Improvement of gene transduction efficiency in T lymphocytes using retroviral vectors

Successful gene transfer into T lymphocytes would provide a useful therapeu tic modality for the treatment of various diseases and a valuable way to st udy T cell functions. Currently, most protocols involving gene transfer int o T lymphocytes utilize amphotropic retroviral vectors. However, transducti on efficiency using these vectors is relatively low because of the high pro portion of resting cells, the concentration-dependent growth manner of T ly mphocytes, and the low titer of retroviral vectors. In this article we defi ne conditions that provide high levels of transduction by using IL-2 presti mulation and LipofectAMINE for both mouse and human T lymphocytes. We compa red the effects of IL-2 prestimulation on transduction efficiencies at diff erent time points and achieved maximum transfer levels at 72 hr after the i ncubation. By combining the best prestimulation time and cationic lipids-Li pofectAMINE at a dose of 0.8 mu M, the transduction efficiencies were incre ased to 45-75 % (62.3 +/- 4.3 %) in human T lymphocytes and to 21-33 % (27 +/- 1.42%) in murine T lymphocytes as determine by FDG staining and X-Gal v isualization, compared with 5% with conventional methods. These results ind icate that transduction efficiencies in T lymphocytes can be significantly improved by a prolonged preincubation with IL-2 and by the addition of Lipo fectAMINE.