Combination Effects of Integrin-linked Kinase and Abelson Kinase Inhibition on Aberrant Mitosis and Cell Death in Glioblastoma Cells.

In cancer cells, inhibition of integrin-linked kinase (ILK) increases centrosome declustering causing mitotic arrest and cell death. Yet, not all cancer cells are susceptible to anti-ILK treatment alone. We investigate a combination drug strategy targeting ILK and another oncogenic kinase, Abelson kinase (ABL). Drug-concentration viability assays (i.e., MTT assays) indicate that ILK and ABL inhibitors in combination decreased the viability of glioblastoma cells over the ILK drug QLT-0267 alone. Combination strategies also increased aberrant mitoses and cell death over QLT-0267 alone. This was evident from an increase in mitotic arrest, apoptosis and a sub-G1 peak following FAC analysis. In vitro, ILK and ABL localized to the centrosome and the putative ILK kinase domain was important for this localization. Increased levels of cytosolic ABL are associated with its transformative abilities. ILK inhibitor effects on survival correlated with its ability to decrease cytosolic ABL levels and inhibit ABL's localization to mitotic centrosomes in glioblastoma cells. ILK inhibitor effects on ABL's centrosomal localization were reversed by the proteasomal inhibitor MG132 (a drug that inhibits ABL degradation). These results indicate that ILK regulates ABL at mitotic centrosomes and that combination treatments targeting ILK and ABL are more effective then QLT-0267 alone at decreasing the survival of dividing glioblastoma cells.

Insulin attenuates apoptosis in neuronal cells by an integrin-linked kinase-dependent mechanism.

Insulin promotes neuronal survival by activating a phosphatidylinositol 3-kinase (PI 3-kinase)/AKT-dependent signaling pathway and reducing caspase activation. We investigated a role for integrin-linked kinase (ILK) in insulin-mediated cell survival in cultured neurons and differentiated R28 cells. We used a serum and depolarization withdrawal model to induce apoptosis in cerebellar granule neurons and a serum withdrawal model to induce apoptosis in differentiated R28 cells. ILK knock-out decreased insulin-mediated protection as did the addition of pharmacological inhibitors of ILK, KP-392 or QLT-0267. Prosurvival effects of insulin were rescued by Boc-Asp (O-methyl)-CH2F (BAF), a pancaspase inhibitor, in the presence of KP-392. Insulin and IGF-1 decreased caspase-3 activation, an effect that was inhibited by KP-392 and QLT-0267. Western blot analysis indicates that insulin-induced stimulation of AKT Ser-473 phosphorylation was decreased after the ILK gene was conditionally knocked-out, following overexpression of AKT-DN or in the presence of QLT-0267. Insulin and IGF-1 stimulated ILK kinase activity in primary neurons and this was inhibited following ILK-DN overexpression. Western blot analysis indicates that insulin exposure upregulated the expression of the cellular inhibitor of apoptosis protein c-IAP2 in an extracellular matrix-dependent manner, an effect blocked by KP-392. These results indicate that ILK is an important effector in insulin-mediated neuroprotection.