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LCR-mediated, long-term tissue-specific gene expression within replicating episomal plasmid and cosmid vectors

Gene Therapy volume 9, pages 327–336 (2002)Cite this article

Abstract

Locus control regions (LCRs) are transcriptional regulatory elements, which possess a dominant chromatin remodelling and transcriptional activating capability conferring full physiological levels of expression on a gene linked in cis, when integrated into the host cell genome. Using the human β-globin LCR (βLCR) as a model, we show that this class of control element can drive high levels of tissue-specific gene expression in stably transfected cultured cells from within an Epstein–Barr virus-based plasmid REV. Furthermore, a 38-kb βLCR minilocus-REV cosmid vector was efficiently retained and maintained therapeutic levels of β-globin transgene expression in the absence of drug selective pressure over a 2-month period of continuous culture equivalent to at least 60 generations. This demonstrates for the first time the feasibility of using REVs for gene therapy of the haemoglobinopathies. Importantly, our results demonstrate that as in the case of integrated transgenes, expression from within REVs is prone to silencing but that the inclusion of the βLCR prevented this repression of gene function. Therefore, appropriate control elements to provide and maintain tissue-specific gene expression, as well as the episomal status of REVs is a crucial feature in vector design. Our data suggest that LCRs can contribute to this vital function.

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Acknowledgements

We thank Bill Sugden for the p220.2 and cos203 EBV-based vectors. This work was initiated with support from Cobra Therapeutics (Keele, UK) and continued with funding under the European Union Framework IV Biomed-2 (contract No. BMH4-CT96-1279) and Biotechnology (contract No. BIO4-CT96-0414) programmes. AA was funded for this work by a European Union Marie Curie Fellowship (contract No. HPMF-CT-1994-00091).

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Author notes

  1. C-M Chow

    Present address: Gene Regulation and Chromatin Group and Membrane Transport Biology Group, MRC Clinical Sciences Centre, Hammersmith Hospital, Du Cane Road, London, W12 0NN, UK

Authors and Affiliations

  1. Division of Medical and Molecular Genetics, Nuclear Biology Group, GKT School of Medicine, Guy's Hospital, London, UK

    C-M Chow, A Athanassiadou, S Raguz, L Psiouri, L Harland, M Malik, MA Aitken & M Antoniou

  2. Department of General Biology, Faculty of Medicine, University of Patras, Patras, Greece

    A Athanassiadou & L Psiouri

  3. Department of Cell Biology, Erasmus University Medical Genetics Centre, Rotterdam, The Netherlands

    F Grosveld

  4. Central Research Division, Bioinformatics Group, Pfizer Limited, Sandwich, Kent, CT13 9NJ, UK

    L Harland

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Chow, CM., Athanassiadou, A., Raguz, S. et al. LCR-mediated, long-term tissue-specific gene expression within replicating episomal plasmid and cosmid vectors. Gene Ther 9, 327–336 (2002). https://doi.org/10.1038/sj.gt.3301654

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