Abstract
Integration-deficient lentiviral vectors (IDLVs) are promising gene delivery tools that retain the high transduction efficiency of standard lentiviral vectors, yet fail to integrate as proviruses and are instead converted into episomal circles. These episomes are metabolically stable and support long-term expression of transgenes in non-dividing cells, exhibiting a decreased risk of insertional mutagenesis. We have embarked on an extensive study to compare the transduction efficiency of IDLVs pseudotyped with different envelopes (vesicular stomatitis, Rabies, Mokola and Ross River viral envelopes) and self-complementary adeno-associated viral vectors, serotype-9 (scAAV-9) in spinal cord tissues after intraspinal injection of mouse embryos (E16). Our results indicate that IDLVs can transduce motor neurons (MNs) at extremely high efficiency regardless of the envelope pseudotype while scAAV9 mediates gene delivery to ~40% of spinal cord motor neurons, with other non-neuronal cells also transduced. Long-term expression studies revealed stable gene expression at 7months post-injection. Taken together, the results of this study indicate that IDLVs may be efficient tools for in utero cord transduction in therapeutic strategies such as for treatment of inherited early childhood neurodegenerative diseases.
Highlights
A key factor in the success of gene therapy is the development of delivery systems that are capable of efficient gene transfer without causing pathogenic effects
We show that Integration-deficient lentiviral vectors (IDLVs) produce better and more selective transduction of motor neurons in comparison to scAAV9, which displayed significant non-neuronal transduction
Intraspinal injection showed robust eGFP fluorescence along the entire spinal cord, with expression observed in spinal ganglia; IDLV-Rabies produced GFP fluorescence in the intercostal nerves
Summary
A key factor in the success of gene therapy is the development of delivery systems that are capable of efficient gene transfer without causing pathogenic effects. Lentiviral vectors, engineered forms of retroviruses from the genus Lentivirus, are promising tools for gene therapy because they transduce dividing and quiescent cells efficiently, their tropism can be changed by replacing the natural envelope of lentiviruses with a heterologous envelope (referred to as pseudotyping) and they lead to strong and long-lasting expression of transgenes [1]. During the transduction process lentiviral vectors integrate their genome in the host cell DNA, which can lead to insertional mutagenesis. Engineering of self-inactivating (SIN) lentiviral vectors by deletion of the U3 region in the 3′-LTR is considered to minimize the probability of interactions between the transfer vector elements and the host genome and the incidence of insertional mutagenesis-related events in transduced cells [5]. Studies with SIN lentiviral vectors containing enhancer-less internal promoters resulted in safety improvements in different preclinical studies [9,10,11,12,13]
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