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A Cdc7 kinase inhibitor restricts initiation of DNA replication and has antitumor activity

Nature Chemical Biology volume 4, pages 357–365 (2008)Cite this article

Abstract

Cdc7 is an essential kinase that promotes DNA replication by activating origins of replication. Here, we characterized the potent Cdc7 inhibitor PHA-767491 (1) in biochemical and cell-based assays, and we tested its antitumor activity in rodents. We found that the compound blocks DNA synthesis and affects the phosphorylation of the replicative DNA helicase at Cdc7-dependent phosphorylation sites. Unlike current DNA synthesis inhibitors, PHA-767491 prevents the activation of replication origins but does not impede replication fork progression, and it does not trigger a sustained DNA damage response. Treatment with PHA-767491 results in apoptotic cell death in multiple cancer cell types and tumor growth inhibition in preclinical cancer models. To our knowledge, PHA-767491 is the first molecule that directly affects the mechanisms controlling initiation as opposed to elongation in DNA replication, and its activities suggest that Cdc7 kinase inhibition could be a new strategy for the development of anticancer therapeutics.

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Figure 1: PHA-767491 inhibits the initiation reaction of DNA replication in HeLa cells.
Figure 2: PHA-767491 inhibits cellular Cdc7 and Cdk9 kinases.
Figure 3: Entry and progression through S phase is affected by PHA-767491.
Figure 4: PHA-767491 has antitumor activity in rodents.
Figure 5: PHA-767491 impairs Mcm2 phosphorylation at Cdc7-dependent sites in tumors.

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Acknowledgements

We are indebted to F. Caprera, R. Giavarini, N. Amboldi and our colleagues in the support units for their contribution to the experimental activities. We thank N. Colombo for NMR analysis and D. Brown, R. Bravo, M. Varasi, N. Mongelli, G. Draetta and F. Colotta for discussion and support. We also thank R. Schilling, M. Hunter, D. Reitz, M. Vazquez, W. Vernier, D. Anderson and D. Phillion for their contribution in early screening activities, compound management and helpful advice. We finally thank A. Migliazza for critical reading of the manuscript.

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Author notes

  1. Pierluigi Tenca & Deborah Brotherton

    Present address: Present addresses: InCura via Staffolo 60, 26040 Vicomoscano (Cremona), Italy (P.T.) and Chroma Therapeutics, 93 Milton Park, Abington OX14 5RY, UK (D.B.).,

Authors and Affiliations

  1. Nerviano Medical Sciences Oncology, Via Pasteur 10, Nerviano, 20014, Italy

    Alessia Montagnoli, Barbara Valsasina, Valter Croci, Maria Menichincheri, Sonia Rainoldi, Vanessa Marchesi, Marcello Tibolla, Clara Albanese, Veronica Patton, Rachele Alzani, Antonella Ciavolella, Francesco Sola, Antonio Molinari, Daniele Volpi, Nilla Avanzi, Francesco Fiorentini, Marina Cattoni, Sandra Healy, Dario Ballinari, Enrico Pesenti, Antonella Isacchi, Jurgen Moll, Ermes Vanotti & Corrado Santocanale

  2. Genomic Vision, Pasteur Biotop, 28 rue du Docteur Roux, Paris, 75724, cedex 15, France

    Aaron Bensimon

  3. National Centre for Biomedical Engineering and Sciences, National University of Ireland Galway, University Road, Galway, Ireland

    Corrado Santocanale

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Contributions

A. Montagnoli, S.R., V.M., P.T., D. Brotherton, C.A. and F.S. set up and performed cell-based assays. B.V., A. Molinari, D.V. and N.A. set up and performed biochemical experiments. M.M., M.T. and E.V. synthesized and performed chemical characterization of PHA-767491. V.C. and E.P. devised antitumor activity experiments. V.P. and R.A. performed IHC analysis. A.C. and D. Ballinari performed cell proliferation assays. A.B. performed DNA combing experiments. F.F. and M.C. worked on the preclinical characterization of PHA-767491. A. Montagnoli, M.M., B.V., S.H., A.I., E.P., J.M., E.V. and C.S. worked on data analysis, formulated the hypothesis and supervised experiments. S.H. and C.S. wrote the paper.

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Correspondence to Corrado Santocanale.

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Competing interests

A.M., B.V., V.C., M.M., S.R., V.M., C.A., V.P., R.A., A.C., F.S., A.M., D.V., N.A., F.F., M.C., D.B., E.P., A.I., J.M. and E.V. declare that they are employees of Nerviano Medical Sciences S.r.l. T.B., P.T., D.B., S.H. and C.S. were former employees of Nerviano Medical Sciences S.r.l. A.B. is co-founder and shareholder of Genomic Vision.

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Montagnoli, A., Valsasina, B., Croci, V. et al. A Cdc7 kinase inhibitor restricts initiation of DNA replication and has antitumor activity. Nat Chem Biol 4, 357–365 (2008). https://doi.org/10.1038/nchembio.90

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