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J Gen Virol 87 (2006), 1349-1356; DOI 10.1099/vir.0.81302-0

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© 2006 Society for General Microbiology

Relationship between retroviral vector membrane and vector stability

M. Carmo1, T. Q. Faria1, H. Falk2, A. S. Coroadinha1, M. Teixeira1, O.-W. Merten3, C. Gény-Fiamma3, P. M. Alves1, O. Danos3, A. Panet2, M. J. T. Carrondo1,4 and P. E. Cruz1,5,6

1 IBET/ITQB, Apartado 12, P-2780-901 Oeiras, Portugal
2 Hebrew University, Hadassah Medical School, Jerusalem, Israel
3 Généthon, 1 bis Rue de l'Internationale, F-91000 Evry, France
4 Laboratório de Engenharia Bioquímica, FCT/UNL, P-2825 Monte da Caparica, Portugal
5 ECBio, Lab. 4.11, Ed. ITQB, Apartado 98, P-2781-901 Oeiras, Portugal
6 Universidade Atlântica, Antiga Fábrica da Pólvora de Barcarena, P-2745-615 Barcarena, Portugal

Correspondence
P. E. Cruz
pcruz{at}itqb.unl.pt

The present work studies the physico-chemical properties of retroviral vector membrane, in order to provide some explanation for the inactivation kinetics of these vectors and to devise new ways of improving transduction efficiency. For this purpose, vectors with an amphotropic envelope produced by TE Fly A7 cells at two culture temperatures (37 and 32 °C) were characterized by different techniques. Electron paramagnetic resonance (EPR) results showed that vectors produced at 32 °C are more rigid than those produced at 37 °C. Further characterization of vector membrane composition allowed us to conclude that the vector inactivation rate increases with elevated cholesterol to phospholipid ratio. Differential scanning calorimetry (DSC) showed that production temperature also affects the conformation of the membrane proteins. Transduction studies using HCT116 cells and tri-dimensional organ cultures of mouse skin showed that vectors produced at 37 °C have higher stability and thus higher transduction efficiency in gene therapy relevant cells as compared with vectors produced at 32 °C. Overall, vectors produced at 37 °C show an increased stability at temperatures below 4 °C. Since vector membrane physico-chemical properties are affected in response to changes in culture temperature, such changes, along with alterations in medium composition, can be used prospectively to improve the stability and the transduction efficiency of retroviral vectors for therapeutic purposes.






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Copyright © 2006 by the Society for General Microbiology.