0754: Persistent but Disturbed Germinal Center Reaction Among 3^rd SARS-CoV-2 Vaccination After Rituximab Exposure

Ana-Luisa Stefanski¹, Hector Rincon-Arevalo¹, Eva Schrezenmeier², Franziska Szelinski¹, Jacob Ritter³, Yidan Chen¹, Christian Meisel⁴, Hubert Schrezenmeier⁵, Andreia C. Lino⁶ and Thomas Dörner¹, ¹Department of Rheumatology and Clinical Immunology, Charité Universitätsmedizin Berlin, Berlin, Germany, ²Department of Nephrology and Medical Intensive Care, Charité Universitätsmedizin Berlin; DRFZ Berlin; Berlin Institute of Health Charité Universitätsmedizin Berlin, Berlin, Germany, ³Department of Rheumatology and Clinical Immunology, Berlin Institute of Health Charité Universitätsmedizin Berlin, Berlin, Germany, ⁴Department of Medical Immunology, Charité University Medicine and Labor Berlin-Charité Vivantes, Berlin, Germany, ⁵Institute of Transfusion Medicine, Ulm University, Ulm, Germany and Institute for Clinical Transfusion Medicine and Immunogenetics, German Red Cross Blood Transfusion Service Baden-Württemberg – Hessen and University Hospital Ulm, Ulm, Germany, ⁶Deutsches Rheuma-Forschungszentrum Berlin, Berlin, Germany

Background/Purpose: Durable vaccine-mediated immunity relies on the generation of long-lived plasma cells and memory B cells (MBCs), differentiating upon germinal center (GC) reactions. SARS-CoV-2 mRNA vaccination induces a strong GC response in healthy volunteers (HC), but limited data is available in patients with medically induced B cell impairment upon rituximab treatment.

Methods: We evaluated humoral as well as B and T cellular responses upon 3rd vaccination in seven patients with rheumatoid arthritis (RA) who initially mounted anti-spike SARS-CoV-2 IgG antibodies after primary 2x vaccination and got re-exposed to RTX 1-2 months after the second vaccination. Ten patients with RA on other therapies and ten HC represented the control groups. As control for known long-lived induced immunity, we analyzed humoral and cellular tetanus toxoid (TT) immune responses in steady-state. To assess subpopulations of circulating antigen-specific, receptor-binding domain (RBD+) B cells before and after 3^rd vaccination (and as control TT+ B cells at steady state), we implemented a high-dimensional flow cytometry analysis using Uniform Manifold Approximation and Projection (UMAP). Antigen-specific T cells were assessed after 2nd and 3rd vaccination.

Results: After 3^rd vaccination, 5/7 RTX patients were IgG seroconverted, revealing lower anti-SARS-CoV-2 IgG titers but similar neutralizing capacity compared with HC. The fine-tuning of high-affinity antibodies as a direct result of somatic hypermutation seems functional also in RTX treated patients. Antibody levels after 2^nd vaccination correlated with titers after 3^rd vaccination, suggesting that upon RTX re-exposure, induction strength of the immune response within recall vaccination, is largely related to pre-existing memory formation. Despite significant reduction of circulating total and antigen-specific B cells in RTX re-exposed patients, we observed the induction of IgG+ MBCs upon 3^rd vaccination (Fig.1 A, B). Notably, only RTX treated patients revealed a high amount of IgA+ memory B cells before 3^rd vaccination with induction of IgA+ plasmablasts (PB) upon boost, suggesting a persistent but disturbed germinal center activity. In contrast, IgA+ B cells were not part of the steady state TT+ B cell pool. With respect to antigen-specific T cells, TNF-secretion and generation of effector memory CD4 spike-specific T cells were significantly boosted upon 3^rd vaccination.

Conclusion: Herein, we describe for the first time humoral and cellular responses upon 3^rd SARS-CoV-2 injection in patients re-exposed to RTX after initially seroconversion upon primary vaccination. While functional aspects of spike-specific CD4 T cells are boosted upon 3^rd vaccination, we report a persistent but disturbed germinal center activity, possibly supported by (semi-primary and /or) additional extrafollicular responses in patients re-exposed to RTX as potential compensatory mechanisms employed in such medically induced B cell impairment.
Distinct B cell subsets before and after 3rd SARS-CoV-2 vaccination. UMAP clustering was performed on a concatenated file of pre-gated antigen specific (RBD+ together with TT+) CD19+ B cells composed of total 1579 events. (A) Cluster overlay of B cells of all groups for subset identification. (B) Distribution of clusters before and after 3rd vaccination, as well as for TT+ B cells in HD, RA and RTX.
Disclosures: A. Stefanski, None; H. Rincon-Arevalo, None; E. Schrezenmeier, None; F. Szelinski, None; J. Ritter, None; Y. Chen, None; C. Meisel, None; H. Schrezenmeier, None; A. Lino, None; T. Dörner, AbbVie, Eli Lilly, Roche/Genentech, Janssen, Novartis, Bristol-Myers Squibb(BMS), UCB.