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Transgene optimization significantly improves SIN vector titers, gp91phox expression and reconstitution of superoxide production in X-CGD cells

Gene Therapy volume 16, pages 111–118 (2009)Cite this article

Abstract

Gene therapy has proven to be of potential value for the correction of inherited hematopoietic disorders. However, the occurrence of severe side effects in some of the clinical trials has questioned the safety of this approach and has hampered the use of long terminal repeat-driven vectors for the treatment of a large number of patients. The development of self-inactivating (SIN) vectors with reduced genotoxicity provides an alternative to the currently used vectors. Our initial attempts to use SIN vectors for the correction of a myeloid disorder, chronic granulomatous disease, failed due to low vector titers and poor transgene expression. The optimization of the transgene cDNA (gp91phox) resulted in substantially increased titers and transgene expression. Most notably, transgene optimization significantly improved expression of a second cistron located downstream of gp91phox. Thus, optimization of the transgene sequence results in higher expression levels and increased therapeutic index allowing the use of low vector copy numbers per transduced cell and weaker internal promoters.

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Acknowledgements

This study was supported by grants from the CGD Research Trust (J4G/04B/GT) to MG and AJT, the Deutsche Forschungsgemeinschaft to SB (BR 2057/3-2), CB, AS and MG (SPP1230) and to CB and AS (excellence cluster REBIRTH) and from the Bundesministerium für Bildung und Forschung to CB, AS and MG (TreatID). AS was further supported by a personal stipend of the Else-Kröner Memorial Foundation. The Georg-Speyer-Haus is supported by the Bundesministerium für Gesundheit and the Hessisches Ministerium für Wissenschaft und Kunst.

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Author notes

  1. B Moreno-Carranza and M Gentsch: These two authors contributed equally to this work.

Authors and Affiliations

  1. Division of Applied Virology and Gene Therapy, Institute for Biomedical Research, Georg-Speyer-Haus, Frankfurt, Germany

    B Moreno-Carranza, S Stein, E Rudolf & M Grez

  2. Department of Pediatrics, University Clinic Carl Gustav Carus, Dresden, Germany

    M Gentsch, M F Ryser & S Brenner

  3. Department of Experimental Hematology, Hannover Medical School, Hannover, Germany

    A Schambach & C Baum

  4. Molecular Immunology Unit, UCL Institute of Child Health and Great Ormond Street Hospital for Children NHS Trust, London, UK

    G Santilli, S Haria & A J Thrasher

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  1. B Moreno-Carranza
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Correspondence to M Grez.

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Supplementary Information accompanies the paper on Gene Therapy website (http://www.nature.com/gt)

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Moreno-Carranza, B., Gentsch, M., Stein, S. et al. Transgene optimization significantly improves SIN vector titers, gp91phox expression and reconstitution of superoxide production in X-CGD cells. Gene Ther 16, 111–118 (2009). https://doi.org/10.1038/gt.2008.143

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