Insertional transformation of hematopoietic cells by self-inactivating lentiviral and gammaretroviral vectors.
Gene transfer vectors may cause clonal imbalance and even malignant cell transformation by insertional upregulation of proto-oncogenes. Lentiviral vectors (LV) with their preferred integration in transcribed genes are considered less genotoxic than gammaretroviral vectors (GV) with their preference for integration next to transcriptional start sites and regulatory gene regions. Using a sensitive cell culture assay and a series of self-inactivating (SIN) vectors, we found that the lentiviral insertion pattern was approximately threefold less likely than the gammaretroviral to trigger transformation of primary hematopoietic cells. However, lentivirally induced mutants also showed robust replating, in line with the selection for common insertion sites (CIS) in the first intron of the Evi1 proto-oncogene. This potent proto-oncogene thus represents a CIS for both GV and LV, despite major differences in their integration mechanisms. Altering the vectors' enhancer-promoter elements had a greater effect on safety than the retroviral insertion pattern. Clinical grade LV expressing the Wiskott-Aldrich syndrome (WAS) protein under control of its own promoter had no transforming potential. Mechanistic studies support the conclusion that enhancer-mediated gene activation is the major cause for insertional transformation of hematopoietic cells, opening rational strategies for risk prevention.
Modlich
Ute
U
Department of Experimental Hematology, Hannover Medical School, Hannover, Germany.
Navarro
Susana
S
Zychlinski
Daniela
D
Maetzig
Tobias
T
Knoess
Sabine
S
Brugman
Martijn H
MH
Schambach
Axel
A
Charrier
Sabine
S
Galy
Anne
A
Thrasher
Adrian J
AJ
Bueren
Juan
J
Baum
Christopher
C
D000818:Animals; D015152:Blotting, Northern; D001854:Bone Marrow Cells / Q000378:metabolism* / Q000821:virology; D017988:Gammaretrovirus / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D016254:Mutagenesis, Insertional / Q000379:methods*; D016133:Polymerase Chain Reaction; D011519:Proto-Oncogenes / Q000235:genetics; D014161:Transduction, Genetic / Q000379:methods*