HIGH-EFFICIENCY GENE-TRANSFER INTO NORMAL AND ADENOSINE DEAMINASE-DEFICIENT T-LYMPHOCYTES IS MEDIATED BY TRANSDUCTION ON RECOMBINANT FIBRONECTIN FRAGMENTS

Primary human T lymphocytes are powerful targets for genetic modificat ion, although the use of these targets in human gene therapy protocols has been hampered by low levels of transduction, We hare shown previo usly that significant increases in the transduction of hematopoietic s tem and progenitor cells with retroviral vectors can be obtained by th e colocalization of the retrovirus and target cells on specific fibron ectin (FN) adhesion domains (H. Hanenberg, X. L. Xiao, D. Dilloo, K. H ashino, I. Kato, and D. A. Williams, Nat. Med. 2:876-882, 1996). We st udied the transfer of genes into primary T lymphocytes by using FN-ass isted retroviral gene transfer. Activated T lymphocytes were infected for three consecutive days on the recombinant FN fragment CH-296 with a retroviral vector encoding the murine B7-1 protein. Transduced lymph ocytes were analyzed for murine B7-1 expression, and it was found that under optimal conditions, 80 to 89% of the CD3(+) lymphocytes were tr ansduced. Gene transfer was predominantly augmented by the interaction between VLA-4 on the T lymphocytes and the FN adhesion site CS-1. Ade nosine deaminase (ADA)-deficient primary T lymphocytes transduced on C H-296 with a retrovirus encoding murine ADA (mADA) exhibited levels of mADA activity severalfold higher than the levels of the endogenous hu man ADA protein observed in normal human T lymphocytes. Strikingly, th e long-term expression of the transgene was dependent on the activatio n status of the lymphocytes. This approach will have important applica tions in human gene therapy protocols targeting primary T lymphocytes.