Fig 5.

Lentiviral transduction of human T cells. A, Schematic representation of the self-inactivating (SIN) perforin lentiviral vector. Δ marks SIN deletion with partially deleted U3 of the 3′ long terminal repeat. ψ, Packaging signal; PGK, phosphoglycerate kinase promoter; ppt, central polypurine tract; SD/SA, splice donor/splice acceptor; U3/R/U5, long terminal repeat elements; WPRE, woodchuck hepatitis virus posttranscriptional regulatory element. B, Representative CD8 T cell phenotype in a patient with FHL-2 in remission and an age-matched healthy control subject before and 5 days after stimulation (day 2) with different multiplicity of infection (MOI). GFP expression can be verified in all CD8 T cell subsets. Results were gated on CD8 T cells. C, Perforin and GFP expression in CD8 T cells in the same patient 72 hours after lentiviral transduction (MOI = 75; for healthy control subjects, see Fig E2, A). A transduction efficiency of 34% verified by GFP expression was achieved. Of CD8 T cells, 20.3% express perforin. D, Cr⁵¹ release assay in the same patient showed improved cytotoxicity in the patient transduced with different MOIs. MOI 75 = GFP, 34%; perforin, 20% (Fig 5, C); MOI 100 = GFP, 41%; perforin, 25%.