Effective gene therapy with nonintegrating lentiviral vectors

Abstract

Retroviral and lentiviral vector integration into host-cell chromosomes carries with it a finite chance of causing insertional mutagenesis. This risk has been highlighted by the induction of malignancy in mouse models, and development of lymphoproliferative disease in three individuals with severe combined immunodeficiency-X1 (refs. 2,3). Therefore, a key challenge for clinical therapies based on retroviral vectors is to achieve stable transgene expression while minimizing insertional mutagenesis. Recent in vitro studies have shown that integration-deficient lentiviral vectors can mediate stable transduction. With similar vectors, we now show efficient and sustained transgene expression in vivo in rodent ocular and brain tissues. We also show substantial rescue of clinically relevant rodent models of retinal degeneration. Therefore, the high efficiency of gene transfer and expression mediated by lentiviruses can be harnessed in vivo without a requirement for vector integration. For therapeutic application to postmitotic tissues, this system substantially reduces the risk of insertional mutagenesis.

Absence of ocular malignant transformation after sub-retinal delivery of rAAV2/2 or integrating lentiviral vectors in p53-deficient mice

“…49 VSV-G pseudotyped HIV-1.SFFV.hrGFP vectors (5Â10 8 TU ml À1 ) were generated and titrated as previously described. 24 …”

“…Immunostaining for RPE65 was performed on 4% paraformaldehyde-fixed 12 mm thick propidium iodide-counterstained cryosections, and RPE65 immunofluorescence detected using fluorescence microscopy as previously described. 24 …”

“…The clinical success of the RPE65 trials, combined with the increasing number of successful pre-clinical studies using rAAV [18][19][20][21] or lentiviruses, [22][23][24][25][26][27][28] suggest that more retinal gene therapy clinical trials are likely to start in the near future. It is, therefore, crucial to determine the oncogenic potential of these gene delivery vehicles in the context of the retina.…”

Tightly regulated gene expression in human hematopoietic stem cells after transduction with helper-dependent Ad5/35 vectors

“…Gene transfer into HSCs by transfection or electroporation is hampered by low efficiency and high cytotoxicity. Nonintegrating lentivirus vector have shown promise for transient expression of transgenes in HSC but suffer from relatively low transduction and expression levels [3]. We have focused on nonintegrating adenovirus (Ad) vectors for gene transfer into human HSC.…”

SIVMAC Vpx improves the transduction of dendritic cells with nonintegrative HIV-1-derived vectors

“…To circumvent this problem, several groups have evaluated the use of integration-incompetent LVs. [3][4][5][6][7][8] Gene expression from these vectors is driven from viral DNA molecules that remain in an episomal form after their entry into the nucleus as 1 or 2 long terminal repeat circles (1-or 2-LTRs, respectively). These episomes are considered as a deadend product in the normal retroviral infection process, but they are a functional substrate for the transcriptional machinery.…”