Benzimidazolyl-pyrazolo[3,4-]pyridinones, Selective Inhibitors of MOLT-4 Leukemia Cell Growth and Sea Urchin Embryo Spiculogenesis: Target Quest.

1,3-Substituted pyrazolo[3,4-]pyridinones - were synthesized by a three-component condensation of Meldrum's acid with aryl aldehydes and 1,3-substituted 5-aminopyrazoles. Their biological activity was evaluated using the phenotypic sea urchin embryo assay and the cytotoxicity screen against human cancer cell lines. In the sea urchin embryo model, 1-benzimidazolyl-pyrazolo[3,4-]pyridinones caused inhibition of hatching and spiculogenesis at sub-micromolar concentrations. These compounds also selectively and potently inhibited growth of the MOLT-4 leukemia cell line. Subsequent structure-activity relationship studies determined the benzimidazolyl fragment as an essential pharmacophore for both effects. We applied numerous techniques for target identification. A preliminary QSAR target identification search did not result in tangible leads. Attempts to prepare a relevant photoaffinity probe that retained potency in both assays were not successful. Compounds were further characterized for their activity in a wild-type versus Notch-mutant leukemia cell lines, and in panels of kinases and matrix metalloproteinases. Using a series of diverse modulators of spiculogenesis as standards, we excluded multiple signaling networks including Notch, Wnt/β-catenin, receptor tyrosine kinases (VEGF/VEGFR, FGF/FGFR), PI3K, and Raf-MEK-ERK as possible targets of . On the other hand, matrix metalloproteinase-9/hatching enzyme was identified as one potential target.