In the loss of immune tolerance in patients with systemic lupus erythematosus (SLE), impaired cell death processes and type I interferon (IFN) signaling pathways play a central role. Data analysis from a phase 2 clinical trial showed that blocking type I IFN signaling with anifrolumab inhibited inflammatory proteins associated with disease activity, improved markers of cardiometabolic disease, and reversed SLE-associated lymphopenia, neutropenia, and thrombocytopenia. In this context, foreign scholars conducted transcriptomic and proteomic analysis of the patient populations in the clinical trials of TULIP-1 and TULIP-2 to further explore the immunomodulatory mechanism of blocking type I IFN receptor subunit 1 in the treatment of SLE with aniglucumab.
Study design
Both the TULIP-1 and TULIP-2 trials were multicenter, double-blind, randomized, placebo-controlled, 52-week phase 3 trials in adults with moderate to severe SLE who received standard of care. On top of standard treatment, patients were randomized to receive intravenous aniglulumab (TULIP-1 included two dose groups, 150 m and 300 mg; The dose group of TULIP-2 is 300 mg administered every 4 weeks for 48 weeks.
Whole genome RNA sequencing (RNA-seq) was used to analyze the expression of 18,017 genes in whole blood (summarized TULIP; anilulumab, n = 244, placebo group, n = 258). Analyze 184 plasma proteins (TULIP-1; Anirumumab, n = 124, placebo group, n = 132). This analysis used only data from the 300mg and placebo groups of anilulumab.
Findings:
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1. Anilulumab vs. placebo: Thousands of genes are differentially expressed, and type I IFN-inducible genes are one of them
Compared with placebo, at week 24, anilulumab was downregulated in 1681 genes (PFDR<0.05) and 1170 genes were upregulated (PFDR<0.05) compared with placebo; At week 52, 2092 genes were downregulated and 1645 genes were upregulated. Among the upregulated genes at weeks 24 and 52, 789 genes overlapped, and among the downregulated genes, 1420 genes overlapped. The type I IFN inducible gene is one of the genes that has been significantly downregulated.
In terms of pathway modulation, aniglulumab significantly downregulated or upregulated multiple pathways at week 52 compared with placebo, and the authors listed the first 25 pathways (expression up-regulation: NES > 0; expression down-regulation: NES < 0; PFDR ≤ 0.001), as shown in Figure 1. Among them, several intracellular signaling pathways downstream of IFN-α/-β29 were inhibited, including Janus kinase/signal transducer and activator of transcription, inosine phosphate 3-kinase and nuclear factor kappa light chain enhancer for activated B cells, and mitogen-activated protein kinase pathway. Multiple pathways associated with IFN-γ-29 are downregulated, including IFN-γ-mediated apoptosis induction and proliferation inhibition and IFN-γ acting on extracellular matrix and cell differentiation.
Figure 1 Pathway analysis of genes downregulated or upregulated by anirumumab at 52 weeks
Note: Pooled analysis from TULIP-1 and TULIP-2; FDR, False Discovery Rate; GSEA, gene set enrichment analysis; NES, standardized enrichment score; PFDR, FDR-corrected p-value
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2. Aniglucumab vs. placebo: 41 proteins were differentially expressed, and IFN-λ was one of them
At 52 weeks, aniglulumab regulated 41 proteins, 34 of which were downregulated (PFDR≤0.05), involving IFN-λ, IFN-γ-induced chemokines, chemokines that regulate and recruit monocytes, cytokines and chemokines with B cell and T cell regulatory functions, and T cell regulatory molecules; Seven proteins, including tumor necrosis factor (TNF)-related apoptosis-inducing ligand receptor 3, were significantly upregulated (Figure 2).
Fig.2 1-year protein level trajectories of patients in anilulumab 300mg group and placebo group (longitudinal linear mixed-effect model)
Note: Data are derived from protein expression in the TULIP-1 assay
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3. Aniglucumab vs. Placebo: Lymphocyte count increased and remained stable until 52 weeks
Compared to placebo, the modules that were significantly downregulated (PFDR≤0.01) at week 52 with aniglulumab included modules associated with IFN activity, and non-IFN pathways included antigen presentation, neutrophil, and monocyte modules. Some modules related to lymphocytes and T cells, as well as modules related to protein modification and synthesis, were significantly upregulated (Figure 3).
Fig.3 Gene set enrichment analysis of transcriptome module at 52 weeks
The estimated proportion of mucosa-associated constant T cells and γδ T cells increased in patients treated with aniglucumab, decreased the estimated proportion of monocytes from baseline to week 52, and remained stable in patients treated with placebo (Figure 4). In the anirulumab-treated group, the complete blood count continued to increase from baseline to week 8 and remained stable until week 52, while this phenomenon was not observed in the placebo group.
Fig.4 Prediction of cell type abundance (A) and lymphocyte count (B)
Note: MAIT, mucosa-associated constant T; NES, standardized enrichment score; PFDR, FDR-adjusted p-value.
Research Discussion
Through transcriptomics and proteomics, this study provides insight into the immunomodulatory mechanism by which anilulumab blocks type I IFN signaling. Blockade of type I IFNs with anirulumab modulates multiple inflammatory pathways, including apoptosis, innate and adaptive immune mechanisms that play a key role in the pathogenesis of SLE immunity. After type I IFN was blocked by anirumumab, apoptosis and neutrophil extracellular trap cell death pathways were inhibited. Aniglulumab can affect both innate and adaptive immunity, down-regulate multiple gene modules in plasma cells and monocytes, upregulate T cell and lymphocyte modules, and increase lymphocyte counts, suggesting that anirumumab's inhibition of type I IFN receptor subunit 1 may help reverse the imbalance of immune cell composition observed in SLE patients.
Conclusions of the study
Multiple pathways downstream of type I IFN receptor subunit 1 are key drivers of SLE pathogenesis, and inhibition of type I IFN signaling pathways can yield favorable clinical outcomes.
Bibliography:
Baker T, Sharifian H, Newcombe PJ, et al. Type I interferon blockade with anifrolumab in patients with systemic lupus erythematosus modulates key immunopathological pathways in a gene expression and proteomic analysis of two phase 3 trials. Ann Rheum Dis. 2024 May 3:ard-2023-225445. doi: 10.1136/ard-2023-225445.
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