Abstract Background: Aplastic anemia (AA) is a bone marrow failure disorder caused by diverse etiologies. AA pathogenesis involves aberrant immune activation and an imbalanced inflammatory bone marrow microenvironment. Relapse following discontinuation of standard immunosuppressive therapy (IST) remains a significant challenge, indicating a dependency on continuous intervention and highlighting the need for deeper mechanistic understanding and improved treatments.

Methods: Based on the therapeutic effects observed in patients with SAA reated with Luspatercept and findings from single-cell sequencing, this study identified an association between the drug and monocyte pyroptosis levels. Subsequently, reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blotting (WB) were employed to detect pyroptosis expression level of THP1 cells and the changes after drug treatment. RNA sequencing was used to identify differentially expressed genes and related pathways. The functional changes of CD8+ T cells and CD14 monocytes were detected by flow cytometry. CD8+ T cells were further co-cultured with monocytes to assess their functional activity and cytokine secretion profile.

Results: The triple therapy (luspatercept + cyclosporine + eltrombopag) significantly restored hematopoietic function in patients with SAA. However, relapse occurred after discontinuation. ScRNA-seq revealed elevated pyroptosis-related genes in monocytes and increased inflammatory cytokines during relapse. Mechanistically, patients with SAA monocytes exhibited heightened NLRP3 inflammasome-dependent inflammatory responses and pyroptosis versus controls. Luspatercept specifically suppressed monocyte pyroptosis by inhibiting the NF-κB pathway, which is reshaped the immune microenvironment, attenuating pro-inflammatory cytokine secretion and cytotoxic molecule expression in CD8+ T cells of patients with AA.

Conclusions: This study demonstrates that luspatercept achieves multi-target immunomodulation in patients with SAA by inhibiting the NF-κB/NLRP3 crosstalk in monocytes. This action mitigates inflammation and pyroptosis, disrupting the pathogenic cycle and promoting hematopoietic recovery.

Keywords: Aplastic Anemia, Luspatercept, Pyroptosis, Monocytes, Immunization therapy

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2025
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