Decreased NOTCH1 Activation Correlates with Response to Ibrutinib in Chronic Lymphocytic Leukemia

Abstract

Purpose: Ibrutinib, a Bruton tyrosine kinase inhibitor (BTKi), has improved the outcomes of chronic lymphocytic leukemia (CLL), but primary resistance or relapse are issues of increasing significance. While the predominant mechanism of action of BTKi is the B-cell receptor (BCR) blockade, many offtarget effects are unknown. We investigated potential interactions between BCR pathway and NOTCH1 activity in ibrutinib-treated CLL to identify new mechanisms of therapy resistance and markers to monitor disease response. Experimental Design: NOTCH activations was evaluated either in vitro and ex vivo in CLL samples after ibrutinib treatment by Western blotting. Confocal proximity ligation assay (PLA) experiments and analyses of down-targets of NOTCH1 by qRT-PCR were used to investigate the crosstalk between BTK and NOTCH1. Results: In vitro ibrutinib treatment of CLL significantly reduced activated NOTCH1/2 and induced dephosphory-lation of eIF4E, a NOTCH target in CLL. BCR stimulation increased the expression of activated NOTCH1 that accumulated in the nucleus leading to HES1, DTX1, and c-MYC transcription. Results of in situ PLA experiments revealed the presence of NOTCH1-ICD/BTK complexes, whose number was reduced after ibrutinib treatment. In ibrutinib-treated CLL patients, leukemic cells showed NOTCH1 activity downregulation that deepened over time. The NOTCH1 signaling was restored at relapse and remained activated in ibrutinib-resistant CLL cells. Conclusions: We demonstrated a strong clinical activity of ibrutinib in a real-life context. The ibrutinib clinical efficacy was associated with NOTCH1 activity downregulation that deepened over time. Our data point to NOTCH1 as a new molecular partner in BCR signaling with potential to further improve CLL-targeted treatments.

High-grade heart block associated with ibrutinib therapy

“…Ibrutinib treatment has been shown to reduce NOTCH1 activation, most pronounced at 12 months of continuous therapy. 13 In the setting of cardiac ischemia, NOTCH1 -activated bone marrow–derived mesenchymal stem cells promote CM survival through an antifibrotic mechanism. 14 Although this hypothesis is predicated on myocardial injury, subclinical ischemia is possible in a cohort with a median age over 75.…”

NOTCH1 Activation Negatively Impacts on Chronic Lymphocytic Leukemia Outcome and Is Not Correlated to the NOTCH1 and IGHV Mutational Status

“…The aberrant NOTCH1 signaling in CLL cells regulates several genes that influence key biological aspects of neoplastic cells, such as apoptosis, cell growth, cell migration, interactions with the microenvironment and B Cell Receptor (BCR) activation. It has been shown that a synergistic cooperation between NOTCH1 and BCR strongly supports the survival/proliferation of CLL cells and contributes to a progression of the disease ( 11 ) but also to the transformation into Richter’s syndrome (RS) ( 12 ). Furthermore, it has been recently shown that ligand-dependent activation of NOTCH1 signaling, via constitutive PI3K/AKT activation, promotes CLL transformation towards RS in Eμ- TCL1 mice in vivo ( 13 ), thus further supporting the association between non-mutational NOTCH1 activation and a poor prognosis.…”

Targeting Nuclear NOTCH2 by Gliotoxin Recovers a Tumor-Suppressor NOTCH3 Activity in CLL

“…RO4929097 inhibited the NOTCH3/NR4A1 axis, upregulated the NOTCH2/FCER2 (CD23) axis, and enhanced the NOTCH2/CSL transcription factor complex ( Figure 5 B, right panel), which stands in sharp contrast to the effect of gliotoxin. The NOTCH1 target gene MYC was downregulated by RO4929097 ( Figure 5 B, right panel) [ 9 , 43 , 44 , 45 ].…”

“…Therefore, compounds that interfere with B-cell activation might affect the expression of both NOTCH receptors in CLL cells. This would explain why CLL cells pre-treated with Duvelisib (a dual PI3K-δ/γ inhibitor) [69] or Ibrutinib (a Bruton's tyrosine kinase inhibitor) [2,44] express lower amounts of CD23 and NOTCH3 and are less sensitive to apoptosis.…”

“…NOTCH1 is not detectable in nuclear NOTCH/CSL transcription factor complexes in CLL cells [ 7 , 8 , 18 , 32 ]. However, NOTCH1 is frequently mutated and/or overexpressed in advanced stage CLL cells, where it has a CLL-driving role by regulating MYC expression [ 8 , 9 , 16 , 17 , 43 , 44 , 45 ]. In this context, NOTCH1 may indirectly account for the relative GSI sensitivity of NOTCH2, keeping in mind that active NOTCH1 is a positive regulator of the NOTCH2 gene in CLL cells ( Figure 5 G) [ 12 ].…”