Journal
ARTHRITIS AND RHEUMATISM
Volume 65, Issue 3, Pages 770-779Publisher
WILEY-BLACKWELL
DOI: 10.1002/art.37818
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Funding
- DFG [650/TP16, 491/7-1]
- UCB Pharma
- UCB
- Immunomedics
- Roche
- Sanofi-Aventis
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Objective CD22 is a surface molecule exclusively expressed on B cells that regulates adhesion and B cell receptor (BCR) signaling as an inhibitory coreceptor of the BCR. Central downstream signaling molecules that are activated upon BCR engagement include spleen tyrosine kinase (Syk) and, subsequently, phospholipase C2 (PLC2), which results in calcium (Ca2+) mobilization. The humanized anti-CD22 monoclonal antibody epratuzumab is currently being tested in clinical trials. This study was undertaken to determine the potential mechanism by which this drug regulates B cell activation. Methods Purified B cells were preincubated with epratuzumab, and the colocalization of CD22 and CD79, without BCR engagement, was assessed by confocal microscopy. The phosphorylation of Syk (Y348, Y352) and PLC2 (Y759) as well as the Ca2+ flux in the cells were analyzed by flow cytometry upon stimulation of the BCR and/or Toll-like receptor 9 (TLR-9). The influence of CD22 ligation on BCR signaling was assessed by pretreating the cells with epratuzumab or F(ab)2 fragment of epratuzumab, in comparison with control cells (medium alone or isotype-matched IgG1). Results Epratuzumab induced colocalization of CD22 and components of the BCR independent of BCR engagement, and also reduced intracellular Ca2+ mobilization and diminished the phosphorylation of Syk and PLC2 after BCR stimulation in vitro. Inhibition of kinase phosphorylation was demonstrated in both CD27 and CD27+ B cells, and this appeared to be independent of Fc receptor signaling. Preactivation of the cells via the stimulation of TLR-9 did not circumvent the inhibitory effect of epratuzumab on BCR signaling. Conclusion These findings are consistent with the concept of targeting CD22 to raise the threshold of BCR activation, which could offer therapeutic benefit in patients with autoimmune diseases.
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