Background: Treatment with autologous CD19 chimeric antigen receptor (CAR) T cells induces deep depletion of B cells in humans. This potential may allow resetting aberrant B-cell immunity in patients with autoimmune diseases such as systemic lupus erythematosus (SLE), idiopathic inflammatory myositis (IIM) and systemic sclerosis (SSc).

Objectives: To investigate the effects of CD19 CAR-T cell treatment on B cell immunity in patients with SLE, IIM and SSc.

Methods: Spectral analyzer-based characterization of B cell subsets in 8 SLE, 2 IIM and 2 SSc patients at baseline, early reconstitution (EaR) 4 months and established reconstitution (EsR) one year after CD19 CAR-T cell therapy. High-throughput single-cell mRNA sequencing of B cells with an Illumina NovaSeq platform in 5 SLE patients before and one year after CD19 CAR-T cell therapy. VH repertoire analysis of B cells based on VDJ library preparation and Illumina MiSeq sequencing in each one patient with SLE, IIM and SSc one year after CD19 CAR-T cell therapy and comparison with VH repertoire of two healthy controls.

Results: Characterization of B cell subsets revealed that reconstituted B cells at EaR and EsR showed a naïve phenotype, while CD19+CD27+ memory B cells were drastically reduced. A limited increase in CD19+CD27+ memory B cells occurred between EaR and EsR, which was predominantly seen in pre-switched IgD+ CD27+ B cells. CD27+CD38+ plasmablasts and SLE-associated activated CD11c+ memory B cells disappeared in SLE patients after B cell reconstitution. Immature CD38+ B cells, indicating reconstitution of B cells from the bone marrow boosted at EaR and declined thereafter. Similar results were also seen in at EaR an EsR of B cells in IIM and SSc. Single cell sequencing-based analysis of heavy chains in SLE patients showed virtual disappearance of IGHG1, IGHG2, IGHG3, IGHG4, IGHA1 and IGHA2 chains, while the expression of IGHM and IGHD increased, resembling a non-class switched B cell receptor phenotype. The expression of distinct chains associated with SLE and autoimmunity, such as immunoglobulin kappa variable (IGKV) 4-1, immunoglobulin heavy variable (IGHV) 4-59 chain and immunoglobulin lambda variable (IGLV) 3-21 chain was downregulated. In addition, a broad and balanced V _H – J _H usage was found in SLE, IIM and SSc patients treated with CAR-T cells without any noticeable clonal expansions, very similar to a normal B cell repertoire of naïve B cells from healthy controls. Analysis of the frequency of somatic mutations in the V _H genes revealed that all IgM sequences had a very low mutation frequency similar to a repertoire from sorted naïve cells from healthy controls.

Conclusion: Altogether the appearance of a naïve non-class-switched B cell system, the disappearance of circulating plasmablasts, the down regulation of specific disease- associated heavy and light chains and the broad and balanced V _H – J _H usage support the concept that CD19 CAR-T cell therapy may have induced a reset of pathologic B cell autoimmunity in patients with systemic autoimmune disease.

REFERENCES: [1] Mougiakakos D et al., CD19-Targeted CAR T Cells in Refractory Systemic Lupus Erythematosus. N Engl J Med 2021;385:567-569.

[2] Mackensen A. et al., Anti-CD19 CAR T cell therapy for refractory systemic lupus erythematosus; Nat Med 2022 Oct; 28(10):2124-2132.

Acknowledgements: NIL.

Disclosure of Interests: None declared.