Protein Kinases in the Era of Precision Medicine

Abstract

Precision medicine signals the beginning of an era in which each patient is a distinct individual from a diagnostic, treatment and health management perspective. In this article, we present a description of a methodology to develop a therapy, based on kinase inhibitors, and a strategy to retarget a therapy in rapid response to a patient's relapse. The idea is to translate the genomic data of an evolving cancer into a three‐dimensional structure to enable a better understanding of the mechanism of resistance, and to then control it with a specific and precise drug targeting therapy based on the SNP resistance mutation. In many cases, a combination of approved drug treatments can assist in controlling the disease. However, the clinical trials selection process is changing from randomised populations to selected populations so that genetic tests can more clearly define the presence of specific mutations in the relapsing patient. This knowledge of the subpopulation will also help identify new small molecules that target multiple kinases based on their specificity profiles. The tendency at present is to define an eligibility criteria based on the genetics of the patients.

Key Concepts

  • The crystallographic characterisation of kinase inhibitor complexes drove the approval of 27 drugs targeting kinome.
  • Organising the kinase three‐dimensional (3D) structures in a uniquely aligned library has enabled us to predict the functionality, in a 3D space, of any variants that are found in a patient's genome.
  • Accurate specificity profiles of approved kinase inhibitors inform a rational combination of inhibitors for a specific genetic subpopulation of patients.
  • Kinase inhibitor therapy can be matched to critical mutations in the kinase 3D structure and a companion diagnostics approach can then be used in specific clinical trial to more accurately select patients.
  • The targeting of both the kinase‐activating mutation and the drug‐resistant mutations can be optimised by the chemistry of the intermediate conformations of the DFG motif.
  • The drug resistance requires frequent genetic monitoring utilising a liquid biopsy.
  • Overcoming the drug resistance on the target is possible by combining approved drugs based on the enhanced knowledge of the highly accurate specificity profiles.

Keywords: kinase; cancer; drug design; drug resistance; Gleevec®; activating mutations; predictive personalised medicine

Figure 1. (a) A sample of the crystal library alignment of 10 kinases. All the 2,147 kinase structures in the library have been uniquely aligned to the conserved ATP‐binding site based on the solved structure of protein kinase A (PKA PDB: 1ATP). Kinases were aligned to PKA because this structure represents the most accurate catalytic description of the in‐line phosphotransfer signalling mechanism. The nine kinases aligned to PKA in this figure are ABL, EGFR, KIT, ERBB2, VEGF2, JAK2, ALK2, RET and BTK, which are the subject of common mutations in cancer, (PDB: 2GLT, 2RGP, 2G0E, 3PP0, 1Y6B, 3IOK, 3H9R, 2IVS, 3OCS, respectively). (b) A unique stabilisation pattern is obtained through a measuring of the peptide backbone RMSD versus the library template sequence (right panel) for Stauroporine (bound to 26 different protein kinases), Imatinib (bound to six different protein kinases, and dasatinib (bound to four different protein kinases). Only the upper beta strand domain is presented in the plots.
Figure 2. (a) The surface of protein kinase A bound to ATP and the respective activation loop (grey) (PDB: 1ATP) has been aligned to Aurora A kinase bound to inhibitors Bisanilinopyrimidine (yellow) and Cl‐Bisanilinopyrimidine (blue) (PDB: 3UO5 and 3UO6 respectively). The chemical difference in inhibitors dictates the conformational change between DFG in/out. (b) The surface of protein kinase A bound to ATP and the respective activation loop (grey) (PDB: 1ATP) has been aligned to ABL1 kinase bound to inhibitors dasatinib (cyan) and imatinib (green) (PDB: 2GQG and 3PYY respectively). There are clear differences observed in both the DFG motif which aspartate is responsible for coordinating the metal sites and the position of the activation loop which is critical for protein substrate positioning. Dasatinib (cyan) binds with the activation loop in a similar position as the ATP, DFG‐in. Imatinib (green), however, binds with the activation loop out preventing substrate positioning, DFG‐out. (c) Comparing the RMSD for each the Aurora A kinases bound with Bisanilinopyrimidine (yellow) and Cl‐Bisanilinopyrimidine (blue) aligned to 1ATP clearly demonstrates a quantitative difference in the orientation of activation loop. The other small variations are addressed to the binding of the ligand to specific key points of the receptor. (d) Comparing the RMSD for each the ABL kinases bound with dasatinib (cyan) and imatinib (green) aligned to 1ATP clearly demonstrates a quantitative difference in the orientation of activation loop. The other small variations are addressed to the binding of the ligand to specific key points of the receptor.
Figure 3. (a) Structure showing ABL1 (cyan surface) co‐crystallised with imatinib (PDB ID: 2HYY). Imatinib in stick format aligned to the other three drugs (alignment to 1ATP using DNA SEQ Alliance crystal library). Red loop is the hinge and blue loop is the DFG activation loop in conformation out. (b) Structure showing ABL1 (cyan surface) co‐crystallised with Nilotinib (PDB ID: 3CS9). Nilotinib in stick format aligned to the other three drugs (alignment to 1ATP using DNA SEQ Alliance crystal library). Red loop is the hinge and blue loop is the DFG activation loop in conformation out. (c) Structure showing ABL1 (cyan surface) co‐crystallised with Dasatinib (PDB ID: 2GQG). Dasatinib in stick format aligned to the other three drugs (alignment to 1ATP using DNA SEQ Alliance crystal library). Red loop is the hinge and blue loop is the DFG activation loop in conformation in. (d) Structure showing ABL1 (cyan surface) co‐crystallised with Bosutinib (PDB ID: 3UE4). Bosutinib in stick format aligned to the other three drugs (alignment to 1ATP using DNA SEQ Alliance crystal library). Red loop is the hinge and blue loop is the DFG activation loop in conformation in. (e) Zoom in on all four drugs aligned with detail on the hydrogen bond connection to the T315 hinge residue usually mutated in cancer resistance. Bosutinib has a different chemical architecture that helps in maintaining the hydrogen bonding or the binding reorganising of its aromatic moiety (highlighted in the square).
Figure 4. (a) PKA (PDB: 1ATP: Science Knighton et al.) in DFG‐IN conformation with ATP and substrate bounded via hydrogen bond; (b) PKA (PDB: 1JBP Madhusudan et al., ) after phosphate is transferred to the substrate, ADP is co‐crystalised as a product of the phosphotransfer. The conformation is in DFG‐IN; however, there are high‐temperature factors present in the upper domain. (c) We represent a conformation of PKA (PDB: 1CTP, see Karlsson et al., ), the upper domain is rotated 15 degrees up and the helix‐C rotates 12 degrees into an open conformation allowing ATP to bind, with the substrate bound to the protein in respect to 1ATP. (d) AurA (PDB: 3UO6, see Martin et al., ) in DFG‐OUT conformation which is incapable of ATP binding. The model is shown with the gatekeeper region, C‐helix, DFG‐activation motif, and upper lobe. These regions identified in the model have also been shown with a cartoon representation as well. (e) This is a model that represents an activating mutation present in the activation loop. The activating mutation causes a loss of regulation of the upper domain and DFG motif, thus is always active. That being the case, the phosotransfer is only dependent on the presence of the substrate. (f) Model shows combined resistance and activating mutation in the kinase.
Figure 5. The figure demonstrates the specificity profiles of (a) dasatinib, (b) imatinib, (c) bosutinib and (d) nilotinib from AMBIT in 2011. Dasatinib and bosutinib both bind in DFG‐in states and thus have a much broader target profile than nilotinib and imatinib which bind in DFG‐out.
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Moiani, Davide, Tabaian, Amir, Suto, Robert K, Doyle, David M, and Sowadski, Janusz M(Feb 2015) Protein Kinases in the Era of Precision Medicine. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000659.pub3]