Heterogeneity of chronic graft-versus-host disease biomarkers: association with CXCL10 and CXCR3+ NK cells

Abstract

Chronic graft-versus-host disease (cGVHD) remains one of the most significant long-term complications after allogeneic blood and marrow transplantation. Diagnostic biomarkers for cGVHD are needed for early diagnosis and may guide identification of prognostic markers. No cGVHD biomarker has yet been validated for use in clinical practice. We evaluated both previously known markers and performed discovery-based analysis for cGVHD biomarkers in a 2 independent test sets (total of 36 cases ≤1 month from diagnosis and 31 time-matched controls with no cGVHD). On the basis of these results, 11 markers were selected and evaluated in 2 independent replication cohorts (total of 134 cGVHD cases and 154 controls). cGVHD cases and controls were evaluated for several clinical covariates, and their impact on biomarkers was identified by univariate analysis. The 2 replications sets were relatively disparate in the biomarkers they replicated. Only sBAFF and, most consistently, CXCL10 were identified as significant in both replication sets. Other markers identified as significant in only 1 replication set included intercellular adhesion molecule 1 (ICAM-1), anti-LG3, aminopeptidase N, CXCL9, endothelin-1, and gelsolin. Multivariate analysis found that all covariates evaluated affected interpretation of the biomarkers. CXCL10 had an increased significance in combination with anti-LG3 and CXCL9, or inversely with CXCR3(+)CD56(bright) natural killer (NK) cells. There was significant heterogeneity of cGVHD biomarkers in a large comprehensive evaluation of cGVHD biomarkers impacted by several covariates. Only CXCL10 strongly correlated in both replication sets. Future analyses for plasma cGVHD biomarkers will need to be performed on very large patient groups with consideration of multiple covariates.

Figure 1

Biomarker selection algorithm. Biomarkers for testing in the replication sets were selected from 2 test sets with discovery-based evaluations using proteomic analysis followed by MRM-MS replication, Luminex, and enzymatic assays. Other markers were selected from communication with collaborators (M.-J.H. and S.G.H.) as well as from recent biomarker studies in the literature (Kitko et al). Using the selection criteria as outlined, only 11 biomarkers were evaluated in the replication sets using ELISA, MRM-MS, Luminex, and enzymatic assays.

Figure 2

Receiver operator characteristic curves. (A) For replication set 1 with CXCL10 and anti-LG3 each alone and in combination; the combination gave the highest AUC and (B) for replication set 2 with CXCL10 and CXCL9 each alone and in combination; the combination gave the highest AUC.