Characterization of the CD4+ T-cell response to MAGE-3191-205.
CD4+ T cells were propagated from donor 4 (HLA-DR11) by weekly restimulation with pool II. (A) Proliferative responses, measured in 2-day microproliferation assays, to each single peptide forming pool II (10 μg/mL) after 4 and 9 weeks of propagation of the line. The data shown are means of triplicate determination ± SDs. The blank (ie, the basal level of proliferation of CD4+ T cells in the presence of autologous PBMCs as APCs) is expressed as B + DR11-PBMC. Responses significantly higher than the blanks are indicated: **.001 < P < .05, ***P < .001 (determined by unpaired, one-tailed Student t test). (B) Responses to pool II (5 μg/mL), in the absence or in the presence of L243 mAb or an irrelevant control mAb (0.5 μg/mL). The blank (ie, the basal level of proliferation of CD4+ T cells in the presence of autologous PBMCs as APCs) is expressed as B + APC. (C-F) Recognition of naturally processed MAGE-3 sequences by CD4+ T cells at 9 or more weeks of propagation. (C) Cytolytic activity, measured in a51Cr release assay, against wild-type DR*1104-LCL and DR*1104-LCL engineered to express MAGE-3 (DR*1104-LCL-M3). (D) Cytolytic activity against HLA-DR–matched (MD TC and OI TC) and HLA-DR–unrelated (SK-Mel 24) melanoma cells. (E-F) Cold target inhibition experiments. (E) Cold targets (MD TC, DR*1104-LCL, and DR*1104-LCL pulsed with MAGE-3191-205) were used to inhibit the lytic activity of CD4+ cytotoxic T cells against hot MD TC (effector-target [E/T] ratio 40:1). The percentage of specific lysis against MD TC in the absence of cold target was 22 ± 1.3. (F) Cold targets (DR*1101-LCL and DR*1101-LCL pulsed with MAGE-3191-205 or MAGE-321-35) were used to inhibit the lytic activity of CD4+ cytotoxic T cells against hot OI TC (E/T ratio 40:1). The percents lytic activity against OI TC and DR*1101-LCL, as negative control, are shown in black symbols. The data presented in panels C through F are representative of at least 3 experiments and are means of triplicate determinations ± SDs.
