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Efficient in vivo regulation of cytidine deaminase expression in the haematopoietic system using a doxycycline-inducible lentiviral vector system

Gene Therapy volume 20, pages 298–307 (2013)Cite this article

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Abstract

Regulated transgene expression may reduce transgene-specific and genotoxic risks associated with gene therapy. To prove this concept, we have investigated the suitability of doxycycline (Dox)-inducible human cytidine deaminase (hCDD) overexpression from lentiviral vectors to mediate effective myeloprotection while circumventing the lymphotoxicity observed with constitutive CDD activity. Rapid Dox-mediated transgene induction associated with a 6–17-fold increase in drug resistance was observed in 32D and primary murine bone marrow (BM) cells. Moreover, robust Dox-regulated transgene expression in the entire haematopoietic system was demonstrated for primary and secondary recipients of hCDD-transduced R26-M2rtTA transgenic BM cells. Furthermore, mice were significantly protected from myelosuppressive chemotherapy as evidenced by accelerated recovery of granulocytes (1.9±0.6 vs 1.3±0.3, P=0.034) and platelets (883±194 vs 584±160 103 per μl, P=0.011). Minimal transgene expression in the non-induced state and no overt cellular toxicities including lymphotoxicity were detected. Thus, using a relevant murine transplant model our data provide conclusive evidence that drug-resistance transgenes can be expressed in a regulated fashion in the lymphohaematopoietic system, and that Dox-inducible systems may be used to reduce myelotoxic side effect of anticancer chemotherapy or to avoid side effects of high constitutive transgene expression.

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Acknowledgements

We thank Matthias Ballmaier (PhD) and his team from the Core-Facility Cell-Sorting of Hannover Medical School for cell sorting, and Doreen Lüttge (Hannover Medical School) for excellent technical assistance. Furthermore, we thank Georg Kensah (Hannover Medical School) for the help in performing the timelapse video studies. This work was supported by grants from the Deutsche Forschungsgemeinschaft: Cluster of Excellence REBIRTH (Exc 62/1), SPP1230 Grant MO 886/3-1 (UM and TM) and by the EU framework programme grant PERSIST (CHB).

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Authors and Affiliations

  1. REBIRTH Cluster-of-Excellence, Research Group Reprogramming, Hannover Medical School, Hannover, Germany

    N Lachmann, S Brennig, N Pfaff, H Schermeier, R Phaltane & T Moritz

  2. Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, REBIRTH Cluster-of-Excellence, Leibniz Research Laboratories for Biotechnology and Artificial Organs, Hannover Medical School, Hannover, Germany

    J Dahlmann & I Gruh

  3. REBIRTH Cluster-of-Excellence, Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany

    U Modlich, A Schambach & C Baum

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  1. N Lachmann
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Lachmann, N., Brennig, S., Pfaff, N. et al. Efficient in vivo regulation of cytidine deaminase expression in the haematopoietic system using a doxycycline-inducible lentiviral vector system. Gene Ther 20, 298–307 (2013). https://doi.org/10.1038/gt.2012.40

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