POS0798 B CELL RECONSTITUTION IN SLE PATIENTS AFTER CAR-T CELL THERAPY IS MARKED BY A NAÏVE PHENOTYPE AND REDUCTION OF EXTRAFOLLICULAR IMMUNE RESPONSES

Background: Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by aberrant B cell activation and autoantibody production. CD19 targeted chimeric antigen receptor (CAR) T-cell therapy has shown promising clinical results in the treatment of refractory SLE, in spite of leading to a rather short and transient, but deep depletion of B cells and allowing for B cell reconstitution after treatment. However, it is poorly understood how the new reconstituting B cell system evolve over time after B cell recovery and whether pathogenic immune phenotypes reemerge. Objectives: To identify and characterize the phenotype of B and T cell subsets in SLE patients before and after treatment with CD19 CAR T-cells, including observation of B cell reconstitution at longer term follow-up. Methods: 18 patients with active, refractory SLE - 15 of them with kidney involvement - received autologous CD19-CAR T-cell therapy at our institution, all further immunosuppressive medication was discontinued. The patients were followed longitudinally, median follow up was 17 months (range: 3 – 36 months). B cells already reconstituted in 14/18 (77,77%) of patients, the first B cell analysis after B cell reappearance was between 28 to 420 days post CAR-T (mean: 206d). Longitudinal, high dimensional B cell phenotyping was performed using spectral flow cytometry from peripheral blood. B cells were analyzed before therapy (N = 18), at first B cell reappearance (N = 14), and then at an early follow-up time point (N = 10) and at a late time point with a minimum of 12 months after cell reconstitution (N = 10). Results were compared to B cells from healthy controls (N = 10). Additionally, phenotyping of peripheral T cells was performed before therapy, during B cell depletion and at early and late follow-up (N=4). Results: Prior to therapy, the B cell compartment of SLE patients showed altered B cell subset frequencies, including a pathognomonic increase in circulating plasmablasts and heightened frequencies of IgD − CD27 − double negative (DN) memory cells. Transient B cell depletion of all CD19+ peripheral B cells was achieved in all patients post therapy. 14 out of 18 patients achieved B cell reconstitution and the absolute B cell count normalized over the course of the follow-up, remaining within normal frequencies comparable to that of healthy donors. At reconstitution and 6 months thereafter, the new B cell compartment after CAR T-cell therapy consisted predominantly of CD19 + CD20 + IgD + CD27 − naïve B-cells. A transient increase in immature and transitional B cells was observed during the early phase of reconstitution. Accordingly, all other B-cell subsets, including switched and unswitched memory and circulating plasmablasts were reduced. Frequencies of IgA- or IgG-switched B-cells were diminished and did not repopulate promptly. Quantification of Tbet + age associated B cells (ABCs) as well as activated naïve B cells and DN type 2 B cells showed increased frequencies before therapy and thorough depletion following CAR-T cell administration. Surprisingly, DN B-cells reappeared one year after CAR-T cell therapy. However, the fresh DN-cells consisted almost exclusively of CXCR5 + CD11c − Tbet − DN1 cells, whereas disease associated CD11c + CXCR5 − Tbet + DN2 cells stayed reduced, constituting less than 5% of peripheral CD19+ cells. Concordantly, new switched memory B cells were mostly CXCR5 + , while CXCR5 − switched memory B cells did not accumulate. New plasmablasts displayed substantially reduced activation levels (CD95 low HLA-DR low ). Within the T cell compartment, elevated frequencies of RORγt+ Th17 cells and exhausted Foxp3+ PD-1+ T cells were found before therapy; those numbers decreased to levels comparable to healthy controls after CAR T-celladministration. Conclusion: The new B-cell compartment after CAR T-cell therapy displays an overall naïve phenotype and reduced activation, indicative of a deep B-cell depletion and repopulation with naïve cells. Disease-associated plasmablasts and extrafollicular DN2 cells remain cleared long after B-cell reconstitution. Accordingly, B cells appear to predominantly repopulate from CXCR5 + cells, suggesting less utilization of extrafollicular B cell maturation pathways. In addition, reduced numbers of Th17 and exhausted T cells suggest less extrafollicular interplay between the two cell types. Taken together, the B- and T-cell compartment after CAR T-cell therapy differs substantially from the aberrant B-cell activation found in active SLE and resembles that of healthy controls, indicating a deep and durable reset of adaptive immunity. REFERENCES: [1] Mougiakakos, D., et al., CD19-Targeted CAR T Cells in Refractory Systemic Lupus Erythematosus. New England Journal of Medicine, 2021. 385 (6): p. 567-569. [2] Mackensen, A., et al., Anti-CD19 CAR T cell therapy for refractory systemic lupus erythematosus. Nature Medicine, 2022. 28 (10): p. 2124-2132. [3] Müller, F., et al., CD19 CAR T-Cell Therapy in Autoimmune Disease — A Case Series with Follow-up. New England Journal of Medicine, 2024. 390 (8): p. 687-700. Acknowledgements: NIL . Disclosure of Interests: None declared . © The Authors 2025. This abstract is an open access article published in Annals of Rheumatic Diseases under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Neither EULAR nor the publisher make any representation as to the accuracy of the content. The authors are solely responsible for the content in their abstract including accuracy of the facts, statements, results, conclusion, citing resources etc.