Enhanced Tyrosine Phosphatase Activity Underlies Dysregulated B Cell Receptor Signaling and Promotes Survival of Human Lupus B Cells.

OBJECTIVE: Systemic lupus erythematosus (SLE) is associated with hyperactivity of B cells and abnormalities of B cell receptor (BCR) signaling. To address the linkage between dysregulated BCR signaling and increased B cell function, we assessed immediate phosphorylation events in lupus B cells. METHODS: B cells from SLE patients and healthy donors were analyzed by flow cytometry to assess phosphorylated CD22, Syk, and Akt as well as the basal expression of the BCR coreceptors CD22 and CD19. Confocal microscopy studies determined the recruitment of CD22 and the tyrosine phosphatase SH2 domain-containing phosphatase 1 to the activated BCR complex. Additionally, phosphatase activity in SLE versus healthy donor B cells was measured. RESULTS: B cells from SLE patients showed diminished Syk phosphorylation and reduced intracellular calcium release after BCR activation as compared to B cells from healthy donors. This was related to an enhanced activity of tyrosine, but not serine/threonine, phosphatases and was corrected by inhibition of tyrosine phosphatase activity. In contrast to reduced Syk phosphorylation after BCR activation, phosphorylation of Akt was significantly increased in SLE B cells. The disturbed balance between Syk and Akt phosphorylation was significantly correlated with B cell survival following BCR engagement. Furthermore, CD27-, but not CD27+, B cells from SLE patients displayed increased expression and phosphorylation of the inhibitory BCR coreceptor CD22. CONCLUSION: These results indicate that an imbalance between serine and tyrosine phosphatases in SLE contributes to an intrinsically disturbed balance of BCR-initiated signaling pathways, resulting in enhanced survival of lupus B cells and differentiation into plasma cells.

Epratuzumab inhibits the production of the proinflammatory cytokines IL-6 and TNF-α, but not the regulatory cytokine IL-10, by B cells from healthy donors and SLE patients.

INTRODUCTION: Cytokines produced by B cells are believed to play important roles in autoimmune diseases. CD22 targeting by epratuzumab has been demonstrated to inhibit phosphorylation of B cell receptor (BCR) downstream signaling in B cells. It has been shown that other sialoadhesin molecules related to CD22 have immunoregulatory functions; therefore, in the present study, we addressed the role of epratuzumab on the production of key cytokines by B cells of patients with systemic lupus erythematosus (SLE) and of healthy donors (HD). METHODS: Peripheral blood B cells were purified and activated by BCR with or without Toll-like receptor 9 (TLR9) stimulation in the presence or absence of epratuzumab. Cytokine production by B cells (interleukin [IL]-6, tumor necrosis factor [TNF]-α and IL-10) in the supernatant and the induction of IL-10(+) B cells from patients with SLE and HD were analyzed. RESULTS: The secretion of the proinflammatory cytokines TNF-α and IL-6 by anti-BCR and BCR- and/or TLR9-activated B cells from HD and patients with SLE was inhibited by epratuzumab. In contrast, the production of IL-10 by B cells was not affected by epratuzumab under either stimulation condition. Consistently, the induction of IL-10-producing B cells in culture was not affected by epratuzumab. CONCLUSIONS: Epratuzumab, by targeting CD22, was able to inhibit the production of the proinflammatory cytokines IL-6 and TNF-α by B cells, in contrast to IL-10, in vitro. These data suggest that targeting CD22 alters the balance between proinflammatory cytokines (TNF-α, IL-6) and the regulatory cytokine IL-10 as another B cell effector mechanism.