Background and aims Belimumab is a monoclonal antibody that inhibits soluble B lymphocyte stimulator (BLyS, also known as BAFF), approved for the treatment of systemic lupus erythematosus (SLE). Here, we sought to identify B cell alterations during belimumab treatment.

Methods Twenty-three SLE patients treated with belimumab were enrolled. Peripheral blood mononuclear cells were collected and cryopreserved at treatment initiation and at regular follow-up visits for up to 3 years. The samples were simultaneously assayed using mass cytometry. Cell counts were adjusted for total lymphocyte counts.

Results CD20⁺ B cells decreased over time (p<0.0001). We observed a rapid reduction of naïve (CD19⁺CD20⁺IgD⁺IgM⁺CD27^-) and age-associated B cells (CD19⁺CD20⁺CD11c⁺CD21^-) already at the first follow-up visit (month 3), followed by a continuous decrease (p<0.0001 for both subsets), while double-negative memory B cells (CD19⁺CD20⁺IgD^-CD27^-) declined significantly first at month 6 (p=0.033) and pre-switching B cells (CD19⁺CD20⁺IgD⁺CD27⁺) showed a trend towards a decrease (p=0.052). Plasma cells (CD138⁺CD38⁺CD27⁺CD19⁺CD3e^-CD20^-) and switched memory B cells (CD19⁺CD20⁺CD27⁺IgD^-) remained stable during the study period, as did T cells and monocytes (p>0.2). Despite continuously decreasing SLE Disease Activity Index, immunological changes correlated with clinical improvements only during early time points (month 0–3). Interestingly, high baseline B cell counts were predictive of non-attaining Lupus Low Disease Activity State at month 24 (area under the ROC-curve: 0.95).

Conclusions B cell alterations betided in two distinct phases, a rapid early and a gradual late phase. Late clinical improvements might reflect preceding immunological changes, implying that early treatment evaluation and discontinuation might underestimate delayed improvements reflecting the late B cell changes.