EVALUATION OF PROTEIN KINASE INHIBITORS WITH PLK4 CROSS-OVER POTENTIAL IN PEDIATRIC EMBRYONAL BRAIN TUMORS

BACKGROUND: In previous work we established that PLK4 is overexpressed and essential for proliferation and survival of pediatric embryonal brain tumors. PLK4 is a cell cycle-regulated protein recruited at the centrosome in dividing cells. Its overexpression triggers centrosome amplification, which is associated with genetic instability and carcinogenesis. We demonstrated that PLK4 inhibition exerts a potent cytostatic effect in EBT. We also established the synergism of the PLK4 inhibitor CFI-400945 with cytotoxic drugs and other signal transduction modulators. METHODS: We studied protein kinase inhibitors (PKI) with PLK4 cross-over potential (CFI-400945, CFI-400437, centrinone, centrinone-B, R-1530, axitinib, alisertib and KW-2449) by assessing their chemical profiles through docking simulations over PLK4 (PDB entry: 3COK) using Gold 5.4 software; evaluating their potential to penetrate the BBB (via logBB calculation by Clark equation), performing LanthaScreen EU Kinase Binding assays (ThermoFisher Scientific) and cell based studies (viability and proliferation) in multiple embryonal tumor cell lines (MON, BT-12, BT-16, DAOY, D283). RESULTS: Kinase binding assays and cell based studies showed that CFI-400437 was the most effective compound, yielding an IC50 <5nM in the binding assay and requiring <100nM of drug to significantly inhibit proliferation in multiple cell lines. However, docking simulations using the PLK4 catalytic cavity and each of the various PKIs demonstrated that all of the inhibitors tested exhibit similar interactions within the binding cavity, engaging in H-bonding with backbone residues Glu-89 and Cys-91. Additional polar and hydrophobic (π-stacking) interactions were also identified for each of the PKIs. LogBB calculations revealed that compound R-1530 has the greatest likelihood to enter the brain (LogBB+0.130). CONCLUSION: Although CFI-400945 is the most effective PLK4 inhibitor among those assayed, it has low potential for BBB exposure. Molecular docking results indicate the potential to develop new chemical entities with higher BBB exposure potential using a similar binding mode to that of CFI-400945.