Expansion of Immature/Transitional B-Lymphocytes in Patients with Active Chronic Graft-Versus-Host Disease.

Background: Chronic graft-versus-host disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (HSCT) and an important cause of non-relapse mortality. The diagnosis of cGVHD depends mainly on clinical signs and histopathological confirmation. CGVHD is known to impair immune reconstitution. Recently, we observed that percentages of both non-class-switched (CD19⁺/IgD⁺/CD27⁺) and class-switched memory B-lymphocytes(CD19⁺/IgD⁻/CD27⁺) were significantly lower in patients with active cGVHD when compared to patients never experiencing cGVHD (p=0.003 and p= 0.001). The ratio of immature/memory B-lymphocytes (CD21⁻/CD27⁺) was significantly higher in patients with active cGVHD when compared to patients never experiencing cGVHD (p=0.0046). In the current study we investigated the influence of cGVHD on the pattern of immune reconstitution of B-cell subpopulations within the first years after HSCT.

Methods: Ninety-nine patients (median age 42 years, range 17–62 years) were analyzed 3 months to 12 years after HSCT. The total of 212 sampling events included 141 samples obtained during active cGVHD and 71 samples from time points without cGVHD. Evidence of cGVHD in at least one organ as defined by the NIH Consensus Development Project was required to assign the sampling event into the active cGVHD group. The series included 10 patients (3 without and 7 developing cGVHD) assessed serially in 3 months’ intervals within the first years after HSCT. Peripheral blood leukocytes were analyzed by multiparameter flow cytometry after staining for CD19, staining for surface Ig and the B-lymphocyte memory marker CD27 as well as staining for CD21, which is absent on immature/transitional B-lymphocytes. The patients were scored for cGVHD activity according to the NIH Consensus Development Project criteria at every sampling event.

Results: While the CD21⁻/CD27⁺ ratio as well as the percentage of immature/transitional B-lymphocytes decreased in the first years after HSCT in patients without cGVHD, these parameters remained significantly higher over the years in patients with active cGVHD (p=0.001). In logistic regression analysis a higher CD21⁻/CD27⁺ ratio and higher percentage of immature B-lymphocytes (CD21⁻) significantly correlated with active cGVHD. The odds ratio increased from 1.028 to 14.532 between the second and seventh year after HSCT. Until the end of the first year after HSCT serially sampled patients without cGVHD and the ones with cGVHD responding to immunosuppressive therapy presented a decrease of immature/transitional B-lymphocytes and increase of class-switched memory B cells whereas this pattern was disturbed in all serial patients with refractory cGVHD.

Conclusions: Our study demonstrates that cGVHD is associated with an impairment of the reconstitution of the B-lymphocyte compartment leading to a long-lasting expansion of immature/transitional B-lymphocytes. Prospective studies with larger patient numbers analyzed serially are warranted for further assessment of the role of B-cell subpopulations in the pathogenesis of cGVHD and as potential biomarkers for cGVHD activity.