacher-Schick A, Petrasch S, Longley BJ, Teschendorf C, Graeven U, Schmiegel W. c-kit mutation and osteopetrosis-like osteopathy in a patient with systemic mast cell disease.Ann Hematol. 1998; 77: 131–134.CrossRefPubMedGoogle Scholar
  17. 17.
    Sperr WR, Walchshofer S, Horny HP, et al. Systemic mastocytosis associated with acute myeloid leukaemia: report of two cases and detection of the c-kit mutation Asp-816 to Val.Br J Haematol. 1998; 103: 740–749.CrossRefPubMedGoogle Scholar
  18. 18.
    Tsujimura T, Furitsu T, Morimoto M, et al. Ligand-independent activation of c-kit receptor tyrosine kinase in a murine mastocy-toma cell line P-815 generated by a point mutation.Blood. 1994; 83: 2619–2626.PubMedGoogle Scholar
  19. 19.
    Tsujimura T, Furitsu T, Morimoto M, et al. Substitution of an aspartic acid results in constitutive activation of c-kit receptor tyrosine kinase in a rat tumor mast cell line RBL-2H3.Int Arch Allergy Immunol. 1995; 106: 377–385.CrossRefPubMedGoogle Scholar
  20. 20.
    Worobec AS, Semere T, Nagata H, Metcalfe DD. Clinical correlates of the presence of the Asp816Val c-kit mutation in the peripheral blood mononuclear cells of patients with mastocytosis.Cancer. 1998; 83: 2120–2129.CrossRefPubMedGoogle Scholar
  21. 21.
    Tsujimura T, Morimoto M, Hashimoto K, et al. Constitutive activation of c-kit in FMA3 murine mastocytoma cells caused by deletion of seven amino acids at the juxtamembrane domain.Blood. 1996; 87: 273–283.PubMedGoogle Scholar
  22. 22.
    Nakahara M, Isozaki K, Hirota S, et al. A novel gain-of-function mutation of c-kit gene in gastrointestinal stromal tumors.Gas-troenterology. 1998; 115: 1090–1095.CrossRefGoogle Scholar
  23. 23.
    Nishida T, Nakamura J,Taniguchi M, et al. Clinicopathological features of gastric stromal tumors.J Exp Clin Cancer Res. 2000; 19: 417–425.PubMedGoogle Scholar
  24. 24.
    Rubin BP, Singer S, Tsao C, et al. KIT activation is a ubiquitous feature of gastrointestinal stromal tumors.Cancer Res. 2001; 61: 8118–8121.PubMedGoogle Scholar
  25. 25.
    Taniguchi M, Nishida T, Hirota S, et al. Effect of c-kit mutation on prognosis of gastrointestinal stromal tumors.Cancer Res. 1999; 59: 4297–4300.PubMedGoogle Scholar
  26. 26.
    Nishida T, Hirota S, Taniguchi M, et al. Familial gastrointestinal stromal tumours with germline mutation of the KIT gene.Nat Genet. 1998; 19: 323–324.CrossRefPubMedGoogle Scholar
  27. 27.
    Beghini A, Tibiletti MG, Roversi G, et al. Germline mutation in the juxtamembrane domain of the kit gene in a family with gastrointestinal stromal tumors and urticaria pigmentosa.Cancer. 2001; 92: 657–662.CrossRefPubMedGoogle Scholar
  28. 28.
    Maeyama H, Hidaka E, Ota H, et al. Familial gastrointestinal stromal tumor with hyperpigmentation: association with a germline mutation of the c-kit gene.Gastroenterology. 2001; 120: 210–215.CrossRefPubMedGoogle Scholar
  29. 29.
    Druker BJ, Tamura S, Buchdunger E, et al. Effects of a selective inhibitor of the Abl tyrosine kinase on the growth of Bcr-Abl positive cells.Nat Med. 1996; 2: 561–566.CrossRefPubMedCentralPubMedGoogle Scholar
  30. 30.
    Druker BJ, Sawyers CL, Kantarjian H, et al. Activity of a specific inhibitor of the BCR-ABL tyrosine kinase in the blast crisis of chronic myeloid leukemia and acute lymphoblastic leukemia with the Philadelphia chromosome.N Engl J Med. 2001; 344: 1038–1042.CrossRefPubMedGoogle Scholar
  31. 31.
    Druker BJ, Talpaz M, Resta DJ, et al. Efficacy and safety of a specific inhibitor of the BCR-ABL tyrosine kinase in chronic myeloid leukemia.N Engl J Med. 2001; 344: 1031–1037.CrossRefPubMedCentralPubMedGoogle Scholar
  32. 32.
    Buchdunger E, Cioffi CL, Law N, et al. Abl protein-tyrosine kinase inhibitor STI571 inhibits in vitro signal transduction mediated by c-kit and platelet-derived growth factor receptors.J Pharmacol Exp Ther. 2000; 295: 139–145.PubMedGoogle Scholar
  33. 33.
    Kovalenko M, Gazit A, Bohmer A, et al. Selective platelet-derived growth factor receptor kinase blockers reverse sis-transformation.Cancer Res. 1994; 54: 6106–6114.PubMedGoogle Scholar
  34. 34.
    Mizushima S, Nagata S. pEF-BOS, a powerful mammalian expression vector.Nucleic Acids Res. 1990; 18: 5322.CrossRefPubMedCentralPubMedGoogle Scholar
  35. 35.
    Ma Y, Zeng S, Metcalfe DD, et al. The c-KIT mutation causing human mastocytosis is resistant to STI571 and other KIT kinase inhibitors: kinases with enzymatic site mutations show different inhibitor sensitivity profiles than wild-type kinases and those with regulatory-type mutations.Blood. 2002; 99: 1741–1744.CrossRefPubMedGoogle Scholar
  36. 36.
    Joensuu H, Roberts PJ, Sarlomo-Rikala M, et al. Effect of the tyrosine kinase inhibitor STI571 in a patient with a metastatic gastrointestinal stromal tumor.N Engl J Med. 2001; 344: 1052–1056.CrossRefPubMedGoogle Scholar
  37. 37.
    van Oosterom AT, Judson I, Verweij J, et al. Safety and efficacy of imatinib (STI571) in metastatic gastrointestinal stromal tumours: a phase I study.Lancet. 2001; 358: 1421–1423.CrossRefPubMedGoogle Scholar
  38. 38.
    Tuveson DA, Willis NA, Jacks T, et al. STI571 inactivation of the gastrointestinal stromal tumor c-KIT oncoprotein: biological and clinical implications.Oncogene. 2001; 20: 5054–5058.CrossRefPubMedGoogle Scholar
  39. 39.
    Tsujimura T, Hashimoto K, Kitayama H, et al. Activating mutation in the catalytic domain of c-kit elicits hematopoietic transformation by receptor self-association not at the ligand-induced dimerization site.Blood. 1999; 93: 1319–1329.PubMedGoogle Scholar
  40. 40.
    Gorre ME, Mohammed M, Ellwood K, et al. Clinical resistance to STI-571 cancer therapy caused by BCR-ABL gene mutation or amplification.Science. 2001; 293: 876–880.CrossRefPubMedGoogle Scholar
  41. 41.
    Weisberg E, Griffin JD. Mechanism of resistance to the ABL tyrosine kinase inhibitor STI571 in BCR/ABL-transformed hemato-poietic cell lines.Blood. 2000; 95: 3498–3505.PubMedGoogle Scholar
  42. 42.
    von Bubnoff N, Schneller F, Peschel C, Duyster J. BCR-ABL gene mutations in relation to clinical resistance of Philadelphia-chromosome-positive leukaemia to STI571: a prospective study.Lancet. 2002; 359: 487–491.CrossRefGoogle Scholar
  43. 43.
    Kovalenko M, Ronnstrand L, Heldin CH, et al. Phosphorylation site-specific inhibition of platelet-derived growth factor beta-receptor autophosphorylation by the receptor blocking tyrphostin AG1296.Biochemistry. 1997; 36: 6260–6269.CrossRefPubMedGoogle Scholar

Copyright information

© The Japanese Society of Hematology 2002

Authors and Affiliations

  • Shuji Ueda
    • 1
  • Hirokazu Ikeda
    • 1
  • Masao Mizuki
    • 1
  • Jun Ishiko
    • 1
  • Itaru Matsumura
    • 1
  • Hirokazu Tanaka
    • 1
  • Hirohiko Shibayama
    • 1
  • Hiroyuki Sugahara
    • 1
  • Emi Takai
    • 1
  • Xian Zhang
    • 1
  • Takashi Machii
    • 1
  • Yuzuru Kanakura
    • 1
  1. 1.Department of Hematology and OncologyOsaka University Graduate School of MedicineOsakaJapan

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