IL-6R Inhibition Reduces Activation Of Different Peripheral Memory B Cell Subsets In RA

Zafar Mahmood¹, Khalid Muhammad¹, Marc Schmalzing², Petra Roll³, Kathrina Eckert¹, Thomas Dörner⁴ and Hans-Peter Tony⁵, ¹Rheumatology and Clinical Immunology, University of Würzburg, Würzburg, Germany, ²Rheumatology/Clinical Immunology, University Hospital Würzburg, Würzburg, Germany, ³Rheumatology and Clinical immunology, University of Würzburg, Würzburg, Germany, ⁴CC12, Dept. Medicine/Rheumatology and Clinical Immunology, Charité University Medicine Berlin, Berlin, Germany, ⁵Rheumatology/Clinical Immunology, University of Würzburg, Würzburg, Germany

Meeting: 2013 ACR/ARHP Annual Meeting

Keywords: B cells, IL-6, Rheumatoid arthritis (RA), states of activation and tocilizumab

Session Information

Title: B cells in Human and Animal Arthritis

Session Type: Abstract Submissions (ACR)

Background/Purpose: Enhanced B cell activity has been proposed as part of the pathogenesis of rheumatoid arthritis also based on the clinical experiences obtained by B cell targeted therapies able to modulate memory B cells. Human peripheral memory B cells can be distinguished by the phenotypic expression of CD27 and IgD defining three major B cell subpopulations: CD27+IgD+ pre-switch, CD27+IgD- post-switch and CD27-IgD- double negative memory B cells. We analyzed these different memory populations in RA and under IL6R blockade.

Methods: B cells from RA patients were phenotypically analyzed by 8 color flow cytometry at baseline, week 12 and week 24 under tocilizumab (TCZ) treatment. Memory B cell subsets were defined by CD27 and isotype surface expression. B cell activation was identified by surface staining with CD95 and intracellular Ki-67 staining. Mutation frequencies of VH gene rearrangements were analyzed by single B cell RT-PCR. Statistical analysis was performed using Mann Whitney U test.

Results: The Ig receptor mutational frequency was highest in class switched CD27+/IgD- memory B cells with 6.1±0.3% compared to CD27-/IgD- (3.5±0.2%) largely containing memory-like B cells and CD27-/IgD+ pre-switch memory B cells (4.1±0.2%). The phenotypically analyzed isotype profile in RA patients (n=40) and healthy donors (n=18) revealed that the memory B cell pool was a heterogeneous population of IgA, IgG and IgM expressing cells. The CD27-/IgD- B cell memory population showed a clear dominance of IgG followed by IgA and IgM (~70%, 20% and 10% respectively), whereas CD27+IgD- class switched B cells had an equal distribution of IgA and IgG. Under IL-6R inhibition by TCZ, the distribution of Ig isotypes remained stable at week 12 and 24. Surface and intracellular staining of B cells showed a significantly higher percentage of CD95 (14.0±1.4%, p=0.016) and Ki-67 expression (3.0±0.3%, p=0.041) in B cells of RA as compared to HD (CD95: 7.5±1.0% and Ki-67: 1.9±0.2% which was highest in post-switched memory B cells in RA (CD95: 38.4±2.1% and Ki-67: 8.1±1.0%). In post-switched memory B cells, IL-6R inhibition significantly decreased the expression of CD95 to 21.1±3.4 % and Ki-67 to 4.8±0.7% at week 12 with p=0.0004 and p=0.0081 respectively.

Conclusion: Our data suggest that the three major peripheral memory B cell populations, pre-, post-switch as well as CD27-/IgD- B cells harbour different numbers of mutations in their Ig receptors. These B cell subsets are activated in RA with enhanced CD95 and Ki-67 expression compared to healthy individuals and which can be reduced by IL-6R inhibition in vivo. The CD27-/IgD- B cell pool displays a significantly higher proportion of IgG bearing cells compared to post-switch B cells which is not modulated by IL-6R inhibition.

Disclosure:

Z. Mahmood,
None;

K. Muhammad,
None;

M. Schmalzing,
None;

P. Roll,
None;

K. Eckert,
None;

T. Dörner,
None;

H. P. Tony,
None.

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