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  • Brief Communication
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Functional transfer of CD40L gene in human B-cell precursor ALL blasts by second-generation SIN lentivectors

Abstract

Three different second-generation lentiviral self-inactivating vectors containing CMV, EF1α and PGK promoter, respectively, and all carrying the exogenous GFP gene, were compared for expression in human B-cell precursor ALL blasts. At a comparable percentage of transduction and vector DNA copy number, CMV clearly showed better efficiency of transcription. Human bone marrow stromal cells were favored compared to the MRC-5 cell line, as support for cell viability during infection. Cells were infected and analyzed after variable culture times ranging from 4 to 12 days, to reduce the possibility of pseudotransduction. In 10/14 samples, we detected more than 20% GFP-positive cells after exposure to high-titer viral supernatants. We then tested a similar vector carrying the human CD40L cDNA and, in similar infection conditions, obtained more than 20% transduction in 6/6 samples. The levels of transduction obtained were sufficient to induce the upregulation of CD83 molecule in cocultured immature dendritic cells.

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Acknowledgements

This work was supported in part by grants from the Associazione Italiana Ricerca sul Cancro (to MI and AB), the Istituto Superiore di Sanità (ISS, Rome, AIDS grants 30C.40, 30D.41 and 40C.66 to MI), the Italian Ministry for University and Research (project FIRB no. RBAU01J2ER and RBAU01H8SX to MI), the Fondazione Tettamanti, Monza, the MIUR – Oncologia (to AB) and Progetti di Ricerca Finalizzata Ministero della Salute RF202/02 (to AB and MB). MB was supported by fellowships from the Fundaçao Ary Frauzino (FAF, Brazil), the Fundaçao PROVITA (Brazil) and the Coordenaçao de Aperfeiçoamento de Pessoal de Nivel Superior (CAPES, Brazil). GD'A was supported by a grant from an FIRC (Fondazione Italiana Ricerca Cancro) fellowship.

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Bonamino, M., Serafini, M., D'Amico, G. et al. Functional transfer of CD40L gene in human B-cell precursor ALL blasts by second-generation SIN lentivectors. Gene Ther 11, 85–93 (2004). https://doi.org/10.1038/sj.gt.3302141

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