Background/Purpose: Systemic lupus erythematosus (SLE) is an autoimmune disease with diverse clinical and immunological manifestations. Bruton’s tyrosine kinase (BTK) is a crucial intracellular kinase in regulating B cell development and function, and also showed pivotal impacts in numerous pathways. Small molecule inhibitors of BTK have been widely used in treating B-cell malignancies and autoimmune diseases (e.g., rheumatoid arthritis, atopic dermatitis, SLE). BTK degraders can eliminate the kinase scaffolding function and remove resistance mutations, displaying a therapeutic potential in SLE. However, the mechanism of action of BTK inhibitors/degraders in SLE is not well understood. The aim of this study is to explore the mechanism of action and identify biomarkers for BTK inhibitor/degrader by comparing cellular and protein-level changes in the mouse SLE model before and after treatment, and to predict its clinical efficacy and mechanism.

Methods: scRNA-seq and protein data from healthy donors and SLE patients, obtained from public databases, were used to narrow down the scope of biomarker research. Adoptive transfer of CD4+T cells from BM12 spleen and lymph node to C57BL/6 mice was performed to induce the lupus-like model. Normal, vehicle, Ibrutinib (BTK inhibitor), and NX-5948 (BTK degrader) were administered daily, and the mice were sacrificed around week 2. The plasma levels of anti-double-stranded DNA (anti-dsDNA) IgM and IgG were detected by ELISA. Cell subtypes, activation, proliferation and BTK expression in splenic cells were characterized by flow cytometry. The cytokines in the plasma were analyzed by Olink technology.

Results: The BM12-induced SLE mouse model generally shares a common set of features that correlate with clinical manifestations of human SLE. Compared to normal mice, the levels of anti-dsDNA IgG and IgM increased in the model group, with an expansion of Tfh, GCB, and plasma cells. Olink analysis results indicate elevated levels of cytokines and chemokines, including TNF, IFN-γ, IL-6, CCL4, and CSF1. Both Ibrutinib and NX-5948 attenuated the BM12-induced lupus-like symptoms, including the reduction of anti-dsDNA IgM and IgG. Compared to Ibrutinib treatment, the BTK protein level in B cell, dendritic cell, monocyte and macrophage were effectively decreased in NX-5948 treatment group. Both Ibrutinib and NX-5948 treatment decreased the number of GCB, plasma cell and the expression of CD69 and CD86 in B cells. The reduction of IL-4 and IL-6 in plasma, along with the downregulation of CD25 expression on B and T cells, suggests that the use of BTK inhibitors and degraders inhibit the interaction and activation of B and T cells.

Conclusion: This study conducted a detailed comparative analysis at the cellular and protein levels among normal mice, SLE model mice, and those treated with a BTK inhibitor or degrader. Our study compared clinical data with disease model data to determine their correlation and used this model to explore the mechanisms of action of BTK inhibitors and degraders. We hope that this research approach can be applied to the study of more diseases.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/exploring-the-mechanism-of-action-and-related-biomarkers-of-btk-inhibitor-degrader-in-systemic-lupus-erythematosus/