CD8⁺T-cell gene-expression signature at aGVHD onset. For this analysis, samples from the 2 cohorts were pooled. All recipients without aGVHD or with aGVHD grade ≥2 were included in the analysis (No GVHD, n = 36; GVHD, n = 22). (A) Gene-expression profiles of CD8⁺ T cells from 22 patients at aGVHD onset compared with 36 patients without aGVHD in cohorts 1 and 2. Orange and light blue dots represent transcripts that were significantly up- or downregulated in recipients at aGVHD onset compared with No GVHD recipients, respectively, with FDR < 0.05. P values were calculated using an unpaired Student t test. Adjusted P values (FDR) were calculated using the Benjamini-Hochberg method to correct for multiple comparisons. (B) Transcript levels of NFKB1, BCL3, NFKBIA, ICOS, CD28, MIF, and GAPDH in CD8⁺ T cells from recipients without aGVHD and at aGVHD onset. These genes showed increased expression at aGVHD onset compared with recipients without GVHD (P < .05; FDR < 0.05). (C) QuSAGE of CD8⁺ T-cell gene-expression profiles in recipients at aGVHD onset compared with recipients without aGVHD from cohorts 1 and 2. For each pathway, the mean fold change and the 95% confidence intervals are plotted and color-coded according to their FDR-corrected P values compared with 0. Red and green bars indicate a statistically significant increased or decreased pathway activity, respectively, in recipients at aGVHD onset compared with recipients without GVHD. (D) Transcript levels of TGFBR2, SMAD3, IGFR2, LAIR1, BTLA, KLRG1, and IL10RA in CD8⁺ T cells from recipients without aGVHD and at aGVHD onset. These genes showed decreased expression at aGVHD onset compared with recipients without aGVHD (P < .05; FDR < 0.05). P values were calculated using an unpaired Student t test. Adjusted P values (FDR) were calculated using the Benjamini-Hochberg method to correct for multiple comparisons. Nominal P values are shown in panels C and D. Tfh, T follicular helper cell.