New vectors for gene therapy

Retrovirus-based vectors are presently the most efficient gene transfer vehicles for introducing genes into human hematopoietic stem and progenitor cells. However, their use for gene therapy is still problematic. A major obstacle is viral sequences such as the tRNA primer binding site or the dimerization and encapsidation signals that are not required for the expression of the therapeutic gene. These sequences can recombine with endogenous and/or exogenous retroviruses to generate new forms of unpredictable retroviruses. Moreover, these sequences are the targets for transcriptional repressors which inhibit the expression of the transduced genes. Therefore we have developed a new generation of retrovirus vectors which self-delete upon integration. The vectors are based on the natural life cycle of retroviruses, involving duplication of the terminal control regions U5 and U3 to generate long terminal repeats, and on the ability of the P1-phage site-specific recombinase (Cre) to excise any sequences positioned between two target sequences (loxP). Thus, while inserting a therapy gene into the genome, the vectors simultaneously excise most proviral sequences that are not required for gene expression.