Expression and Activity of the NF-κB Subunits in Chronic Lymphocytic Leukaemia: A Role for RelB and Non-Canonical Signalling

SIMPLE SUMMARY: Chronic lymphocytic leukaemia (CLL) patients frequently experience drug-resistance. Signalling via the transcription factor NF-κB is a major contributor to resistance due to its ability to regulate the expression of genes that confer growth and survival of CLL cells. NF-κB signalling comprises ‘canonical’ and ‘non-canonical’ pathways involving the subunits RelA and RelB, respectively. RelA has been widely investigated in resistance and outcome in CLL but RelB activity and therapeutic implications are less well understood. We sought to examine RelB expression and function using a large cohort of comprehensively annotated patient-derived CLL cells. We demonstrate that RelB activity is associated with specific subsets of patients and with poorer outcome. Use of CD40L-expressing cells to represent the in vivo microenvironment led to RelB activation and CLL cell proliferation. Strategies to develop novel agents that target non-canonical NF-kB signalling will enable patient stratification and the development of more personalized therapy for CLL patients. ABSTRACT: Background: Canonical NF-κB signalling by p65 (RelA) confers chemo-resistance and poor survival in chronic lymphocytic leukaemia (CLL). The role of non-canonical NF-κB signalling (leading to RelB and p52 subunit activation) in CLL is less understood, but given its importance in other B-cell tumour types, we theorised that RelB and p52 may also contribute to the pathology of CLL. Methods: DNA binding activity of all five NF-kB subunits, p65, p50, RelB, p52, and c-Rel, was quantified using ELISA and correlated to ex vivo chemoresistance, CD40L-stimulated signalling (to mimic the lymph node microenvironment), and clinical data. Results: Importantly, we show for the first time that high basal levels of RelB DNA binding correlate with nuclear RelB protein expression and are associated with del(11q), ATM dysfunction, unmutated IGHV genes, and shorter survival. High levels of nuclear p65 are prevalent in del(17p) cases (including treatment-naïve patients) and also correlate with the outcome. CD40L-stimulation resulted in rapid RelB activation, phosphorylation and processing of p100, and subsequent CLL cell proliferation. Conclusions: These data highlight a role for RelB in driving CLL cell tumour growth in a subset of patients and therefore strategies designed to inhibit non-canonical NF-κB signalling represent a novel approach that will have therapeutic benefit in CLL.