title: Viral interference during simultaneous transduction with two independent helper-free retroviral vectors.
abstract: The ability to stably transduce a single cell with two independent retroviral vectors would have distinct advantages for gene therapy. We determined that cells can be transduced with two distinct retroviral vectors and have quantitated transduction efficiencies in cells infected sequentially and simultaneously. Two amphotropic, helper virus-free, retroviral vectors, a murine Moloney sarcoma virus-based vector containing the nuclear beta-galactosidase and neomycin resistance genes (MMSVn beta-gal/neoR) and a Harvey virus-derived vector containing the human multidrug resistance gene (HaMDR) were introduced into NIH-3T3 cells, pig keratinocytes, and primary pig fibroblasts simultaneously and sequentially. Analytical flow cytometry was utilized to determine retroviral transduction efficiency by assessing the percentage of cells transduced by either one or both retroviruses, in the absence of selection. Simultaneous retroviral transductions were infrequent events. In addition, transduction of previously infected cells (sequential transductions) occurred at lower than expected frequencies. Our data suggest that there is quantifiable viral interference in sequential retroviral transductions. This interference occurs by a mechanism that appears to be independent of the amphotropic retroviral receptor. Thus, such dual transductions will likely require in vitro selection or the use of a single retrovirus which contains both desired genes on the same genome.
authors.affiliation: Dermatology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-1908, USA.
authors.forename: P S
authors.initials: PS
authors.lastname: Walker
authors.forename: U R
authors.initials: UR
authors.lastname: Hengge
authors.forename: M C
authors.initials: MC
authors.lastname: Udey
authors.forename: I
authors.initials: I
authors.lastname: Aksentijevich
authors.forename: J C
authors.initials: JC
authors.lastname: Vogel
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002460:Cell Line; D005347:Fibroblasts / Q000502:physiology; D019450:Genes, MDR; D005822:Genetic Vectors; D006251:Harvey murine sarcoma virus / Q000235:genetics*; D006378:Helper Viruses / Q000235:genetics; D006801:Humans; D015603:Keratinocytes / Q000502:physiology; D051379:Mice; D008980:Moloney murine sarcoma virus / Q000235:genetics*; D013552:Swine; D013997:Time Factors; D014161:Transduction, Genetic; D014762:Viral Interference; D001616:beta-Galactosidase / Q000235:genetics
chemical_list: D001616:beta-Galactosidase