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

Oncogene volume 27, pages 5797–5807 (2008)Cite this article

Abstract

Among the ten pharmacological inhibitors of cyclin-dependent 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.

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Accession codes

Accessions

GenBank/EMBL/DDBJ

Abbreviations

CDK:

cyclin-dependent kinase

CK1:

casein kinase 1

DMSO:

dimethylsulfoxide

GSK-3:

glycogen synthase kinase-3

LDH:

lactate dehydrogenase

MTS:

3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium

PBS:

phosphate-buffered saline

PP1α:

protein phosphatase 1α

Rb:

retinoblastoma protein

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Acknowledgements

We are grateful to J Boix, C Guillouzo and J Mester for cell lines, G Ponzio for the human foreskin primary fibroblasts, Dr Michael Kubbutat (ProQinase, Freiburg, Germany) and Sir Philip Cohen (Dundee, UK) for the selectivity panel assays and to the beamline scientists at the ESRF (ID14-EH-2 and ID14-EH-1) for providing excellent facilities for crystal data collection. This research was supported by grants from the EEC (FP6-2002-Life Sciences and Health, PRO-KINASE Research Project) (LM, JE), the ‘Cancéropole Grand-Ouest’ grant (LM), the ‘Institut National du Cancer’ (INCa ‘Cancer Détection d’innovations 2006′) (LM), the ‘Association France-Alzheimer Finistère’ (LM) and the ‘Ligue Nationale contre le Cancer (Comité du Finistère)’ (LM). KB was supported by a fellowship from the ‘Ministère de la Recherche’ and from the ‘Association pour la Recherche sur le Cancer’. NO benefited from a PhD fellowship from the ‘Institut National du Cancer’. The structures of pCDK2/cyclin A/(R)-roscovitine, and pCDK2/cyclin A/(R)-CR8 have been deposited in the PDB with accession number codes 3DDQ and 3DDP, respectively.

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Authors and Affiliations

  1. CNRS, Cell Cycle Group, Station Biologique, Bretagne, France

    K Bettayeb, Y Ferandin & L Meijer

  2. Laboratoire de Chimie Organique 2, INSERM U 648, Université Paris—Descartes, Paris cedex, France

    N Oumata & H Galons

  3. Department of Biochemistry, Laboratory of Molecular Biophysics, University of Oxford, Oxford, UK

    A Echalier & J A Endicott

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  1. K Bettayeb
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  2. N Oumata
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  3. A Echalier
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  6. H Galons
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  7. L Meijer
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Correspondence to H Galons or L Meijer.

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Supplementary information accompanies the paper on the Oncogene website (http://www.nature.com/onc)

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Bettayeb, K., Oumata, N., Echalier, A. et al. CR8, a potent and selective, roscovitine-derived inhibitor of cyclin-dependent kinases. Oncogene 27, 5797–5807 (2008). https://doi.org/10.1038/onc.2008.191

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