Expression profile of genes flanking the LC15 retroviral vector integration sites. (A) Semiquantitative RT-PCR was performed on RNA extracted from LC15 or control PBLs that were obtained from the same patient used to generate LC15. CD8 cells were isolated from PBLs by negative selection, and RNA was extracted immediately after purification or after an 8-hour exposure to 10 ng IL-15/mL. A fraction of the PBLs were stimulated with OKT3 and cultured with IL-15 (10 ng/mL) for 7 days prior to RNA extraction. Primers were designed to amplify ACVR1C, PSCDBP, FOXP1, PGBD4, and C15orf29 which are genes flanking the retroviral integration sites in LC15. 18S rRNA primers were used as an internal control. (B) Gene expression was also examined using a cDNA array. Untreated CD8⁺ T cells were used as a reference for all other samples. Gene expression in IL-15–exposed CD8⁺ cells (left), OKT3-stimulated CD8⁺ cells cultured in IL-15 (10 ng/mL) for 7 days (middle), and LC15 (right) is depicted. The abscissa represents the relative gene expression minus background in the CD8⁺ cDNA control. The ordinate represents the relative gene expression minus background of the experimental conditions listed above. The darkened lines on each plot delineate 2-fold differential gene expression. The darkened spots represent ACVR1C, PSCDPB, FOXP1 splice variant 1, FOXP1 splice variant 2, and 2 unique oligos corresponding to C15orf29. OKT3-stimulated CD8⁺ cells cultured in IL-15, and LC15 exhibited down-regulation of splice variant 1 of FOXP1.