Stabilization of gammaretroviral and lentiviral vectors: from production to gene transfer.
BACKGROUND
The low stability of gammaretroviral and lentiviral vectors affects their production, making high quality clinical preparations a difficult goal to achieve. Recently, our laboratory has shown that the main inactivation mechanism for both these vectors is the loss of their capacity to perform reverse transcription. The present study aimed to increase the stability of gammaretroviral and lentiviral at 37 degrees C and at 4 degrees C.


METHODS
Inactivation studies were performed with gammaretroviral and lentiviral vectors at 37 and 4 degrees C, with and without several stabilizing compounds. The residual viral infectivity and reverse transcription capacity of these samples were tested.


RESULTS
The results obtained demonstrate that it is possible to increase the stability of reverse transcription and the infectivity stability of purified gammaretroviral vectors by adding recombinant human albumin (rHSA) to the storage buffer, both at 37 degrees C and at 4 degrees C. For lentiviral vectors, it was observed that further protection was needed. This was achieved by adding lipids to the storage buffer, using a mixture of lipoproteins and rHSA. The difference of stabilization between gammaretroviral and lentiviral vectors was validated by performing stabilization tests with vectors possessing different envelope proteins and produced by different cell lines.


CONCLUSIONS
The presented study reveals that it is possible to increase the half-life of purified gammaretroviral and lentiviral vectors at 37 degrees C and at 4 degrees C, but the two vectors have different stabilization requirements: for retroviral vectors, the addition of rHSA is enough and, for lentiviral vectors, it is necessary to add both lipoproteins and rHSA. The increase of the stability of the reverse transcription process was shown to have a high impact with respect to the increase of the stability of infectivity.
Carmo
Marlene
M
IBET/ITQB-UNL, Oeiras, Portugal.
Alves
Ana
A
Rodrigues
Ana Filipa
AF
Coroadinha
Ana Sofia
AS
Carrondo
Manuel Jose Teixeira
MJ
Alves
Paula Marques
PM
Cruz
Pedro Estilita
PE
D002460:Cell Line; D017988:Gammaretrovirus / Q000235:genetics*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D048348:Reverse Transcription; D014759:Viral Envelope Proteins / Q000378:metabolism; D038301:Virus Inactivation
D014759:Viral Envelope Proteins