IN SILICO AND SPECTROSCOPIC INVESTIGATION OF THE INTERACTION BETWEEN ABL KINASE AND ANTILEUKEMIC INDOLES

Introduction: Chronic Myeloid Leukemia (CML) is primarily caused by the fusion of AB kinase and BCR genes in the Philadelphia chromosome. ABL kinase catalyzes tyrosine phosphorylation of peptides/proteins. Transcription of the ABL-BCR fusion gene produces a hyperactive protein that disrupts cellular metabolism, including apoptosis regulation. Consequently, the inhibition of ABL kinase constitutes the main therapeutic approach for CML. It has been shown that indole-based and structurally related heterocyclic compounds interact with ABL kinase, eliciting a biological response in cellular assays (Med Chem Res 2019, 28(5), 633). In this work, we characterized the interaction of indole derivatives actives in cellular assays, synthesized by our group, with ABL kinase, using in silico and NMR spectroscopy. Materials and Methods: a) in silico studies: Semi-flexible molecular docking protocols were applied and validated by analyzing root mean square deviation (RMSD) values in redocking and crossdocking simulations with imatinib and nilotinib used as ligands. The drugs and the indoles derivatives were docked in ABL kinase using the software Vina 1.2. The analysis of the indole derivatives-ABL docking simulations was carried out by visually inspecting the ligand binding modes, comparing the docking energy score and the ligand-ABL formation enthalpy (\Delta H_f°), and identifying the compound's interaction profile. The \Delta H_f° in each ABL-ligand complex was calculated with the software Molpac using the PM7 semi-empirical method; b) NMR spectroscopy: ABL containing the SH2, SH3, and kinase domains, was produced by heterologous expression in E. coli BL21DE3 cells and purified by affinity and size exclusion chromatography; ABL kinase structure was inspected by 1H 1D NMR experiments and the indole derivatives interaction with ABL kinase was monitored by 1H 1D saturation transfer difference NMR experiments (STD-NMR). Results: The docking scores and \Delta H_f° indicated thermodynamically favorable ligand-ABL complexes. The indoles occupied the same region of the pyridine and pyrimidine rings of nilotinib, forming hydrogen bonds with ABL residues GLU266 and ASP361. STD-NMR experiments confirmed the interactions of indole derivatives with the ABL kinase with a dissociation constant, Kd, within the micro to millimolar range. Interestingly, for two compounds the interaction with ABL was just seen in 1H 1D NMR spectra, indicating sub-micromolar Kd. Further NMR experiments are being performed to identify ligand binding sites on ABL kinase structure. Conclusion: The results presented in this work suggest the potential ABL inhibition by indole-based compounds and could contribute to the development of optimized ABL inhibitor candidates by optimization strategies based on structural information of the molecular target.

This work was supported by CNPq, FAPEMIG, and CAPES.