CR8, a potent and selective, roscovitine-derived inhibitor of cyclin-dependent kinases

Abstract

Among the ten pharmacological inhibitors of cyclindependent kinases (CDKs) currently in clinical trials, the purine roscovitine (CYC202, Seliciclib) is undergoing phase 2 trials against non-small-cell lung and nasopharyngeal cancers. An extensive medicinal chemistry study, designed to generate more potent analogues of roscovitine, led to the identification of an optimal substitution at the N6 position (compound CR8). An extensive selectivity study (108 kinases) highlights the exquisite selectivity of CR8 for CDK1/2/3/5/7/9. CR8 was 2-to 4-fold more potent than (R)-roscovitine at inhibiting these kinases. Cocrystal structures of (R)-CR8 and (R)-roscovitine with pCDK2/cyclin A showed that both inhibitors adopt essentially identical positions. The cellular effects of CR8 and (R)-roscovitine were investigated in human neuroblastoma SH-SY5Y cells. CR8 inhibited the phosphorylation of CDK1 and 9 substrates, with a 25-50 times higher potency compared to (R)-roscovitine. CR8 was consistently more potent than (R)-roscovitine at inducing apoptotic cell death parameters: 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium reduction (40-fold), lactate dehydrogenase release (35-fold), caspases activation (68-fold) and poly-(ADP-ribose)polymerase cleavage (50-fold). This improved cell death-inducing activity of CR8 over (R)-roscovitine was observed in 25 different cell lines. Altogether these results show that second-generation analogues of (R)-roscovitine can be designed with improved antitumor potential.

CR8, a Novel Inhibitor of CDK, Limits Microglial Activation, Astrocytosis, Neuronal Loss, and Neurologic Dysfunction after Experimental Traumatic Brain Injury

“…[4][5][6][8][9][10]27 CR8, an N6-biarylsubstituted derivative of the selective CDK inhibitor, roscovitine, was synthesized to generate a second-generation analog with greater therapeutic potential. 28 CR8 not only exhibits higher solubility and 50-fold greater potency in vitro, 28 but also 10 or 20 times higher potency in vivo. 10 In order to simulate a more clinically relevant treatment paradigm, we administered CR8 systemically at 3 hours post injury and investigated its long-term neuroprotective effects on LFP injury-induced neurologic deficits, neurodegeneration, and neuroinflammation.…”

N-&-N, a new class of cell death-inducing kinase inhibitors derived from the purine roscovitine

“…Crystals were briefly cryoprotected in 8 mol/L sodium formate before being cryocooled in liquid nitrogen. The crystal structure of phospho-CDK2/cyclin A with (R)-roscovitine will be described in details elsewhere (40).…”

“…These observations were confirmed when similar experiments were carried out using extracts of SH-SY5Y cells (data not shown). 8 A full description of the roscovitine-CDK2/cyclin A cocrystal structure will be presented elsewhere (40). Both inhibitors occupy the CDK2 ATP-binding site and make two hydrogen bonds to the CDK2 backbone within the hinge sequence that links the two lobes of the kinase.…”

Inhibition of NF-κB–Mediated Signaling by the Cyclin-Dependent Kinase Inhibitor CR8 Overcomes Prosurvival Stimuli to Induce Apoptosis in Chronic Lymphocytic Leukemia Cells

“…Flavopiridol and SNS-032 are currently in clinical trials for CLL (18). Although roscovitine progressed through phase II clinical trials for non-small cell lung and nasopharyngeal cancers due to its strong selectivity for CDKs and its relative lack of toxicity (13), its weak potency and short half-life in vivo led to the development of more potent analogs (19). Here, we show that CR8, a novel roscovitine analog, possesses enhanced potency over roscovitine for inducing apoptosis in CLL cells, which is not reduced by mouse fibroblast L cell (NT-L)-mediated cytoprotection.…”