Stable Integration of Human Immunodeficiency Virus-Based Retroviral Vectors into the Chromosomes of Nondividing Cells

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Abstract
Human immunodeficiency virus type 1 (HIV-1)-based vectors are thought to be useful for gene transfer into nondividing cells. We examined whether HIV vectors can really integrate into the chromosomes of nondividing cells. CD4+HeLa cells arrested at the G2 or G1/S phase were incubated with the HIV vector pseudotyped with the HIV envelope. The transduction efficiency of the HIV vector in these nondividing cells was comparable to that in proliferating cells. Sequencing of the polymerase chain reaction-amplified fragments containing the junction sites showed that the HIV vector was stably integrated into the chromosomal DNA. It was also demonstrated that terminally differentiated human macrophages and nonproliferating NT neurons could be transduced by the HIV vector after adenovirus-mediated expression of CD4. These results suggest that the HIV vector may be useful not only for gene therapy of AIDS but also for a variety of gene therapy protocols targeting nondividing cells. Human immunodeficiency virus (HIV) vectors pseudotyped with the vesicular stomatitis virus (VSV) envelope were shown to infect nondividing cells. However, it was not completely clear whether the HIV vector integrates into the host cell genome. In this work, nondividing CD4+ cells were transduced with HIV vectors pseudotyped with the HIV envelope. The genomic DNA fragments containing the junction sites were amplified by linker-mediated polymerase chain reaction, cloned into plasmid, and sequenced. The results showed that HIV vector could really integrate as a retroviral vector into the chromosomal DNA of nondividing cells.