Structural Basis for the Synthesis of Indirubins as Potent and Selective Inhibitors of Glycogen Synthase Kinase-3 and Cyclin-Dependent Kinases

Abstract

Pharmacological inhibitors of glycogen synthase kinase-3 (GSK-3) and cyclin-dependent kinases have a promising potential for applications against several neurodegenerative diseases such as Alzheimer's disease. Indirubins, a family of bis-indoles isolated from various natural sources, are potent inhibitors of several kinases, including GSK-3. Using the cocrystal structures of various indirubins with GSK-3beta, CDK2 and CDK5/p25, we have modeled the binding of indirubins within the ATP-binding pocket of these kinases. This modeling approach provided some insight into the molecular basis of indirubins' action and selectivity and allowed us to forecast some improvements of this family of bis-indoles as kinase inhibitors. Predicted molecules, including 6-substituted and 5,6-disubstituted indirubins, were synthesized and evaluated as CDK and GSK-3 inhibitors. Control, kinase-inactive indirubins were obtained by introduction of a methyl substitution on N1.

Low-molecular-weight inhibitors of cell differentiation enable efficient growth of mouse iPS cells under feeder-free conditions

“…CHIR99021 is reported as the inhibitor of glycogen synthase kinase 3b (GSK-3b) signaling, which has a large impact to maintain pluripotency of ES and iPS cells effectively (Sato et al 2004;Besser 2004;Polychronopoulos et al 2004;Ying et al 2008). PD0325901 targets mitogen-activated protein kinase (MAPK/ERK kinase) (Sebolt-Leopold and Herrera 2004; Bain et al 2007).…”

Cyclin-dependent kinase inhibitor indirubin-3′-oxime selectively inhibits human papillomavirus type 16 E7-induced numerical centrosome anomalies

“…In order to test first whether the ability of IO to abrogate abnormal centriole duplication depends on its ability to inhibit protein kinase activity, we treated cells with 1-methyl-indirubin-3 0 -oxime (MeIO), a kinase-inactive analogue of IO (Figure 2a). N1-methylation of IO, leading to MeIO, prevents the cyclic nitrogen of the lactam ring of IO from donating a hydrogen bond to the peptidyl carbonyl oxygen of CDK2's Glu81, and thus from binding the ATP-binding pocket of the kinase and inhibiting its kinase activity (Hoessel et al, 1999;Davies et al, 2001;Meijer et al, 2003;Polychronopoulos et al, 2004). U-2 OS/centrin-GFP cells were transiently transfected with empty vector or HPV-16 E7 and treated either with 1 mM IO or 1 mM MeIO for 24 h. As previously observed (Figure 1d), IO led to a significant reduction of cells with abnormal centriole numbers in HPV-16 E7 transfected cells (3.1-fold, Pp0.05).…”

“…Inhibitor treatment and in vitro kinase assay IO and MeIO were synthesized as described elsewhere (Hoessel et al, 1999;Meijer et al, 2003;Polychronopoulos et al, 2004). Cells were treated with IO dissolved in dimethyl sulfoxide (DMSO) for 24 h at the indicated concentrations.…”

Indirubin-3′-monoxime inhibits autophosphorylation of FGFR1 and stimulates ERK1/2 activity via p38 MAPK

“…Furthermore, indirubin-3 0 -monoxime, but not indirubin, was able to inhibit CDK9 in a cell-free kinase assay and to inhibit replication of HIV-1 in blood mononuclear cells and macrophages (Heredia et al, 2005). Additionally, it was recently reported that indirubin derivatives were able to block Stat3 signaling by inhibiting the Src kinase activity and in this way induce apoptosis in human cancer cells (Nam et al, 2005), and also, bind to and inhibit glycogen synthetase kinase-3 (Leclerc et al, 2001;Polychronopoulos et al, 2004), aryl hydrocarbon receptor (Adachi et al, 2001), muscle glycogen phosphorylase b (Kosmopoulou et al, 2004) and c-Jun NH2-terminal kinase (JNK) (Xie et al, 2004). More recently, indirubin-3 0 -monoxime was found to inhibit the activation of nuclear factor-kB resulting in enhancement of apoptosis induced by tumor necrosis factors in human leukemic cells (Sethi et al, 2006).…”