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Development of formulations that enhance physical stability of viral vectors for gene therapy

Gene Therapy volume 8, pages 1281–1290 (2001)Cite this article

Abstract

This study summarizes our initial efforts to address an issue that is critical to the success of any multicenter gene therapy clinical trial – maintenance of vector viability during shipping and storage at remote test sites. We have identified formulation and processing factors that influence stability of viral preparations such as selection of appropriate buffer systems, cryoprotectants, and storage conditions. Adenovirus and adeno-associated virus expressing E. coli beta-galactosidase (lacZ) were suspended in blends of complex carbohydrates, cyclodextrins and various surfactants. X-gal stains of 293 and 84-31 cells were used to determine infectious titer of all preparations. Potassium phosphate-buffered preparations consistently maintained high viral titers after storage at −20 and 4°C. Blends of sucrose, mannitol, and surfactant showed negligible loss of titer for 35 days at 4°C. Formulations of sucrose and cyclodextrin were stable for 2 years at −20°C. Negligible loss in titer was observed in unit-dose viral preparations lyophilized in sucrose and stored at 4°C for 1 year after an initial loss of 0.5 log due to processing. Studies with lyophilized sucrose/mannitol blends have shown that viral recovery after processing is directly related to the final moisture content of the dried product. Virus concentration also plays a significant role in recovery after processing with highly concentrated preparations showing minimal loss in titer after lyophilization. In summary, lyophilized preparations that can be shipped and stored at 25°C offer a solution to the current problem of distribution of viral vectors for clinical trials.

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Acknowledgements

The authors sincerely thank Dr Andrew McGee and Mr William Romanow of the University of Pennsylvania Department of Material Science Engineering and the Laboratory for Research on the Structure of Matter for assistance with differential scanning calorimetry and thermogravimetric analysis. We would also like to thank Animal Models Group (Marcia Houston-Leslie, Rosalind Barr, Jeanna Stabinski, Holly Clouse) of the Institute for Human Gene Therapy for expert technical assistance with vector administration. This work was funded by the Cystic Fibrosis Foundation, the NIH (NHLBI P01 HL59407, NIDDK P30 DK47757), and Genovo, Inc., a biotechnology company which Dr Wilson founded and has equity in. MAC is a recipient of a National Research Service Award.

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Authors and Affiliations

  1. Division of Pharmaceutics, The University of Texas at Austin College of Pharmacy, Austin, TX, USA

    MA Croyle & X Cheng

  2. Institute for Human Gene Therapy and Department of Molecular and Cellular Engineering, University of Pennsylvania, Philadelphia, PA, USA

    JM Wilson

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  1. MA Croyle
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Croyle, M., Cheng, X. & Wilson, J. Development of formulations that enhance physical stability of viral vectors for gene therapy. Gene Ther 8, 1281–1290 (2001). https://doi.org/10.1038/sj.gt.3301527

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Keywords

  • adenovirus
  • AAV
  • physical stability
  • formulation
  • lyophilization

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