Allosteric interactions between the myristate- and ATP-site of the Abl kinase.
Abl kinase inhibitors targeting the ATP binding pocket are currently employed as potent anti-leukemogenic agents but drug resistance has become a significant clinical limitation. Recently, a compound that binds to the myristate pocket of Abl (GNF-5) was shown to act cooperatively with nilotinib, an ATP-competitive inhibitor to target the recalcitrant "T315I" gatekeeper mutant of Bcr-Abl. To uncover an explanation for how drug binding at a distance from the kinase active site could lead to inhibition and how inhibitors could combine their effects, hydrogen exchange mass spectrometry (HX MS) was employed to monitor conformational effects in the presence of both dasatinib, a clinically approved ATP-site inhibitor, and GNF-5. While dasatinib binding to wild type Abl clearly influenced Abl conformation, no binding was detected between dasatinib and T315I. GNF-5, however, elicited the same conformational changes in both wild type and T315I, including changes to dynamics within the ATP site located approximately 25 Å from the site of GNF-5 interaction. Simultaneous binding of dasatinib and GNF-5 to T315I caused conformational and/or dynamics changes in Abl such that effects of dasatinib on T315I were the same as when it bound to wild type Abl. These results provide strong biophysical evidence that allosteric interactions play a role in Abl kinase downregulation and that targeting sites outside the ATP binding site can provide an important pharmacological tool to overcome mutations that cause resistance to ATP-competitive inhibitors.
Department of Chemistry & Chemical Biology, and The Barnett Institute for Chemical and Biological Analysis, Northeastern University, Boston, Massachusetts, United States of America.
Roxana E
RE
Iacob
Jianming
J
Zhang
Nathanael S
NS
Gray
John R
JR
Engen
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000494:Allosteric Regulation; D000970:Antineoplastic Agents / Q000494:pharmacology; D001665:Binding Sites; D000069439:Dasatinib; D006801:Humans; D013058:Mass Spectrometry; D019814:Myristic Acid / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D016315:Proto-Oncogene Proteins c-abl / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D011743:Pyrimidines / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology
D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D013844:Thiazoles; D019814:Myristic Acid; D000255:Adenosine Triphosphate; D016315:Proto-Oncogene Proteins c-abl; C498826:nilotinib; D000069439:Dasatinib
Nature. 2010 Jan 28;463(7280):501-6
20072125
Blood. 2010 May 27;115(21):4206-16
20299508
Cancer Res. 2006 Dec 1;66(23):11314-22
17114238
Nat Rev Cancer. 2009 Jan;9(1):28-39
19104514
Nat Struct Mol Biol. 2008 Oct;15(10):1109-18
18794843
Nat Med. 1996 May;2(5):561-6
8616716
Nat Chem Biol. 2006 Feb;2(2):95-102
16415863
Curr Top Med Chem. 2008;8(10):905-21
18673174
Nat Rev Drug Discov. 2007 Oct;6(10):834-48
17853901
Science. 2001 Aug 3;293(5531):876-80
11423618
Cancer Res. 2007 Sep 1;67(17):7987-90
17804707
Nat Struct Mol Biol. 2004 Dec;11(12):1192-7
15543157
J Am Soc Mass Spectrom. 2006 Dec;17(12):1700-3
16931036
Q Rev Biophys. 1983 Nov;16(4):521-655
6204354
Mol Cancer Ther. 2010 Jul;9(7):1956-67
20571069
N Engl J Med. 2006 Dec 7;355(23):2408-17
17151364
Science. 2004 Jul 16;305(5682):399-401
15256671
Biopolymers. 2007 Mar;85(4):333-48
17167796
Cancer Cell. 2005 Feb;7(2):129-41
15710326
Chem Biol Drug Des. 2007 Sep;70(3):171-81
17718712
Cell. 2003 Mar 21;112(6):831-43
12654249
Cancer Cell. 2009 Nov 6;16(5):401-12
19878872
J Med Chem. 2006 Nov 16;49(23):6819-32
17154512
J Biol Chem. 1951 Mar;189(1):123-36
14832224
Cancer Res. 2006 Jun 1;66(11):5790-7
16740718
Cancer Cell. 2002 Aug;2(2):117-25
12204532
Nat Rev Cancer. 2005 Mar;5(3):172-83
15719031
J Biol Chem. 2009 Oct 16;284(42):29005-14
19679652
Proc Natl Acad Sci U S A. 2009 Feb 3;106(5):1386-91
19164531
Proc Natl Acad Sci U S A. 1987 Dec;84(23):8200-4
3317402
Rapid Commun Mass Spectrom. 2006;20(13):1989-94
16755610
Curr Opin Oncol. 2006 Nov;18(6):578-83
16988578
Curr Mol Med. 2005 Nov;5(7):615-23
16305488
Anal Chem. 2008 Sep 1;80(17):6815-20
18672890
J Med Chem. 2009 Aug 13;52(15):4743-56
19572547
Mass Spectrom Rev. 2006 Jan-Feb;25(1):158-70
16208684
Proc Natl Acad Sci U S A. 2008 Apr 8;105(14):5507-12
18367669
Curr Protoc Protein Sci. 2009 Nov;Chapter 17:Unit 17.6.1-17
19937720
J Biol Chem. 2008 Jun 27;283(26):18292-302
18434310
Anal Chem. 1996 Jul 15;68(14):2340-4
8686927
Protein Sci. 2005 Dec;14(12):3135-9
16260764
Mol Cell. 2006 Mar 17;21(6):787-98
16543148
Cancer Res. 2006 Jan 15;66(2):1007-14
16424036
Cancer Res. 2009 Mar 15;69(6):2384-92
19276351
Nat Rev Drug Discov. 2009 Sep;8(9):709-23
19629074
J Med Chem. 2004 Dec 30;47(27):6658-61
15615512