The genotoxic potential of retroviral vectors is strongly modulated by vector design and integration site selection in a mouse model of HSC gene therapy.
gamma-Retroviral vectors (gammaRVs), which are commonly used in gene therapy, can trigger oncogenesis by insertional mutagenesis. Here, we have dissected the contribution of vector design and viral integration site selection (ISS) to oncogenesis using an in vivo genotoxicity assay based on transplantation of vector-transduced tumor-prone mouse hematopoietic stem/progenitor cells. By swapping genetic elements between gammaRV and lentiviral vectors (LVs), we have demonstrated that transcriptionally active long terminal repeats (LTRs) are major determinants of genotoxicity even when reconstituted in LVs and that self-inactivating (SIN) LTRs enhance the safety of gammaRVs. By comparing the genotoxicity of vectors with matched active LTRs, we were able to determine that substantially greater LV integration loads are required to approach the same oncogenic risk as gammaRVs. This difference in facilitating oncogenesis is likely to be explained by the observed preferential targeting of cancer genes by gammaRVs. This integration-site bias was intrinsic to gammaRVs, as it was also observed for SIN gammaRVs that lacked genotoxicity in our model. Our findings strongly support the use of SIN viral vector platforms and show that ISS can substantially modulate genotoxicity.
Montini
Eugenio
E
San Raffaele-Telethon Institute for Gene Therapy, via Olgettina 58, 20132 Milan, Italy. montini.eugenio@hsr.it
Cesana
Daniela
D
Schmidt
Manfred
M
Sanvito
Francesca
F
Bartholomae
Cynthia C
CC
Ranzani
Marco
M
Benedicenti
Fabrizio
F
Sergi
Lucia Sergi
LS
Ambrosi
Alessandro
A
Ponzoni
Maurilio
M
Doglioni
Claudio
C
Di Serio
Clelia
C
von Kalle
Christof
C
Naldini
Luigi
L
D000818:Animals; D001483:Base Sequence; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000172:deficiency / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D017988:Gammaretrovirus / Q000235:genetics* / Q000472:pathogenicity*; D019942:Genes, p16; D015316:Genetic Therapy / Q000009:adverse effects* / Q000379:methods; D005822:Genetic Vectors / Q000009:adverse effects*; D018380:Hematopoietic Stem Cell Transplantation / Q000009:adverse effects; D016086:Lentivirus / Q000235:genetics / Q000472:pathogenicity; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D009154:Mutation; D009374:Neoplasms, Experimental / Q000209:etiology / Q000235:genetics; D020079:Terminal Repeat Sequences
C417743:Cdkn2a protein, mouse; D019941:Cyclin-Dependent Kinase Inhibitor p16; D017931:DNA Primers