Ectopic expression of the neuron-specific inositol-1,4,5-trisphosphate-3-kinase A (ITPKA) in lung cancer cells increases their metastatic potential because the protein exhibits two actin regulating activities; it bundles actin filaments and regulates inositol-1,4,5-trisphosphate (InsP(3))-mediated calcium signals by phosphorylating InsP(3). Thus, in order to inhibit the metastasis-promoting activity of ITPKA, both its actin bundling and its InsP(3)kinase activity has to be blocked. In this study, we performed a high throughput screen in order to identify specific and membrane-permeable substances against the InsP(3)kinase activity. Among 341,44 small molecules, 237 compounds (0.7%) were identified as potential InsP(3)kinase inhibitors. After determination of IC50-values, the three compounds with highest specificity and highest hydrophobicity (EPPC-3, BAMB-4, MEPTT-3) were further characterized. Only BAMB-4 was nearly completely taken up by H1299 cells and remained stable after cellular uptake, thus exhibiting a robust stability and a high membrane permeability. Determination of the inhibitor type revealed that BAMB-4 belongs to the group of mixed type inhibitors. Taken together, for the first time we identified a highly membrane-permeable inhibitor against the InsP(3)kinase activity of ITPKA providing the possibility to partly inhibit the metastasis-promoting effect of ITPKA in lung tumor cells. (C) 2013 Elsevier Inc. All rights reserved.