Improved Methods of HIV Vector Mediated Gene Transfer

HIV vectors are capable of targeted gene transfer into CD4+ cells. However, extensive testing of HIV vectors in gene therapy applications is hampered by the low titer of current HIV vector could be concentrated by approximately 20 times by sulfonated cellulose column chromatography. No replication competent cytopathic HIV was detected in the concentrated vector preparation. When the vector preparation and the target cells were centrifuged at transduction, about a five-fold increase in the apparent titer was achieved. Accordingly, by combining these two techniques the overall titer was increased by approximately two orders of magnitude. Using this high efficiency strategy, we transduced human primary lymphocytes that are refractory to transduction with currently available viral vectors. Amplification and sequencing of the integration sites showed that HIV vectors could stably integrate into the chromosomes of CD4 enriched human peripheral blood mononuclear cells. These findings indicate that HIV vectors are useful for the development of gene therapy targeting lymphocytes.