RECOMBINASE-DNA NUCLEOPROTEIN FILAMENTS AS GENE-THERAPY VECTORS

In december 1991, we devised a new approach to Gene Therapy based on p remade DNA-recombinase nucleoprotein filaments. Aimed at repairing mut ant genes (inherited diseases) and at inactivating deleterious or vita l viral genes (cancer, hepatitides, AIDS), the idea is to generate, in vitro, human recombinase-DNA complexes analogous to the bacterial Rec A presynaptic filament and mimic, upon transfection, recombinational D NA repair that occurs in most dividing mammalian cells. The aim is to substitute chromosomal DNA segments by exogenous homologous genomic DN A from wild type (gene repair) or mutant(gene inactivation) origin. Ou r actual strategies deal with human RAD51 protein that has been shown to mediate in vitro, in the presence of replication protein A, the ATP -dependent homologous DNA pairing-strand exchange reaction. We describ e how, in order to avoid unspecific targeting to repetitive chromosoma l elements and to bypass potential arrests of the homologous DNA pairi ng-strand exchange reaction by therapeutic heterologous bases, we have designed presynaptic filaments that comprise a double-stranded DNA co re. From transfecting cell lines in vitro toward targeting specific so matic cells in vivo, our ultimate goal would be to develop a recombina se-mediated gene therapy that would substitute to the present minigene expression approach.