title: Tyrosine kinase inhibitors induce down-regulation of c-Kit by targeting the ATP pocket.
abstract: The stem cell factor receptor (SCF) c-Kit plays a pivotal role in regulating cell proliferation and survival in many cell types. In particular, c-Kit is required for early amplification of erythroid progenitors, while it must disappear from cell surface for the cell entering the final steps of maturation in an erythropoietin-dependent manner. We initially observed that imatinib (IM), an inhibitor targeting the tyrosine kinase activity of c-Kit concomitantly down-regulated the expression of c-Kit and accelerated the Epo-driven differentiation of erythroblasts in the absence of SCF. We investigated the mechanism by which IM or related masitinib (MA) induce c-Kit down-regulation in the human UT-7/Epo cell line. We found that the down-regulation of c-Kit in the presence of IM or MA was inhibited by a pre-incubation with methyl-β-cyclodextrin suggesting that c-Kit was internalized in the absence of ligand. By contrast to SCF, the internalization induced by TKI was independent of the E3 ubiquitin ligase c-Cbl. Furthermore, c-Kit was degraded through lysosomal, but not proteasomal pathway. In pulse-chase experiments, IM did not modulate c-Kit synthesis or maturation. Analysis of phosphotyrosine peptides in UT-7/Epo cells treated or not with IM show that IM did not modify overall tyrosine phosphorylation in these cells. Furthermore, we showed that a T670I mutation preventing the full access of IM to the ATP binding pocket, did not allow the internalization process in the presence of IM. Altogether these data show that TKI-induced internalization of c-Kit is linked to a modification of the integrity of ATP binding pocket.
authors: Institut Cochin, Département d'Immunologie-Hématologie, Paris, France.
authors: Diane
authors: D
authors: D'allard
authors: Julie
authors: J
authors: Gay
authors: Clotilde
authors: C
authors: Descarpentries
authors: Emilie
authors: E
authors: Frisan
authors: Kevin
authors: K
authors: Adam
authors: Frederique
authors: F
authors: Verdier
authors: Célia
authors: C
authors: Floquet
authors: Patrice
authors: P
authors: Dubreuil
authors: Catherine
authors: C
authors: Lacombe
authors: Michaela
authors: M
authors: Fontenay
authors: Patrick
authors: P
authors: Mayeux
authors: Olivier
authors: O
authors: Kosmider
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D001549:Benzamides / Q000494:pharmacology*; D001665:Binding Sites / Q000187:drug effects; D002460:Cell Line; D015536:Down-Regulation / Q000187:drug effects*; D004900:Erythroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D000068877:Imatinib Mesylate; D008024:Ligands; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D010879:Piperazines / Q000494:pharmacology*; D010880:Piperidines; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D021381:Protein Transport / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D059748:Proteolysis / Q000187:drug effects; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism*; D011725:Pyridines; D011743:Pyrimidines / Q000494:pharmacology*; D013844:Thiazoles / Q000494:pharmacology*
chemical_list: D001549:Benzamides; D008024:Ligands; D010879:Piperazines; D010880:Piperidines; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D011743:Pyrimidines; D013844:Thiazoles; D000068877:Imatinib Mesylate; D000255:Adenosine Triphosphate; D011505:Protein-Tyrosine Kinases; D019009:Proto-Oncogene Proteins c-kit; C526575:masitinib
references: Mol Biol Cell. 1999 Apr;10(4):961-74
references: 10198050
references: Clin Genitourin Cancer. 2013 Jun;11(2):134-40
references: 23058498
references: Blood. 2005 Jan 1;105(1):226-32
references: 15315962
references: Biochem J. 2006 Oct 1;399(1):59-67
references: 16780420
references: J Clin Oncol. 2002 Mar 15;20(6):1692-703
references: 11896121
references: Physiol Rev. 2012 Oct;92(4):1619-49
references: 23073628
references: Blood. 2010 Aug 19;116(7):1114-23
references: 20484085
references: Blood. 2012 Feb 9;119(6):1532-42
references: 22160620
references: Mol Cell Proteomics. 2002 May;1(5):376-86
references: 12118079
references: PLoS One. 2009 Sep 30;4(9):e7258
references: 19789626
references: J Biol Chem. 2003 Oct 24;278(43):42457-65
references: 12930839
references: Blood. 1999 Sep 15;94(6):1979-86
references: 10477727
references: Mol Cancer Ther. 2012 Aug;11(8):1770-80
references: 22665524
references: Clin Cancer Res. 2007 Aug 15;13(16):4874-81
references: 17699867
references: J Biol Chem. 2004 Jul 23;279(30):31655-63
references: 15123710
references: EMBO Rep. 2005 Oct;6(10):942-8
references: 16113650
references: Blood. 2009 Nov 5;114(19):4197-208
references: 19734451
references: Cell. 1999 Mar 5;96(5):677-87
references: 10089883
references: Blood. 1995 Jul 15;86(2):572-80
references: 7541668
references: Oncogene. 2003 Feb 6;22(5):660-4
references: 12569358
references: Oncogene. 2006 Oct 5;25(45):6140-6
references: 16751810
references: Cancer Cell. 2005 Dec;8(6):467-78
references: 16338660
references: Nat Biotechnol. 2008 Dec;26(12):1367-72
references: 19029910
references: Proc Natl Acad Sci U S A. 2012 Aug 21;109(34):E2276-83
references: 22652566
references: J Biol Chem. 2000 Jun 16;275(24):18375-81
references: 10849444
references: J Biol Chem. 1994 Dec 16;269(50):31991-8
references: 7527401
references: Blood. 2005 Jul 15;106(2):721-4
references: 15790786
references: J Cell Physiol. 1988 Nov;137(2):293-8
references: 3263970
references: Proc Natl Acad Sci U S A. 1999 Jun 8;96(12):6775-80
references: 10359788
references: Exp Cell Res. 2007 Nov 1;313(18):3935-42
references: 17904548
references: Blood. 2010 Oct 14;116(15):2665-75
references: 20595514
references: Blood. 2005 Jan 15;105(2):600-8
references: 15358619
references: Cancer Res. 1991 Jan 1;51(1):341-8
references: 1824823
references: J Biol Chem. 1999 Aug 6;274(32):22151-4
references: 10428778
references: Gastroenterology. 2004 Jul;127(1):294-9
references: 15236194
references: Blood. 1995 Feb 1;85(3):790-8
references: 7530509
references: Cell. 2007 Jul 27;130(2):323-34
references: 17662946
references: Cancer Res. 2007 Aug 15;67(16):7695-702
references: 17699773