Lentivirus Vectors
Lentiviruses are a subclass of retroviruses. The viral genome in the form of RNA is reverse-transcribed when the virus enters the cell to produce DNA, which is then inserted into the genome at a random position by the viral integrase enzyme. Lentiviruses have been adapted as vectors thanks to their ability integrate into the genome of non-dividing as well as dividing cells. The vector can be used to provide highly effective gene therapy as lentiviruses can change the expression of their target cell’s gene for up to six months. They can be used for nondividing or terminally differentiated cells such as neurons, macrophages, hematopoietic stem cells, retinal photoreceptors, and muscle and liver cells, cell types for which previous gene therapy methods could not be used. The only cells lentiviruses cannot gain access to are quiescent cells (in the G0 state) because this blocks the reverse transcription step.
For safety reasons lentiviral vectors never carry the genes required for their replication. Lentiviral vectors are usually created in a transient transfection system in which a cell line is transfected with three separate plasmid expression systems. These include the transfer vector plasmid ( portions of the HIV provirus), the packaging plasmid or construct, and a plasmid with the heterologous envelop gene (ENV) of a different virus. The three-plasmid components of the vector are put into a packaging cell which is then inserted into the HIV shell. The virus portions of the vector contain insert sequences so that the virus cannot replicate inside the cell system.