BAFF/Blys Levels Correlate with Disease Activity and Alter Peripheral B Cell Subsets in Patients with Chronic GVHD.

Patients with chronic graft versus host disease (cGVHD) after allogeneic hematopoietic stem cell transplantation (HSCT) have been found to have high titers of allo-reactive antibodies, but a role for B cells in the pathology of this disease remains undefined. B Cell Activating Factor, BAFF/Blys (BAFF), promotes differentiation and expansion of antigen-activated B cells and contributes to loss of B cell tolerance in animal models. We hypothesized that the BAFF/BAFF receptor (BAFF-R) pathway may perpetuate potentially allo- or auto-reactive antigen-experienced CD27+ B cells in patients with cGVHD after HSCT. Soluble BAFF was measured in plasma from 104 patients after HSCT. Median BAFF levels were statistically different when groups were compared using a two-sided Wilcoxon-Rank-Sum test (see Table below). Logistic regression analysis revealed that higher BAFF levels were associated with active cGVHD after adjusting for other GVHD prognostic factors (p=0.0007). Patients with ≥10ng/ml BAFF levels had ten-fold increased odds of having cGVHD compared to patients with BAFF levels of <10ng/ml (OR of 10.8, p=<0.0001). High dose prednisone reduced median plasma BAFF levels (3.8ng/ml in patients receiving ≥30mg daily prednisone versus 14ng/ml for those receiving <30mg or no prednisone). Serial BAFF measurements revealed peak BAFF levels at 6 months post-HSCT in patients who later developed limited cGVHD. 81% of patients with BAFF levels >10ng/ml at 6 months subsequently developed cGVHD (median BAFF was 20ng/ml) compared to 39% of patients with BAFF levels <10ng/ml at 6 months (p=0.002). We also found that BAFF-R expression on B cells was down-regulated in vitro in the presence of BAFF. Consistent with this finding flow cytometry revealed very low BAFF-R expression on B cells in patients with active cGVHD. BAFF-R expression on peripheral B cells correlated with BAFF levels (p=0.0001), suggesting that BAFF signals via BAFF-R in cGVHD. We used 5-color FACS to characterize peripheral B cell subsets in 68 post-HSCT patients. Compared to patients without cGVHD, the proportion of antigen-experienced CD27+ B cells was increased in patients with limited cGVHD (n=20, p=0.04). The extensive cGVHD patient group was smaller (n=11) with greater variability in CD27+ B cell frequency resulting in no statistical difference (p=0.27). The proportion of CD27+ post-germinal center B cells was also increased in patients with active cGVHD (p=0.04 and p=0.03 extensive and limited cGVHD, respectively). High BAFF levels correlated with increased total numbers of CD27+ B cells (p=0.05), but not with total or naïve B cell numbers, suggesting that BAFF plays a role in perpetuation of circulating antigen-experienced and memory B cells in cGVHD patients. Our results suggest that high levels of BAFF after HSCT help break peripheral B cell tolerance and contribute to cGVHD pathobiology.