FK778 induces T-cell anergy and suppressive capacity after allogeneic stimulation. PBMCs were stimulated with allogeneic stimulator cells for 7 days. Cells were harvested and rested for 2 to 3 days. Viable cells were recovered by density gradient centrifugation. (A) Proliferation of recovered cells treated with 0 μg/mL (•), 10 μg/mL (▵), or 50 μg/mL (○) FK778 during primary allogeneic stimulation was investigated in a secondary MLC (in the absence of FK778). During the secondary MLC, the recovered cells (2 × 10⁴) were restimulated with allogeneic stimulator cells (10⁵) alone (left panel) or in the presence of 12.5 U/mL IL-2 (right panel); *P < .05 for differences between untreated and FK778 (50 μg/mL)–treated cells. (B) The proliferative capacity of PBMCs primed in the presence of FK506 (left panel), rapamycin (middle panel), or MPA (right panel) was analyzed in a secondary MLC, as described in panel A, in the absence of the respective drugs. (C) The suppressive potential of FK778-treated cells was investigated in a coculture suppression assay. Proliferation of naive PBMCs (5 × 10⁴) stimulated with allogeneic stimulator cells (5 × 10⁴) was studied in the absence or presence of increasing numbers of untreated (•) or FK778 (50 μg/mL)–treated (○) cells (*P < .05; ** P < .01 for differences between untreated and FK778-treated cells. One representative experiment of 4 is presented. (D) PBMCs were stimulated with αCD3/αCD28 mAb–coated beads instead of allogeneic stimulator cells in the presence or absence of FK778. The proliferation of the recovered cells (2 × 10⁴), untreated (•) or treated with 50 μg/mL FK778 (○) during the primary MLC, was examined during the secondary culture. At the peak of proliferation, *P < .01 for differences between untreated and FK778-treated cells. One representative experiment of 3 is presented. Error bars represent SD counts.