Induction of CTL responses against peptides derived from viral and tumor antigens using CD40–B cells as APCs.

(A) Peptide-specific lysis of T2 cells pulsed with specific peptide (filled circles), irrelevant HLA-A2–binding control peptide (empty circles), or no peptide (empty squares). Influenza A MP58-specific lysis (MP58) is shown for day 18 of T-cell culture; all other experiments for MART-1 (E26), hTERT (I540), and HIV RTpol (RTpol476) were performed on day 25. For each peptide, a representative peptide-specific CTL line is shown here. (B) Tetramer analysis using peptide-specific tetramers was always controlled with the tetramer specific for the immunodominant HTLV-1 tax peptide. The percentage of CD8⁺ tetramer⁺ T cells is shown. Analysis was performed after 3 (Influenza A [Inf A]) or 4 weeks of CTL expansion (all other antigens). Shown here is a representative peptide-specific CTL line. (C) Endogenous processing was tested for MART-1 by means of A2⁺ MART-1⁺ Malme-3M cells (filled circles) and HLA-A2⁻SK-MEL-2 cells (empty circles); for hTERT by means of HLA-A2⁺ hTERT⁺U266 cells (filled circles) and HLA-A2⁻ HSS cells (empty circles) (second graph), IFN-γ–pretreated (72 hours) HLA-A2⁺ hTERT-transfected U2OS cells (filled circles), or HLA-A2⁺ wild-type U2OS cells (empty circles) (third graph); and for RTpol by means of HLA-A2⁺ RTpol⁺ COS cells (filled circles) and HLA-A*0201⁺ RTpol⁻COS cells (empty circles) as target cells. (D) Generation of peptide-specific CTL lines in healthy donors and cancer patients is expressed as individual T-cell cultures that induced a peptide-specific response out of the total number of T-cell cultures performed. (E) Generation of RTpol476-specific CTL lines using CD40L-matured monocyte-derived DCs. Peptide-specific lysis of T2 cells pulsed with specific peptide (●) and HLA-A2–binding control peptide (○) (upper panel); percentage of CD8⁺tetramer⁺ T cells (middle panel); and recognition of endogenously processed peptide with the use of HLA-A2⁺RTpol⁺ COS cells (●) and HLA-A*0201⁺RTpol⁻ COS cells (○) as target cells (lower panel).