GENE TRANSFER INTO NON DIVIDING CELLS BY MEANS OF HIV

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We have developed the gene transfer system into non-dividing cells using HIV vectors. CD4+HeLa cells were arrested at the G2 phase by gamma irradiation and at the cytometry analysis. These non-dividing cells were incubated with the HIV based vector containing the beta-gal gene and stained with X-gal after two days. Approximately the same numbers of blue cells were detected in both G2 and G1/S arrested CD4+HeLa cells. The transduction efficiency of HIV vectors on these non-dividing cells was comparable to that on dividing cells. Moreover, we analyzed the integration sites of HIV vectors in non-dividing cells by the linker mediated PCR (LMPCR) technique. The junction sequences chracteristic for retrovirus integration was detected in some clones, indicating that HIV vectors are able to integrate stably into the chromosome of non-dividing cells.The next step, we have developed the two step gene transfer system using an adenoviral vector containing the CD4 gene and HIV vectors to expand the host range of HIV vectors. Adenoviral mediated transient expression of CD4 was efficiently render various non-T cells susceptible to HIV mediated stable gene transfer. Since both adenovirus and HIV vectors can transduce non-dividing cells, the combination of these two vectors may be used as a general strategy for gene transfer into non-dividing cells. Therefor, HIV vector may be useful not only for gene therapy of AIDS but also for a variety of gene therapy protocols targeting non-dividing cells.