Neddylation blockade regulates DC-mediated T-cell activation. (A) Survival of BALB/C animals lethally irradiated and transplanted with BM and CD90.2⁺ T cells from either syngeneic BALB/C or allogeneic C57BL/6 donors. Following lethal irradiation (8 Gy) on day 1, MLN4924 (20 mg/kg) was administered in 5 daily doses, day 1 to day 3 relative to BMT. Data are combined from 2 independent experiments (n = 10 to 12 animals in the allogeneic groups). (B-C) Abb animals were lethally irradiated (10 Gy) on day 1 and WT B6 BMDC (10 × 10⁶) pretreated with vehicle or MLN4924 overnight and subsequently transferred in 2 doses separated by 24 hours (day 1 and day 0). CD90.2⁺ T cells (2 × 10⁶) from syngeneic WT-C57BL/6 or allogeneic bm12 animals were transferred on day 0. Following sacrifice on day 6, spleens were analyzed for CD69 (B) and Tbet (C). (D) Quantification via ELISA of TNF-α (left) and IL-6 (right) released from human moDCs stimulated with LPS in the presence or absence of MLN4924 or dexamethasone at the indicated doses. Cells receiving treatment were preincubated with vehicle, MLN4924, or dexamethasone for 2 hours followed by concurrent LPS stimulation (0.5 µg/mL) for 4 hours. One representative experiment of 3 is shown of groups treated in triplicate. (E) Human PBMCs (1 × 10⁵/well) and moDCs were obtained from 2 healthy donors and cocultured at 1:1 and 10:1 ratios in an MLR in the presence or absence of MLN4924 for 96 and 120 hours.