In vitro activity: MELK-8a is a novel and potent MELK inhibitor. MELK kinase has been implicated in playing an important role in tumorigenesis. MELK is involved in the regulation of cell cycle and its genetic depletion leads to growth inhibition in a subset of high MELK-expressing basal-like breast cancer cell lines. MELK inhibitors 8a recapitulate the cellular effects observed by short hairpin ribonucleic acid (shRNA)-mediated MELK knockdown in cellular models. It was found that a novel fluorine-induced hydrophobic collapse that locked the ligand in its bioactive conformation and led to a 20-fold gain in potency. These novel pharmacological inhibitors achieved high exposure in vivo and were well tolerated, which may allow further in vivo evaluation.

Kinase Assay: MELK-8a hydrochloride is a novel maternal embryonic leucine zipper kinase (MELK) inhibitor with an IC50 of 2 nM.

Cell Assay: MELK is involved in the regulation of cell cycle and its genetic depletion leads to growth inhibition in a subset of high MELK-expressing basal-like breast cancer cell lines. MDA-MB-468 and MCF7 cells are seeded in growth medium into 96-well plates at 1000 and 4000 cells/well, respectively. Sixteen hours after plating, MELK-8a are added and incubated for 7 days. For each well, ATPLite reagent is added and incubated. Luminescence is measured on an multilabel plate reader