Plasma Biomarkers in Graft-versus-Host Disease: A New Era?

Antibody array heatmap of discovery set samples. This heatmap depicts relative protein values obtained from antibody microarrays after the removal of batch effects due to 3 separate analyses. Samples from 21 GVHD^– patients (A) and 21 GVHD⁺ patients (B) are represented. Only the antibodies giving the 35 smallest P values for differences between GVHD⁺ and GVHD^– patient plasma are shown. The P values compare the GVHD⁺ and GVHD^– samples. (Reproduced with permission [29].)

ROC curves of 4 individual discriminator proteins and the composite panel in the training set. Individual ROC curves for IL-2Rα, TNFR1, HGF, and IL-8 and the composite panel. (Reproduced with permission [29]).

Low-risk and high-risk groups correlated with GVHD grade. The blue boxes represent the low-risk groups, and the red boxes represent the high-risk groups, in the training (A) and validation (B) sets. The solid blue represents GVHD grade 0; the solid red, GVHD grade I-IV.

NRM and OS stratified by the biomarker panel in the training set (A) and validation set (B). In (A), the cumulative incidence of NRM and OS (determined by Kaplan-Meier) are plotted according to the predicted probability of acute GVHD: low (—; n = 193) and high (---; n = 89) (P = .001 and .006, adjusted for age, donor type, HLA match, and intensity of conditioning, for differences in NRM and OS, respectively). NRM at 3.5 years is 15% (95% CI = 9% to 21%) for the low-risk group and 36% (95% CI = 24% to 48%) for the high-risk group. OS at 3.5 years is 53% (95% CI = 45% to 63%) for the low-risk group and 33% (95% CI = 22% to 48%) for the high-risk group. In (B), the cumulative incidence of NRM and OS of the 2 groups are plotted for the validation set: low (—; n = 93) and high (---; n = 49) (P < .001 and .02, adjusted as before, for differences in NRM and OS, respectively). NRM at 3.5 years is 11% (95% CI = 4% to 19%) for the low-risk group and 38% (95% CI = 23% to 53%) for the high-risk group. OS at 3.5 years is 59% (95% CI = 49% to 72%) for the low-risk group and 44% (95% CI = 31% to 63%) for the high-risk group. (Reproduced with permission [29].)

Intact protein analysis system workflow and in-depth analysis of plasma proteins. Plasma pooled from 10 patients with GVHD was labeled with the heavy isotope and compared with plasma pooled from 10 patients with no GVHD labeled with the light isotope. The specimens were then subjected to extensive fractionation (by ion-exchange chromatography and reverse-phase chromatography) before individual fractions were analyzed. This decreased the complexity of individual fractions subjected to analysis by liquid chromatography-MS/MS. (Adapted from [30].)