In Chronic Lymphocytic Leukemia the JAK2/STAT3 Pathway Is Constitutively Activated and Its Inhibition Leads to CLL Cell Death Unaffected by the Protective Bone Marrow Microenvironment

Severin Severin,Facco Facco,Carraro Carraro,Semenzato Semenzato,Trentin Trentin,Trimarco Trimarco,Tibaldi Tibaldi,Piazza Piazza,Brunati Brunati,Frezzato Frezzato,Visentin Visentin,Martini Martini

doi:10.3390/cancers11121939

Severin Severin, Facco Facco + Show 10 more

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https://doi.org/10.3390/cancers11121939

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Abstract

The bone marrow microenvironment promotes proliferation and drug resistance in chronic lymphocytic leukemia (CLL). Although ibrutinib is active in CLL, it is rarely able to clear leukemic cells protected by bone marrow mesenchymal stromal cells (BMSCs) within the marrow niche. We investigated the modulation of JAK2/STAT3 pathway in CLL by BMSCs and its targeting with AG490 (JAK2 inhibitor) or Stattic (STAT3 inhibitor). B cells collected from controls and CLL patients, were treated with medium alone, ibrutinib, JAK/Signal Transducer and Activator of Transcription (STAT) inhibitors, or both drugs, in the presence of absence of BMSCs. JAK2/STAT3 axis was evaluated by western blotting, flow cytometry, and confocal microscopy. We demonstrated that STAT3 was phosphorylated in Tyr705 in the majority of CLL patients at basal condition, and increased following co-cultures with BMSCs or IL-6. Treatment with AG490, but not Stattic, caused STAT3 and Lyn dephosphorylation, through re-activation of SHP-1, and triggered CLL apoptosis even when leukemic cells were cultured on BMSC layers. Moreover, while BMSCs hamper ibrutinib activity, the combination of ibrutinib+JAK/STAT inhibitors increase ibrutinib-mediated leukemic cell death, bypassing the pro-survival stimuli derived from BMSCs. We herein provide evidence that JAK2/STAT3 signaling might play a key role in the regulation of CLL-BMSC interactions and its inhibition enhances ibrutinib, counteracting the bone marrow niche.

Highlights

Janus Family Kinases (JAKs)/Signal Transducer and Activator of Transcription (STAT) signaling is one of the most investigated and important pathways involved in tumorigenesis and cancer sustainment
No significant differences emerged for Jak2 mRNA, we found a higher amount of Stat3
By Western Blotting (WB), we evaluated STAT3 phosphorylation on Tyr705 in different conditions: (i) freshly isolated chronic lymphocytic leukemia (CLL) cells; (ii) CLL cells cultured in medium alone for 24 h; (iii) CLL cells cultured for 24 h on a layer of bone marrow mesenchymal stromal cells (BMSCs)

Summary

Introduction

JAK/Signal Transducer and Activator of Transcription (STAT) signaling is one of the most investigated and important pathways involved in tumorigenesis and cancer sustainment. Deregulation of JAK/STAT signaling pathways, leading to inappropriate activation of STATs, has been recognized in several hematological malignancies as a critical factor supporting survival, proliferation and metabolism of neoplastic cells [2,3,4,5]. The survival of the neoplastic clone is supported by intrinsic defects of CLL cells, such as the activation of the B-cell receptor (BCR), the overexpression and activation of the Src kinase Lyn [8,9], Bruton tyrosine kinase (BTK) [10] and Bcl-2 [11] proteins, and extrinsic microenvironmental factors derived from the bone marrow or lymph nodes [12].

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