Inflammatory Markers and Autoantibodies That Correlate with Early and Late Onset of New Onset Pediatric Chronic Graft-Versus-Host Disease (GVHD).
Authors: Fujii Hisaki , She Kevin , Aslanian Soudabeh , Shimizu Hiromi , Krailo Mark , Chen Zhengjia , Goldman Frederick , Wall Donna A. , Gilman Andrew L. , Schultz Kirk R. 
Published: October 2019

Abstract
Chronic graft-vs-host disease (cGVHD) has previously been characterized by elevation of a number of inflammatory markers and autoantibodies. We hypothesized that evaluation of such markers would contribute to a biological classification of new onset cGVHD in children and possibly aid in early diagnosis. The COG study ASCT0031 is a phase III cGVHD therapeutic trial that included the biological evaluation of 52 newly diagnosed cGVHD pts and 34 time-matched post BMT control patients (2 – 18 yrs) with no clinical evidence of cGVHD. Chronic GVHD was divided into an early onset between 3 to 8 mos post BMT (n=37, mean 5.7, range 3 – 8 months ) and late onset ≥9 months post BMT (n=15, mean 14.3, range 9 to 30 months) groups and 34 non-cGVHD controls evaluated at 6 and 12 months. Soluble B-cell activating factor (sBAFF), soluble IL-2 receptor alpha (sIL-2Ralpha), transforming growth factor-beta (TGF-beta), interferon-alpha (IFN-alpha), monocyte chemoattractant protein-1 (MCP-1), IL-6, anti-nuclear antibody (ANA), anti-dsDNA antibody, anti-mitochondrial antibody and anti-cardiolipin antibody were evaluated by sandwich ELISA. We found an increased level of sBAFF in cGVHD patients with both early (7.2± 5.3 vs. 3.7 ± 2.5 ng/mL, p = 0.008, t-test) and late (3.2 ± 2.7 vs. 1.5 ± 0.8 ng/mL, p = 0.04) onset cGVHD compared with controls. Although the anti-dsDNA concentrations were considered to be in the normal range, they were significantly higher in both early (2.4 ± 2.5 vs. 0.1±0.3 IU/mL; p < 0.0001) and late (4.1 ± 3.1 vs. 0.2 ± 0.3 IU/mL; p < 0.0001) cGVHD compared to non-cGVHD controls. We found that early onset cGVHD compared to patients without cGVHD at 6 months had an elevation in sIL-2Ralpha (10.1±5.4 vs. 7.0±3.4 pg/mL; p < 0.0001) but not in late onset cGVHD. ANA, TGF-beta, IFNalpha, IL-6, MCP-1, and anti-cardiolipin antibodies were not associated with new onset cGVHD. Acute GVHD, a major factor for the development of cGVHD, did not correlate with any of the three markers. We found that the ANA (2.3±2.1 vs. 0.9±0.7 absorbance ratio, p = 0.0004) and anti-dsDNA (anti-dsDNA; 3.8±2.9 vs. 2.4±2.5 IU/mL, p = 0.04) were significantly elevated in de novo onset cGVHD patients compared to those with previous acute GVHD. Soluble BAFF and sIL-2Ralpha are associated with Th1 responses and B cell activation/expansion, whereas anti-dsDNA is associated with specific B cell responses. The elevation of sBAFF and sIL-2Ralpha are consistent with both high levels of T cell and B cell activation as part of early cGVHD. The association of later onset cGVHD with only sBAFF and anti-dsDNA suggests that B cell activation is more predominant in that setting and is consistent with our previous observation of an association of cGVHD with activated Toll-like receptor 9 expressing B cells. These findings suggest early and late cGVHD may be classified by different patterns of T and B cell activation.
Early onset cGVHD (3–8 mo. Post BMT) Late Onset cGVHD (>=9 mo. post BMT) Marker cGVHD (N=37) No cGVHD at 6 mo. (N=26) p value cGVHD (N=14) No cGVHD at 12 mo. (N=15) p value Anti-dsDNA (IU/mL) 2.4±2.5 0.1±0.3 <0.001 4.1±3.1 0.2 ± 0.3 <0.001 sBAFF (ng/mL) 7.2± 5.3 3.7 ± 2.5 0.008 3.2 ± 2.7 1.5 ± 0.8 0.04 sIL-2Ralpha (pg/mL) 10.1±5.4 7.0±3.4 <0.001 6.9±3.3 5.6±3.7 NS