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Retroviral vectors for clinical immunogene therapy are stable for up to 9 years

Cancer Gene Therapy volume 15, pages 268–274 (2008)Cite this article

Abstract

Recombinant retroviruses are one of the most commonly used gene transfer vehicles for therapeutic gene delivery. The stability of viral vectors upon long-term storage, anticipated to be short lived, is expected to impact timeline and financial course of clinical immunogene therapy. However, to date little is known about vector stability. Therefore, we analyzed the stability of retroviral vectors produced in culture supernatants (RTVsup) for ex vivo gene therapy upon long-term storage. We have generated RTVsups derived from two packaging cell lines, PG13 and Phoenix(Ampho). Both lines produced murine leukemia virus-derived SFG-scFv(G250)-CD4γ vector, which were pseudotyped with the gibbon ape leukemia virus envelope and amphotropic envelope, respectively. The supernatants were stored at −80 or −196 °C. To date, the PG13-derived RTVsups have been evaluated over a period of 9 years (1998–2007). In addition, a clinical batch of Phoenix(Ampho)-derived RTVsup has been evaluated over a period of 5 years (2002–2007). Here, we show that both RTVsups, when stored up to 9 and 5 years, respectively, do not show any sign of decay in their capacity to functionally transduce primary human T cells. These data provide evidence that in terms of ‘life expectancy’ the production and storage of clinical batches of RTVsup for gene therapy warrants the corresponding professional and financial risks.

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References

  1. McTaggart S, Al Rubeai M . Retroviral vectors for human gene delivery. Biotechnol Adv 2002; 20: 1–31.

    Article  CAS  PubMed  Google Scholar 

  2. Thomas CE, Ehrhardt A, Kay MA . Progress and problems with the use of viral vectors for gene therapy. Nat Rev Genet 2003; 4: 346–358.

    Article  CAS  PubMed  Google Scholar 

  3. Lamers C, van Elzakker P, Langeveld S, Sleijfer S, Gratama J . Process validation and clinical evaluation of a protocol to generate gene-modified T lymphocytes for imunogene therapy for metastatic renal cell carcinoma: GMP-controlled transduction and expansion of patient's T lymphocytes using a carboxy anhydrase IX-specific scFv transgene. Cytotherapy 2006; 8: 542–553.

    Article  CAS  PubMed  Google Scholar 

  4. Le Doux JM, Davis HE, Morgan JR, Yarmush ML . Kinetics of retrovirus production and decay. Biotechnol Bioeng 1999; 63: 654–662.

    Article  CAS  PubMed  Google Scholar 

  5. Pizzato M, Merten OW, Blair ED, Takeuchi Y . Development of a suspension packaging cell line for production of high titre, serum-resistant murine leukemia virus vectors. Gene Therapy 2001; 8: 737–745.

    Article  CAS  PubMed  Google Scholar 

  6. Beer C, Meyer A, Muller K, Wirth M . The temperature stability of mouse retroviruses depends on the cholesterol levels of viral lipid shell and cellular plasma membrane. Virology 2003; 308: 137–146.

    Article  CAS  PubMed  Google Scholar 

  7. Carmo M, Faria TQ, Falk H, Coroadinha AS, Teixeira M, Merten OW et al. Relationship between retroviral vector membrane and vector stability. J Gen Virol 2006; 87: 1349–1356.

    Article  CAS  PubMed  Google Scholar 

  8. Coroadinha AS, Ribeiro J, Roldao A, Cruz PE, Alves PM, Merten OW et al. Effect of medium sugar source on the production of retroviral vectors for gene therapy. Biotechnol Bioeng 2006; 94: 24–36.

    Article  CAS  PubMed  Google Scholar 

  9. Coroadinha AS, Silva AC, Pires E, Coelho A, Alves PM, Carrondo MJ . Effect of osmotic pressure on the production of retroviral vectors: enhancement in vector stability. Biotechnol Bioeng 2006; 94: 322–329.

    Article  CAS  PubMed  Google Scholar 

  10. Coroadinha AS, Alves PM, Santos SS, Cruz PE, Merten OW, Carrondo MJ . Retrovirus producer cell line metabolism: implications on viral productivity. Appl Microbiol Biotechnol 2006; 72: 1125–1135.

    Article  CAS  PubMed  Google Scholar 

  11. Kaptein LC, Greijer AE, Valerio D, Van Beusechem VW . Optimized conditions for the production of recombinant amphotropic retroviral vector preparations. Gene Therapy 1997; 4: 172–176.

    Article  CAS  PubMed  Google Scholar 

  12. Higashikawa F, Chang L . Kinetic analyses of stability of simple and complex retroviral vectors. Virology 2001; 280: 124–131.

    Article  CAS  PubMed  Google Scholar 

  13. Wikstrom K, Blomberg P, Islam KB . Clinical grade vector production: analysis of yield, stability, and storage of GMP-produced retroviral vectors for gene therapy. Biotechnol Prog 2004; 20: 1198–1203.

    Article  PubMed  Google Scholar 

  14. Lamers CH, Sleijfer S, Vulto AG, Kruit WH, Kliffen M, Debets R et al. Treatment of metastatic renal cell carcinoma with autologous T-lymphocytes genetically retargeted against carbonic anhydrase IX: first clinical experience. J Clin Oncol 2006; 24: e20–e22.

    Article  PubMed  Google Scholar 

  15. Lamers CH, Willemsen RA, Luider BA, Debets R, Bolhuis RL . Protocol for gene transduction and expansion of human T lymphocytes for clinical immunogene therapy of cancer. Cancer Gene Ther 2002; 9: 613–623.

    Article  CAS  PubMed  Google Scholar 

  16. Lamers CH, Willemsen RA, van Elzakker P, van Krimpen BA, Gratama JW, Debets R . Phoenix-ampho outperforms PG13 as retroviral packaging cells to transduce human T cells with tumor-specific receptors: implications for clinical immunogene therapy of cancer. Cancer Gene Ther 2006; 13: 503–509.

    Article  CAS  PubMed  Google Scholar 

  17. Uemura H, Okajima E, Debruyne FMJ, Oosterwijk E . Internal image antiidiotype antibodies related to renal-cell carcinoma-associated antigen G250. Intel J Cancer 1994; 56: 609–614.

    Article  CAS  Google Scholar 

  18. Byun J, Kim JM, Kim SH, Yim J, Robbins PD, Kim S . A simple and rapid method for the determination of recombinant retrovirus titer by G418 selection. Gene Therapy 1996; 3: 1018–1020.

    CAS  PubMed  Google Scholar 

  19. Reeves L, Smucker P, Cornetta K . Packaging cell line characteristics and optimizing retroviral vector titer: the National Gene Vector Laboratory experience. Hum Gene Ther 2000; 11: 2093–2103.

    Article  CAS  PubMed  Google Scholar 

  20. Kwon YJ, Hung G, Anderson WF, Peng CA, Yu H . Determination of infectious retrovirus concentration from colony-forming assay with quantitative analysis. J Virol 2003; 77: 5712–5720.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  21. Sanburn N, Cornetta K . Rapid titer determination using quantitative real-time PCR. Gene Therapy 1999; 6: 1340–1345.

    Article  CAS  PubMed  Google Scholar 

  22. Husemoen LL, Gram GJ, Hansen JE . Application of the polymerase chain reaction in determination of recombinant retrovirus titers as fifty percent endpoints. APMIS 2000; 108: 38–44.

    Article  CAS  PubMed  Google Scholar 

  23. Merten OW . State-of-the-art of the production of retroviral vectors. J Gene Med 2004; 6 (Suppl 1): S105–S124.

    Article  CAS  PubMed  Google Scholar 

  24. Aloia RC, Jensen FC, Curtain CC, Mobley PW, Gordon LM . Lipid composition and fluidity of the human immunodeficiency virus. Proc Natl Acad Sci USA 1988; 85: 900–904.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Aloia RC, Tian H, Jensen FC . Lipid composition and fluidity of the human immunodeficiency virus envelope and host cell plasma membranes. Proc Natl Acad Sci USA 1993; 90: 5181–5185.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Beer C, Pedersen L, Wirth M . Amphotropic murine leukaemia virus envelope protein is associated with cholesterol-rich microdomains. Virol J 2005; 2: 36.

    Article  PubMed Central  PubMed  Google Scholar 

  27. Beer C, Buhr P, Hahn H, Laubner D, Wirth M . Gene expression analysis of murine cells producing amphotropic mouse leukaemia virus at a cultivation temperature of 32 and 37 degrees C. J Gen Virol 2003; 84: 1677–1686.

    Article  CAS  PubMed  Google Scholar 

  28. Cruz PE, Silva AC, Roldao A, Carmo M, Carrondo MJ, Alves PM . Screening of novel excipients for improving the stability of retroviral and adenoviral vectors. Biotechnol Prog 2006; 22: 568–576.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank C Groot-van Ruyven and B van Krimpen for technical assistance. This work was supported by the Dutch Cancer Foundation (grant DDHK99-1865), the European Commission grant QLK3-1999-01262 and the Cancer Research Institute, New York, NY (clinical investigation grant ‘Immuno-gene therapy of metastatic renal cell cancer patients’).

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Authors and Affiliations

  1. Department of Medical Oncology, Laboratory of Clinical and Tumor Immunology, Erasmus MC—Daniel den Hoed Cancer Center, Rotterdam, The Netherlands

    C H J Lamers, P van Elzakker, B A Luider, S C L van Steenbergen, R Debets & J W Gratama

  2. Department of Medical Oncology, Erasmus MC—Daniel den Hoed Cancer Center, Rotterdam, The Netherlands

    S Sleijfer

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  1. C H J Lamers
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  2. P van Elzakker
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Correspondence to C H J Lamers.

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Lamers, C., van Elzakker, P., Luider, B. et al. Retroviral vectors for clinical immunogene therapy are stable for up to 9 years. Cancer Gene Ther 15, 268–274 (2008). https://doi.org/10.1038/sj.cgt.7701114

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