New J Chem.2016, 40(10):8296-8304. DOI: 10.1039/C6NJ01909A

Two conformers of a tyrosine kinase inhibitor (AG-1478) disclosed using simulated UV-Vis absorption spectroscopy

Muhammad Khattab,a   Subhojyoti Chatterjee,b   Andrew H. A. Clayton*a and   Feng Wang*b
a Centre for Micro-Photonics, Faculty of Science, Engineering and Technology, Swinburne University of Technology, Melbourne, Australia. E-mail:
b Molecular Model Discovery Laboratory, Department of Chemistry and Biotechnology, School of Science, Faculty of Science, Engineering and Technology, Swinburne University of Technology, Melbourne, Australia. E-mail:
* Corresponding authors
AG-1478 (N-(3-chlorophenyl)-6,7-dimethoxy-4-quinazolinamine) shows promising in vitro and in vivo antiproliferative activity and has gained global interest due to its potent and broad biopharmaceutical activities. An important step towards understanding its spatial and temporal distribution is to determine whether the inhibitors have spectral signatures that might assist in determining the relevant targets and interactions. Its UV-Vis absorption spectra in various solutions have been measured [Khattab et al., Spectrochimica Acta A, 2016, 164, 128]. The present study correlates the UV-Vis spectral signatures with the structure of the drug. Two stable conformers AG-1478B and AG-1478A with close energy values (ΔE = 1.58 kcal mol−1) were located on the potential energy surface through rotation of the single C–N bond of the C–NH–C chain of the drug. The present density functional theory (DFT) study reveals that both conformers contribute to the measured UV-Vis absorption spectrum of AG-1478. The conformers, AG-1478B and AG-1478A, were subjected to further study using molecular orbital theory. It is found that although the conformers are close in energy, the anisotropic properties, such as the shape in three dimensional (3D) space, the dipole moment and the orbitals, are apparently different. The excess orbital energy spectrum (EOES) indicates that six core orbitals exhibit significant conformational changes, exhibiting the signatures of the N atoms, i.e., the NH linker N(25) and the quinazoline N(12). The valence orbitals with significant configurational changes are either due to the local distribution (30a) or delocalization (46a, 76a and 82a (highest occupied molecular orbital (HOMO))).


Cancer arises, in part, from the uncontrolled growth of cells. Alterations to the cells, sometimes through genetic changes, give the cells a growth advantage and allow them to evade normal quality control processes that would prevent the cells from growing abnormally.  For some cancers, such as brain, head, neck and breast cancer, this growth advantage can arise from proteins on the cell surface called receptors. These receptors normally send a growth signal to the cell from other cells during phases of development but in cancer these receptors are overproduced or altered in a way that they send a message for the cell to grow unremittingly. 

An important class of receptors is called tyrosine kinase receptors. Tyrosine kinase receptors work by catalysing the transfer of a phosphate group to a tyrosine residue. The phosphorylated tyrosine then acts as a docking site for intracellular effectors and other enzymes which form a bucket brigade allowing transmission of the growth signal from outside of the cell to inside the cell. Tyrosine kinase inhibitors are a class of drugs which block the action of tyrosine kinases. These normally bind to a pocket inside the receptor preventing the binding of ATP or blocking substrate access.  Although there are a few tyrosine kinase inhibitors in the clinic, they suffer from two problems. The first is they can cause undesirable side effects (skin rash) and the second is that they can lead to drug resistance. Therefore there is a continued development of new tyrosine kinase inhibitors.

Understanding the structure of the tyrosine kinase inhibitor in complex with its tyrosine kinase is very important for drug development. Having a good complementarity of the protein surface to the shape of the drug is important for specific binding to avoid side-effects or allergic reactions. The main technique for such studies is x-ray crystallography which is a highly specialised technique. First, for this approach to work the scientist must be able to form crystals in which the protein and inhibitor are in complex. Second, the scientist must be able to obtain high enough resolution data to discern the conformation of the inhibitor and its surroundings.

We have sought a different approach to determining inhibitor conformation and protein binding site environment which can be performed in solution without crystallization and using relatively simple instrumentation. The approach combines computational chemistry with optical spectroscopy. Candidate conformations of the inhibitor are optimised in a computer and the theoretical optical absorption spectrum of these conformations are computed using quantum chemistry. The experimental conformer can then be determined by measuring the absorption spectrum of the inhibitor in solution and comparing it to the theoretical absorption spectra of the different inhibitor conformers.

Using this approach we have identified two conformations of the tyrosine kinase inhibitor AG1478, in different solvents. 

The next step will be to apply this approach to inhibitor protein complexes. Understanding which conformer is bound in the inhibitor-kinase complex will enable us to design a drug with optimised geometry.






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