Activation of tyrosine kinases by mutation of the gatekeeper threonine.
Protein kinases targeted by small-molecule inhibitors develop resistance through mutation of the 'gatekeeper' threonine residue of the active site. Here we show that the gatekeeper mutation in the cellular forms of c-ABL, c-SRC, platelet-derived growth factor receptor-alpha and -beta, and epidermal growth factor receptor activates the kinase and promotes malignant transformation of BaF3 cells. Structural analysis reveals that a network of hydrophobic interactions-the hydrophobic spine-characteristic of the active kinase conformation is stabilized by the gatekeeper substitution. Substitution of glycine for the residues constituting the spine disrupts the hydrophobic connectivity and inactivates the kinase. Furthermore, a small-molecule inhibitor that maximizes complementarity with the dismantled spine (compound 14) inhibits the gatekeeper mutation of BCR-ABL-T315I. These results demonstrate that mutation of the gatekeeper threonine is a common mechanism of activation for tyrosine kinases and provide structural insights to guide the development of next-generation inhibitors.
Azam
Mohammad
M
Karp research building, 7th floor, Division of Pediatric Hematology/Oncology, Children's Hospital of Boston, Massachusetts 02115, USA.
Seeliger
Markus A
MA
Gray
Nathanael S
NS
Kuriyan
John
J
Daley
George Q
GQ
D001665:Binding Sites; D000081247:CSK Tyrosine-Protein Kinase; D002460:Cell Line; D018360:Crystallography, X-Ray; D004789:Enzyme Activation / Q000235:genetics; D066246:ErbB Receptors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics*; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016315:Proto-Oncogene Proteins c-abl / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017479:Receptors, Platelet-Derived Growth Factor / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013912:Threonine / Q000235:genetics* / Q000378:metabolism*; D019061:src-Family Kinases
D013912:Threonine; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D017479:Receptors, Platelet-Derived Growth Factor; D000081247:CSK Tyrosine-Protein Kinase; D016315:Proto-Oncogene Proteins c-abl; D019061:src-Family Kinases; C000631346:CSK protein, human
Science. 2001 Aug 3;293(5531):876-80
11423618
Methods Enzymol. 1997;276:307-26
27754618
Cancer Cell. 2005 Apr;7(4):387-97
15837627
Nat Genet. 2005 Dec;37(12):1315-6
16258541
Proc Natl Acad Sci U S A. 2006 Dec 19;103(51):19466-71
17164333
Structure. 2007 Mar;15(3):299-311
17355866
J Exp Med. 1999 May 3;189(9):1399-412
10224280
Mol Cell Biol. 1986 Nov;6(11):4133-5
3025637
Cancer Res. 2005 Jun 1;65(11):4500-5
15930265
Science. 2000 Sep 15;289(5486):1938-42
10988075
Acta Crystallogr D Biol Crystallogr. 2005 Apr;61(Pt 4):458-64
15805601
Proc Natl Acad Sci U S A. 2008 Feb 12;105(6):2070-5
18227510
N Engl J Med. 2003 Mar 27;348(13):1201-14
12660384
Chem Biol. 2006 Jul;13(7):779-86
16873026
Nat Rev Drug Discov. 2004 Dec;3(12):1001-10
15573099
Proc Natl Acad Sci U S A. 1996 Dec 10;93(25):14845-50
8962143
Proc Natl Acad Sci U S A. 2006 Nov 21;103(47):17783-8
17095602
Cancer Cell. 2003 May;3(5):459-69
12781364
Cancer Res. 2005 Oct 1;65(19):8968-74
16204070
Proc Natl Acad Sci U S A. 2006 Jun 13;103(24):9244-9
16754879
Proc Natl Acad Sci U S A. 2006 Nov 28;103(48):18101-6
17114285
Mol Diagn Ther. 2006;10(2):67-76
16669605
Cell. 2003 Mar 21;112(6):831-43
12654249
Oncogene. 2004 Feb 12;23(6):1214-20
14647454
Cell. 1987 Apr 10;49(1):1-4
3030562
J Biol Chem. 2002 Aug 30;277(35):32214-9
12077114
Blood. 2000 Dec 1;96(12):3907-14
11090077
Cell. 2003 Mar 21;112(6):737-40
12654240
Acta Crystallogr D Biol Crystallogr. 1997 May 1;53(Pt 3):240-55
15299926
Cancer Cell. 2002 Aug;2(2):117-25
12204532
J Clin Oncol. 2002 Mar 15;20(6):1692-703
11896121
Cell. 2006 Jun 16;125(6):1137-49
16777603
Nat Rev Cancer. 2004 Sep;4(9):718-27
15343278
Cell. 2003 Mar 21;112(6):845-57
12654250
Proc Natl Acad Sci U S A. 1988 Dec;85(23):9312-6
3143116
Acta Crystallogr D Biol Crystallogr. 2005 Jul;61(Pt 7):850-5
15983406
Nature. 2004 Nov 18;432(7015):294-7
15549090
Nature. 1997 Feb 13;385(6617):602-9
9024658
N Engl J Med. 2005 Feb 24;352(8):786-92
15728811
Genes Dev. 2003 Dec 15;17(24):2998-3010
14701871
Nat Rev Drug Discov. 2002 Apr;1(4):309-15
12120282
Science. 2004 Mar 19;303(5665):1800-5
15031492
Cell. 2002 May 3;109(3):275-82
12015977
Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32
15572765
Mol Cell Biol. 1986 Dec;6(12):4155-60
2432397
Mol Cell Biol. 2006 Aug;26(16):6082-93
16880519
Cell. 2003 Mar 21;112(6):859-71
12654251
Blood. 2007 Jul 15;110(2):727-34
17405907
Nature. 1997 Feb 13;385(6617):595-602
9024657
Bioorg Med Chem. 1998 Aug;6(8):1219-26
9784863
Nat Med. 1996 May;2(5):561-6
8616716
Gastroenterology. 2004 Jul;127(1):294-9
15236194
Cell. 1983 Mar;32(3):881-90
6299580
Nature. 1997 Feb 13;385(6617):650-3
9024665
Cancer Cell. 2007 Mar;11(3):217-27
17349580
PLoS Biol. 2006 May;4(5):e144
16640460
J Pharmacol Exp Ther. 2000 Oct;295(1):139-45
10991971