Immunomodulation Induced By The Minor Subpocket Of Cxcr4 In B Cells
GIORGIUTTI S. 1,2, ROTTURA J. 1, VOLLMER O. 1,2, SOULAS-SPRAUEL P. 1,2, HERBEUVAL J. 3, KORGANOW A. 1,2, GIES V. 1,2
1 Université de Strasbourg, Inserm UMR - S1109, Strasbourg, France; 2 Department of Clinical Immunology and Internal Medicine, National Reference Center for Systemic Autoimmune Diseases (CNR RESO), Tertiary Center for Primary Immunodeficiency, Strasbourg University Hospital , Strasbourg, France; 3 CNRS UMR-8601, Chemistry & Biology, Modeling & Immunology for Therapy, Paris, France
Objectives: A novel function of CXCR4 has recently been described through the modulation of pro-inflammatory cytokine production by Toll-like Receptor (TLR) activated innate immune cells. The binding of a specific region (minor pocket) of CXCR4 with small molecules, Minor Pocket Agonists (MiPAs), drives a profound anti-IFN-I signal. MIPAs injection in a pristane-induced lupus mouse model reduced disease progression and circulating anti-dsDNA antibodies, suggesting an effect on B cells. Here, we aim to describe the effect of CXCR4 minor pocket activation on B cells.
Methods: Human sorted B cells were used for in vitro experiments. B cell activation was evaluated after 24 hours of stimulation with TLR agonists or BCR stimulation, using flow cytometry and cytokine multiplex immunoassays. Plasmablast differentiation was evaluated in flow cytometry and with immunoglobulins quantification in the supernatants (ELISA). The effect of MIPAs on cell migration was also evaluated. CXCR4 knock-out and HA-CXCR4-labelled B cell lines were also generated for further mechanistic investigations.
Results: After 24 hours of TLR7 or TLR9 stimulation, B cell activation was strongly impaired in the MiPAs-treated group. Pro-inflammatory cytokines in supernatants were also barely detectable, suggesting an early inhibition of B cell activation in vitro. We found that B cell plasmablast differentiation was abolished in presence of MiPAs, resulting in a low production of immunoglobulins in the supernatants. MiPAs have however no effect on cell migration induced by CXCL12 chemotaxis in a Transwell migration assays demonstrating that MIPAs do not interfere with CXCL12 signaling, providing competitive advantage for potential MiPAs-based therapeutic strategies.
Conclusions: CXCR4 minor pocket activation by MiPAs induces a specific profound immunomodulation of B cells activated by TLR agonists. Understanding the underlying molecular mechanisms using generated transgenic B cell lines and transcriptomic approach is an ongoing challenge, in the hope of making them a new therapeutic target in autoimmune diseases.