A combinatorial in silico and cellular approach to identify a new class of compounds that target VEGFR2 receptor tyrosine kinase activity and angiogenesis

Background and purpose: Vascular endothelial growth factor receptor 2 (VEGFR2/KDR/Flk-1) is an attractive therapeutic target for the treatment of diseases such as Cancer. Small-molecule VEGFR2/KDR/Flk-1 inhibitors of a variety of chemical classes are currently under development or in clinical use. In this study, we describe the de novo design of a new generation pyrazole-based molecule (JK-P3) that targets VEGFR2/KDR/Flk-1 kinase activity and angiogenesis.

Experimental approach: JK-P compound series were designed using de novo structure-based identification methods. Compounds were tested in an in vitro VEGFR2/KDR/Flk-1 kinase assay. Using primary endothelial cells, JK-P compounds were assessed for their ability to inhibit VEGF-A-stimulated VEGFR2/KDR/Flk-1 activation and intracellular signalling. We tested these compounds in cell migration, proliferation and angiogenesis assays.

Key results: JK-P3 and JK-P5 were predicted to bind the VEGFR2/KDR/Flk-1 kinase domain with high affinity, and both compounds showed pronounced inhibition of endogenous VEGFR2/KDR/Flk-1 kinase activity in primary human endothelial cells. Only JK-P3 inhibited VEGF-A-stimulated VEGFR2/KDR/Flk-1 activation and intracellular signalling. Interestingly, JK-P3 inhibited endothelial monolayer wound closure and angiogenesis but not endothelial cell proliferation. Both compounds inhibited Fibroblast Growth Factor receptor kinase activity in vitro, but not basic fibroblast growth factor-mediated signalling in endothelial cells.

Conclusions and implications: This is the first report that describes an anti-angiogenic inhibitor based on such a pyrazole core. Using a de novo structure-based identification approach is an attractive method to aid such drug discovery. These results thus provide an important basis for the development of multi-tyrosine kinase inhibitors for clinical use in the near future.