THERAPEUTIC GENE DELIVERY IN HUMAN BETA-LYMPHOBLASTOID CELLS BY ENGINEERED NON-TRANSFORMING INFECTIOUS EPSTEIN-BARR-VIRUS

The B-lymphotrophic human herpes Epstein-Barr virus (EBV) is a 160-kil obase double-stranded DNA episomal virus carried in a persistent asymp tomatic state by more than 90% of the worldwide adult population. We e ngineered a helper-dependent mini-EBV, with the minimal cis-EBV elemen ts for episomal replication, viral amplification and packaging, for us e as a gene delivery system. The therapeutic potential of this system was established by stably transducing B-lymphoblastoid cells from a Fa nconi anaemia group C (FA-C) patient with a mini-EBV constitutively ex pressing the normal FACC cDNA and showing in vitro correction of the F A phenotype. In the absence of selective pressure, episomal expression persisted with a half-life of 30 days in actively growing transduced cells, indicating a retention rate of 98% expression per cell doubling . This work demonstrates the generation of an infectious non-transform ing viral vector that can potentially deliver large therapeutic genes efficiently and selectively into human B cells.