Spontaneous EBV-Reactivation during B Cell Differentiation as a Model for Polymorphic EBV-Driven Lymphoproliferation

Epstein-Barr virus (EBV)-driven B cell neoplasms arise from the reactivation of latently infected B cells. In a subset of patients, EBV was seen to drive a polymorphous lymphoproliferative disorder (LPD) in which B cell differentiation was retained. In this work, spontaneous EBV reactivation following B cell mitogen stimulation was shown to provide a potential model of polymorphic EBV-driven LPD. Here, we developed an in vitro model of plasma cell (PC) differentiation from peripheral blood memory B cells. To assess the frequency and phenotypes of EBV-associated populations derived during differentiation, we analysed eight differentiations during the PC stage with a targeted single-cell gene expression panel. We identified subpopulations of EBV-gene expressing cells with PC and/or B cell expression features in differentiations from all tested donors. EBV-associated cells varied in frequency, ranging from 3-28% of cells. Most EBV-associated cells expressed PC genes such as XBP1 or MZB1, and in all samples these included a quiescent PC fraction that lacked cell a cycle gene expression. With increasing EBV-associated cells, populations with B cell features became prominent, co-expressing a germinal centre (GC) and activating B cell gene patterns. The presence of highly proliferative EBV-associated cells was linked to retained MS4A1/CD20 expression and IGHM and IGHD co-expression, while IGHM class-switched cells were enriched in quiescent PC fractions. Thus, patterns of gene expression in primary EBV reactivation were shown to include features related to GC B cells, which was also observed in EBV-transformed lymphoblastoid cell lines. This suggests a particular association between spontaneously developing EBV-expansions and IgM+ IgD+ non-switched B cells.