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Structure of the Kinase Domain of an Imatinib-Resistant Abl Mutant in Complex with the Aurora Kinase Inhibitor VX-680

Matthew A. Young, Neil P. Shah, Luke H. Chao, Markus Seeliger, Zdravko V. Milanov, William H. Biggs, III, Daniel K. Treiber, Hitesh K. Patel, Patrick P. Zarrinkar, David J. Lockhart, Charles L. Sawyers and John Kuriyan

Cancer Res 2006; 66(2): 1007-14   Local Copy

Summary / Figures / Table / PDB


Summary: We present a high-resolution (2.0 Å) crystal structure of the catalytic domain of a mutant form of the Abl tyrosine kinase (H396P; Abl-1a numbering) that is resistant to the Abl inhibitor imatinib. The structure is determined in complex with the small-molecule inhibitor VX-680 (Vertex Pharmaceuticals, Cambridge, MA), which blocks the activity of various imatinib-resistant mutant forms of Abl, including one (T315I) that is resistant to both imatinib and BMS-354825 (dasatinib), a dual Src/Abl inhibitor that seems to be clinically effective against all other imatinib-resistant forms of BCR-Abl. VX-680 is shown to have significant inhibitory activity against BCR-Abl bearing the T315I mutation in patient-derived samples. The Abl kinase domain bound to VX-680 is not phosphorylated on the activation loop in the crystal structure but is nevertheless in an active conformation, previously unobserved for Abl and inconsistent with the binding of imatinib. The adoption of an active conformation is most likely the result of synergy between the His396Pro mutation, which destabilizes the inactive conformation required for imatinib binding, and the binding of VX-680, which favors the active conformation through hydrogen bonding and steric effects. VX-680 is bound to Abl in a mode that accommodates the substitution of isoleucine for threonine at residue 315 (the "gatekeeper" position). The avoidance of the innermost cavity of the Abl kinase domain by VX-680 and the specific recognition of the active conformation explain the effectiveness of this compound against mutant forms of BCR-Abl, including those with mutations at the gatekeeper position.


Figures (Click on the small image to view the bigger one):


Figure 1. Structure of VX-680 complex.

  1. structure of Abl kinase domain bound to imatinib (left; PDB code 1OPJ;) and VX-680 (right). Hydrogen atoms are not shown.
  2. chemical structure VX-680 (28) and 2Fo-Fc electron density for VX-680 and the DFG motif contoured at 1.9σ.
  3. mode of binding of VX-680 to the Abl kinase domain. The three hydrogen bonds to the hinge region (yellow dashed lines) and the hydrogen bond to Asp381 (red dashed line; distances in Å).


Figure 2. Comparison of VX-680 complex with other structures.

  1. superposition of the Abl:VX-680 complex (blue) with the structure of the active form of the Lck kinase domain (PDB code 3LCK; ref. 41; orange).
  2. conformation of the activation loop of Abl:VX-680 (blue) superimposed on the structure of the active and phosphorylated activation loop of Lck (3LCK; orange).
  3. active conformation of the DFG loop in the VX-680 complex (blue) compared with active conformation of Lck (3LCK; orange; ref. 41), the inactive conformation in the complex with PD173955 (red; 1M52; ref. 14), and inactive conformation in complex with imatinib (dark blue; 1OPJ; ref. 14).


Figure 3. Mode of binding of VX-680.

  1. extent of burial of VX-680 (left) and imatinib (right).
  2. interactions between the phosphate binding loop (P-loop) of the kinase domain and VX-680 (left) and imatinib (right).
  3. residues that make side chain contact with VX-680 (defined as having any side-chain atom within 5.0 Å of the drug; red spheres). Residues in which only the Cα atom or the backbone make contact with VX-680 are not shown (except for glycine). Residues that are substituted in Aurora-A are indicated by naming the Aurora-A residue in parentheses.
  4. interaction between VX-680 and Aurora-A (yellow; 1MQ4; ref. 43), modeled on the basis of the Abl:VX-680 structure. The position of VX-680 within Aurora was obtained by aligning the two kinase domain on the residues of the hinge region. Leu210 in Aurora-A, located at the gatekeeper position, is shown with spheres for the side-chain atoms. Interaction between VX-680 and Abl (T315I; blue), modeled by substituting Thr315 in the crystal structure with isoleucine.


Figure 4. Inhibition of purified kinase domain of Abl by VX-680 and imatinib.

  1. T315I Abl kinase activity is shown not to be affected by imatinib, whereas VX-680 binds less tightly to wild-type Abl than T315I Abl.
  2. IC50 values for wild-type and T315I Abl for imatinib and VX-680.


Figure 5. VX-680 inhibits BCR-Abl/T315I in primary CML cells. Exposure of peripheral blood mononuclear cells isolated from a patient known to harbor BCR-Abl/T315I to the concentrations of VX-680 or BMS-354825 followed by Western immunoblotting with a CrkL antibody. Migration of phospho-CrkL species.


Figure 6. Two conformations of residue 396: gold-wildtype Abl (H396) crystallized in the inactive conformation; red-the H396P mutant crystallized in the active conformation. Contour plots indicate the distribution of Phi-Psi values for all His (left) and Pro (right) residues found in the PDB. Pro cannot adopt the conformation found in the inactive conformation and the H396P mutation thus destabilizes the inactive conformation.


Table 1. Crystallographic statistics and refinement
PDB: 2F4J.pdb PDB infor at RCSB