Abstract
Background/Objectives: Chronic lymphocytic leukemia (CLL) is the most common adult leukemia in Western countries and it can progress to Richter's syndrome (RS), a more aggressive condition. The NF-κB pathway is pivotal in CLL pathogenesis, driven mainly by B-cell receptor (BCR) signaling. However, recent evidence indicates that BCR signaling is reduced in RS, raising questions about whether and how NF-κB activity is maintained in RS. This study aims to elucidate the triggers and dynamics of NF-κB activation and the progression from CLL to RS. Methods: Integrated single-cell RNA sequencing data from peripheral blood samples of four CLL-RS patients were analyzed. NF-κB pathway activity and gene expression profiles were assessed to determine changes in NF-κB components and their targets. Tumor microenvironment composition and cell-cell communication patterns were inferred to explore NF-κB regulatory mechanisms. Results: RS samples showed increased proportions of malignant cells expressing NF-κB components, including NFKB1, NFKB2, RELA, IKBKG, MAP3K14, CHUK, and IKBKB, with significantly higher expression levels than in CLL. Enhanced NF-κB pathway activity in RS cells was associated with targets involved in immune modulation. The tumor microenvironment in RS displayed significant compositional changes, and signaling inference revealed enhanced cell-cell communication via BAFF and APRIL pathways, involving interactions with receptors such as BAFF-R and TACI on RS cells. Conclusions: The findings from this study reveal an active state of NF-κB in RS and suggest that this state plays a critical role in the evolution of CLL to RS, which is modulated by alternative signaling pathways and the influence of the tumor microenvironment.