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

Stefano Baldoni,Chiara Rompietti,Debora Cecchini,Carlo Sorrentino,Marta Di Nicola,Bianca Fabi,Raffaella Giancola,Valerio Guarente,Paolo Sportoletti,Emanuela Rosati,Arianna Stella,Daniele Sorcini,Erica Dorillo,Filomena De Falco,Beatrice Del Papa,Mauro Di Ianni,Elena Mondani,Francesco Maria Adamo,Francesco Guardalupi,Francesca Ulbar,Michele Marchioni,Lorenzo Moretti,Manuel Nogarotto,Maria Grazia Mameli,Estevão Carlos Silva Barcelos ,S Plebani

doi:10.3389/fonc.2021.668573

Abstract

NOTCH1 mutations and deregulated signal have been commonly found in chronic lymphocytic leukemia (CLL) patients. Whereas the impact of NOTCH1 mutations on clinical course of CLL has been widely studied, the prognostic role of NOTCH1 activation in CLL remains to be defined. Here, we analyzed the activation of NOTCH1/NOTCH2 (ICN1/ICN2) and the expression of JAGGED1 (JAG1) in 163 CLL patients and evaluated their impact on TTFT (Time To First Treatment) and OS (Overall Survival). NOTCH1 activation (ICN1+) was found in 120/163 (73.6%) patients. Among them, 63 (52.5%) were NOTCH1 mutated (ICN1+/mutated) and 57 (47.5%) were NOTCH1 wild type (ICN1+/WT). ICN1+ patients had a significant reduction of TTFT compared to ICN1-negative (ICN1−). In the absence of NOTCH1 mutations, we found that the ICN1+/WT group had a significantly reduced TTFT compared to ICN1− patients. The analysis of IGHV mutational status showed that the distribution of the mutated/unmutated IGHV pattern was similar in ICN1+/WT and ICN1− patients. Additionally, TTFT was significantly reduced in ICN1+/ICN2+ and ICN1+/JAG1+ patients compared to ICN1−/ICN2− and ICN1−/JAG1− groups. Our data revealed for the first time that NOTCH1 activation is a negative prognosticator in CLL and is not correlated to NOTCH1 and IGHV mutational status. Activation of NOTCH2 and JAGGED1 expression might also influence clinical outcomes in this group, indicating the need for further dedicated studies. The evaluation of different NOTCH network components might represent a new approach to refine CLL risk stratification.

Highlights

NOTCH1 mutations and deregulated signal have been commonly found in chronic lymphocytic leukemia (CLL) patients (1, 2)
Western blot (WB) analysis was performed on protein lysates (20 μg) extracted from CLL cells freshly isolated from peripheral blood samples collected at diagnosis, using the Cleaved NOTCH1 Val1744 monoclonal antibodies (moAbs) (Cell Signaling Technology, Beverly, MA, USA) to detect activated NOTCH1 intracellular domain (ICN1+), the polyclonal NOTCH2 antibody Cleaved-Val1697
To exclude technical issues causing false intracellular domain of NOTCH1 (ICN1)−, in each WB session, we evaluated the quality of ICN1 staining, loading Molt[4] cell line lysate as positive control for delCT mutation, or Jurkat cell line lysate as positive control for NOTCH1 activation

Summary

INTRODUCTION

NOTCH1 mutations and deregulated signal have been commonly found in chronic lymphocytic leukemia (CLL) patients (1, 2). Clonal NOTCH1 mutations have been detected in up to 20% of CLL while recent evidence showed that the NOTCH1 pathway can be constitutively activated independently of mutation in about 50% of patients (3, 4). Whereas the impact of NOTCH1 mutations on the clinical course of CLL has been widely studied (5, 6), the role of NOTCH1 activation remains to be defined. We previously showed that CLL cells exhibit a constitutively activated NOTCH2 and express the JAGGED1 ligand which is involved in IL4-induced CLL cell survival (2, 7–9) whose prognostic role in CLL is largely unknown. In the present retrospective study, we analyzed the role of NOTCH1/NOTCH2 activation and JAGGED1 expression in the outcome of CLL patients and weighed up their impact in comparison with NOTCH1 and IGHV mutational status

MATERIALS AND METHODS

RESULTS AND DISCUSSION

ETHICS STATEMENT