Structural Analysis Identifies Imidazo[1,2-b]Pyridazines as PIM Kinase Inhibitors with In vitro Antileukemic Activity

Abstract

Much attention has recently been focused on PIM kinases as potential targets for the treatment of hematopoietic malignancies and some solid cancers. Using protein stability shift assays, we identified a family of imidazo [1,2-b]pyridazines to specifically interact with and inhibit PIM kinases with low nanomolar potency. The high-resolution crystal structure of a PIM1 inhibitor complex revealed that imidazo[1,2-b]pyrridazines surprisingly interact with the NH 2 -terminal lobe helix AC rather than with the kinase hinge region. Thus, the identified inhibitors are ATP competitive but not ATP mimetic compounds, explaining their enhanced selectivity with respect to conventional type I kinase inhibitors. One of the identified imidazo[1,2-b]pyridazines (K00135) was further tested in several hematopoietic cellular systems. First, K00135 dosedependently impaired survival of murine Ba/F3 cells that have been rendered cytokine independent by overexpression of human PIMs. Second, K00135 impaired survival and clonogenic growth of a panel of human acute leukemia cells. Third, exposure of K00135 significantly suppressed in vitro growth of leukemic blasts from five acute myelogenous leukemia patients but not of normal umbilical cord blood mononuclear cells. In vitro kinase assays and immunoblotting using lysates from human MV4;11 leukemic cells showed inhibition of phosphorylation of known PIM downstream targets, such as BAD and eukaryotic translation initiation factor 4E-binding protein 1, by K00135. Taken together, we report a family of small molecules that selectively interact and block PIM kinases and could serve as a lead to develop new targeted antileukemic therapeutics. [Cancer Res 2007;67(14):6916-24]

Dissection of PIM serine/threonine kinases in FLT3-ITD–induced leukemogenesis reveals PIM1 as regulator of CXCL12–CXCR4-mediated homing and migration

“…To further test the hypothesis that PIM1 kinase activity is involved in regulation of CXCR4 surface expression, we used a recently identified potent small molecule inhibitor of human PIM1 (K00486; Pogacic et al, 2007). Human JURKAT acute lymphoblastic leukemia cells that express high levels of PIM1 and surface CXCR4 were used in this experiment.…”

Pim-1 controls NF-κB signalling by stabilizing RelA/p65

“…Indeed, small molecule inhibitors of the Pim-1 kinase have been recently described in vitro and in cell-based systems. [33][34][35] Targeting the Pim-1 kinase may be a beneficial addition to a traditional cancer chemotherapy regimen.…”

PIM Kinases Are Essential for Chronic Lymphocytic Leukemia Cell Survival (PIM2/3) and CXCR4-Mediated Microenvironmental Interactions (PIM1)

“…Previous studies have demonstrated the importance of the microenvironment, and especially the presence of stromal cells, for the survival of CLL cells (10,42). The interaction between BMSCs and CLL cells induces a protective microenvironment blocking the proapoptotic effect of various chemotherapeutical agents and spontaneous apoptosis in vitro.…”

“…K00135 and K00486, two imidazo[1,2-b]pyridazines, show low IC 50 values for inhibition of PIM1 and PIM2 kinase activity, and have shown proapoptotic effects in AML cells (37,42). The third PIM kinase inhibitor, A47, is based on another chemical structure, a furan thiazolidin, and inhibits all three PIM kinases in low micromolar concentrations including PIM3 (39).…”