Transcriptional enhancers induce insertional gene deregulation independently from the vector type and design.
The integration characteristics of retroviral (RV) vectors increase the probability of interfering with the regulation of cellular genes, and account for a tangible risk of insertional mutagenesis in treated patients. To assess the potential genotoxic risk of conventional or self-inactivating (SIN) gamma-RV and lentiviral (LV) vectors independently from the biological consequences of the insertion event, we developed a quantitative assay based on real-time reverse transcriptase--PCR on low-density arrays to evaluate alterations of gene expression in individual primary T-cell clones. We show that the Moloney leukemia virus long terminal repeat (LTR) enhancer has the strongest activity in both a gamma-RV and a LV vector context, while an internal cellular promoter induces deregulation of gene expression less frequently, at a shorter range and to a lower extent in both vector types. Downregulation of gene expression was observed only in the context of LV vectors. This study indicates that insertional gene activation is determined by the characteristics of the transcriptional regulatory elements carried by the vector, and is largely independent from the vector type or design.
Maruggi
Giulietta
G
Department of Biomedical Sciences, University of Modena and Reggio Emilia, Modena, Italy.
Porcellini
Simona
S
Facchini
Giulia
G
Perna
Serena K
SK
Cattoglio
Claudia
C
Sartori
Daniela
D
Ambrosi
Alessandro
A
Schambach
Axel
A
Baum
Christopher
C
Bonini
Chiara
C
Bovolenta
Chiara
C
Mavilio
Fulvio
F
Recchia
Alessandra
A
D004742:Enhancer Elements, Genetic / Q000235:genetics* / Q000502:physiology; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008979:Moloney murine leukemia virus / Q000235:genetics; D016254:Mutagenesis, Insertional / Q000235:genetics*; D012190:Retroviridae / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020079:Terminal Repeat Sequences / Q000235:genetics / Q000502:physiology