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Investigational New Drugs

, Volume 22, Issue 3, pp 253–262 | Cite as

A novel mammalian cell-based approach for the discovery of anticancer drugs with reduced cytotoxicity on non-dividing cells

  • Valeria Gonzalez-Nicolini
  • Cornelia Fux
  • Martin Fussenegger
Article

Abstract

A key asset of cytotoxic drugs in cancer therapeutics is their ability to discriminate between proliferating and mitotically inert cells and eliminate preferentially neoplastic ones. We have designed a high throughput-compatible mammalian cell-based assay for the discovery of cytotoxic drugs, which selectively kill proliferation-competent target cells. This cytotoxic drug discovery assay is based on a transgenic CHO-K1-derived cell line engineered for a conditional G1-specific growth arrest following tetracycline-responsive overexpression of the human cyclin-dependent kinase inhibitor p27Kip1. The CHO-derived cell line CHO-p27Kip1 shows wild type proliferation rates and can be expanded in the presence of tetracycline antibiotics when p27Kip1 expression is repressed. Upon withdrawal of regulating antibiotics CHO-p27Kip1 differentiates into a 1:1 mixed population consisting of two different proliferation phenotypes: (i) a G1-arrested cell population induced by heterologous expression of p27Kip1 which mimics mitotically inactive terminally differentiated cells and (ii) a proliferation-competent cell population which eliminated the p27Kip1 expression unit and imitates neoplastic cell characteristics. Addition of chemical or metabolic libraries to CHO-p27Kip1 populations cultivated in tetracycline-free medium followed by scoring for cell viability will reveal cytotoxic drug candidates associated with a high viability ratio of proliferation-competent/arrested populations.

We have validated the cell-based cytotoxic drug discovery assay using the clinically licensed cancer drugs mitomycin C, doxorubicin, etoposide and 5-fluorouracil. Comparative proof-of-concept studies showed that these top-prescribed cancer therapeutics preferentially eliminate proliferating cells while showing less interference with the viability of G1-arrested cell populations. These results demonstrate the CHO-p27Kip1-based cytotoxic drug finder technology is ready-to-apply for high throughput screenings of chemical as well as metabolic libraries to discover novel cancer therapeutics which show reduced cytotoxicity on terminally differentiated cells.

TET system p27Kip1 drug screening etoposide doxorubicin mitomycin C 5-fluorouracil 

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Copyright information

© Kluwer Academic Publishers 2004

Authors and Affiliations

  • Valeria Gonzalez-Nicolini
    • 1
  • Cornelia Fux
    • 1
  • Martin Fussenegger
    • 1
  1. 1.Institute of BiotechnologySwiss Federal Institute of TechnologyZurichSwitzerland

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