Anti-ICOS inhibits GVHD mediated by alloantigen-specific Tg T cells. (A) CB6 mice were sublethally irradiated (6.5 cGy Cs), infused with 2C and TEa LN adjusted for Tg T-cell content (4 × 10⁶ T cells each), and irrelevant mAb (•) or anti-ICOS (○) was administered from day -1 through day +28. Radiation controls (^*) did not receive Tg T cells or Ab. Survival is shown (n = 8/group; P = .0002, irrel mAb vs anti-ICOS). (B) Spleens were harvested from mice described in panel A on day 4, counted, and proportion of Tg T cells was determined by flow cytometric analysis. Shown is average of absolute counts of 2C CD8⁺ (left) and TEa CD4⁺ Tg T cells (right) per spleen ± 1 SEM (n = 19/group, pool of 4 separate experiments; P = .0027 for 2C CD8⁺, P = .0062 for TEa CD4⁺, rIgG, ▪, vs anti-ICOS, ▦). (C) Spleens from mice treated as described in panels A-B were phenotyped for indicated cell surface activation antigens and intracellular cytotoxic effector molecule (granzyme B). 2C (left column) and TEa Tg cells (right column) were gated on to obtain histograms. Dotted line indicates negative control. Bold solid line and thin solid line indicate anti-ICOS and rIgG treatment, respectively. The dashed line shown in the CD28, L Sel, and CD44 histograms indicates the level of expression on freshly harvested Tg T cells. Shown is a representative sample.