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Reticulin fibrosis and nestin positive (nestin + ) cells before and after treatment with mirabegron. (A) Bone marrow histology of a patient before (week 0) and at the end (week 24) of treatment with mirabegron. (Top) Reticulin fibers are stained black by silver impregnation (Gömöri). (Bottom) Immunohistochemical staining with a monoclonal antibody against human nestin protein. Note decrease of reticulin fibrosis and increase of nestin + cells (brown staining) after 24 weeks of treatment. Magnification: 200x. (B) Single patient evolutional curves of the grade of reticulin fibrosis (left) and nestin + mesenchymal cells/mm 2 (right) at study inclusion and after 24 weeks of mirabegron. n: number; PMF: primary myelofibrosis; ET: essential thrombocythemia; PV: polycythemia vera; MF: myelofibrosis.
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The β-3 sympathomimetic agonist BRL37344 restored nestin-positive cells within the stem cell niche, and thereby normalized blood counts and improved myelofibrosis in a mouse model of JAK2-V617F-positive myeloproliferative neoplasms. We therefore tested the effectiveness of mirabegron, a β-3 sympathomimetic agonist, in a phase II trial including 39...
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Context 1
... slight decrease in reticulin fiber content from a median grade of 1.0 (IQR 0-3) to 0.5 (IQR 0-2) (P=0.01) and an increase in the nestin + MSCs cells from a median of 1.09/mm 2 (IQR 0.38-3.27) to 3.95/mm 2 (IQR 1.98-8.79) (P<0.0001) were observed (Figure 2). The mean change in the nestin + cells from baseline to week 24 was 3.52/mm 2 [95% confidence interval (CI): 1.65-5.39]. ...
Context 2
... some of the effects observed in the preclinical JAK2-V617F mouse model treated with BRL37344, i.e. increase in nestin + bone marrow MSCs and decrease in myelofibrosis, were also seen in our mirabegron study: BM biopsies performed in a subset of 20 patients revealed a significant increase in the nestin + MSCs and a decrease in reticulin fibrosis (Figure 2). Although the beneficial effect of mirabegron on reticulin fibrosis was moderate, the duration of treatment was also rather short (24 weeks), as it was mainly designed to assess the primary end point of reduction of allele burden. ...
Citations
... On the other hand, risk factors for MF in the context of ET (post-ET MF) are advanced age, leukocytosis, anemia, reticulin fibrosis, absence of JAK2 V617F , use of anagrelide, and the presence of ASXL1 mutation. Growth factors, cytokines, and chemokines, such as CXCL4, CXCL10, FGF, PDGFA, TGF-β, VEGF, as well as loss of β3 sympathetic innervation and nestin niches have also been found to play an important role in progressive MF [59,[61][62][63][64]. The so-called "megakaryocytic activation" (M-ACT), characterized by megakaryocytic emperipolesis, clustering, and fibers surrounding megakaryocytes, has been suggested to predict the development of MF in MPN. ...
... With respect to mechanisms of MF in MPN, proinflammatory monocytes overproduce growth-regulated oncogene α (CXCL1), and its levels predict respective progression in ET [62]. Of some practical importance, clinical trial data suggest potential applicability of the β3 sympathomimetic agonist mirabegron to restore nestin-positive niches and reduce reticulin BM fibrosis in PMF and PV [64]. ...
Disease progression in myelodysplastic syndromes (MDS), myelodysplastic-myeloproliferative neoplasms (MDS/MPN), and myeloproliferative neoplasms (MPN), altogether referred to as myeloid neoplasms (MN), is a major source of mortality. Apart from transformation to acute myeloid leukemia, the clinical progression of MN is mostly due to the overgrowth of pre-existing hematopoiesis by the MN without an additional transforming event. Still, MN may evolve along other recurrent yet less well-known scenarios: (1) acquisition of MPN features in MDS or (2) MDS features in MPN, (3) progressive myelofibrosis (MF), (4) acquisition of chronic myelomonocytic leukemia (CMML)-like characteristics in MPN or MDS, (5) development of myeloid sarcoma (MS), (6) lymphoblastic (LB) transformation, (7) histiocytic/dendritic outgrowths. These MN-transformation types exhibit a propensity for extramedullary sites (e.g., skin, lymph nodes, liver), highlighting the importance of lesional biopsies in diagnosis. Gain of distinct mutations/mutational patterns seems to be causative or at least accompanying several of the above-mentioned scenarios. MDS developing MPN features often acquire MPN driver mutations (usually JAK2), and MF. Conversely, MPN gaining MDS features develop, e.g., ASXL1, IDH1/2, SF3B1, and/or SRSF2 mutations. Mutations of RAS-genes are often detected in CMML-like MPN progression. MS ex MN is characterized by complex karyotypes, FLT3 and/or NPM1 mutations, and often monoblastic phenotype. MN with LB transformation is associated with secondary genetic events linked to lineage reprogramming leading to the deregulation of ETV6, IKZF1, PAX5, PU.1, and RUNX1. Finally, the acquisition of MAPK-pathway gene mutations may shape MN toward histiocytic differentiation. Awareness of all these less well-known MN-progression types is important to guide optimal individual patient management.
... Importantly, they highlight the need for greater standardization between trial designs as MF research evolves. Early evidence of a progression toward nontraditional primary end points is encouraging, 32,33 and defining key modifiers will help to establish greater uniformity. ...
The development of targeted therapies for the treatment of myelofibrosis highlights a unique issue in a field that has historically relied on symptom relief, rather than survival benefit or modification of disease course, as key response criteria. There is, therefore, a need to understand what constitutes disease modification of myelofibrosis to advance appropriate drug development and therapeutic pathways. Here, the authors discuss recent clinical trial data of agents in development and dissect the potential for novel end points to act as disease modifying parameters. Using the rationale garnered from latest clinical and scientific evidence, the authors propose a definition of disease modification in myelofibrosis. With improved overall survival a critical outcome, alongside the normalization of hematopoiesis and improvement in bone marrow fibrosis, there will be an increasing need for surrogate measures of survival for use in the early stages of trials. As such, the design of future clinical trials will require re‐evaluation and updating to incorporate informative parameters and end points with standardized definitions and methodologies.
... This remodeling requires direct cell-cell interaction and changes in the expression levels of TGF-β, Notch, and other inflammatory signaling. In addition, in the case of JAK2-driven MPN, it has been shown that in patients and in animal models, the disease is driven by a consistent reduction in the numbers of sympathetic nerve fibers, supporting Schwann cells, which regulates the HSC supporting functions of a subpopulation of nestin + mesenchymal stem cells [48,49]. ...
Serum levels of inflammatory cytokines are currently investigated as prognosis markers in myelofibrosis, the most severe Philadelphia-negative myeloproliferative neoplasm. We tested this hypothesis in the Gata1low model of myelofibrosis. Gata1low mice, and age-matched wild-type littermates, were analyzed before and after disease onset. We assessed cytokine serum levels by Luminex-bead-assay and ELISA, frequency and cytokine content of stromal cells by flow cytometry, and immunohistochemistry and bone marrow (BM) localization of GFP-tagged hematopoietic stem cells (HSC) by confocal microscopy. Differences in serum levels of 32 inflammatory-cytokines between prefibrotic and fibrotic Gata1low mice and their wild-type littermates were modest. However, BM from fibrotic Gata1low mice contained higher levels of lipocalin-2, CXCL1, and TGF-β1 than wild-type BM. Although frequencies of endothelial cells, mesenchymal cells, osteoblasts, and megakaryocytes were higher than normal in Gata1low BM, the cells which expressed these cytokines the most were malignant megakaryocytes. This increased bioavailability of proinflammatory cytokines was associated with altered HSC localization: Gata1low HSC were localized in the femur diaphysis in areas surrounded by microvessels, neo-bones, and megakaryocytes, while wild-type HSC were localized in the femur epiphysis around adipocytes. In conclusion, bioavailability of inflammatory cytokines in BM, rather than blood levels, possibly by reshaping the HSC niche, correlates with myelofibrosis in Gata1low mice.
... As perturbations in the relationship between clonal and nonclonal hematopoietic cells and components of the marrow stem cell niche are gradually elucidated using sophisticated murine models of myeloid malignancies, 55,56,65-68 therapeutic strategies targeting the mediators of tumor cell survival, proliferation, and chemoresistance are beginning to emerge. [69][70][71][72] Translating these findings to human disease and validating novel therapies will require a concerted effort to move from the conventional, subjective, and laborious description of tissue morphologic features by pathologists to objective, quantitative, and automated descriptions of marrow constituents and their interactions. ...
Philadelphia-negative myeloproliferative neoplasms (MPNs) are an excellent tractable disease model of a number of aspects of human cancer biology, including genetic evolution, tissue-associated fibrosis, and cancer stem cells. In this review, we discuss recent insights into MPN biology gained from the application of a number of new single-cell technologies to study human disease, with a specific focus on single-cell genomics, single-cell transcriptomics, and digital pathology.
... In a phase II trial, 39 patients with a JAK2V617F-mutated MPN were treated for 24 weeks with an oral β-3 adrenergic agonist (mirabegron). Although the primary end point (>50% reduction in JAK2V617F allelic burden) was not reached for any of the patients, BM evaluation (available for 20 out of 39 patients) showed an increase of the nestin+ MSCs, a slight decrease in the reticulin fibers and a trend towards a reduction of megakaryocyte clusters [170]. ...
The Philadelphia-negative myeloproliferative neoplasms (MPNs) are malignancies of the hematopoietic stem cell (HSC) arising as a consequence of clonal proliferation driven by somatically acquired driver mutations in discrete genes (JAK2, CALR, MPL). In recent years, along with the advances in molecular characterization, the role of immune dysregulation has been achieving increasing relevance in the pathogenesis and evolution of MPNs. In particular, a growing number of studies have shown that MPNs are often associated with detrimental cytokine milieu, expansion of the monocyte/macrophage compartment and myeloid-derived suppressor cells, as well as altered functions of T cells, dendritic cells and NK cells. Moreover, akin to solid tumors and other hematological malignancies, MPNs are able to evade T cell immune surveillance by engaging the PD-1/PD-L1 axis, whose pharmacological blockade with checkpoint inhibitors can successfully restore effective antitumor responses. A further interesting cue is provided by the recent discovery of the high immunogenic potential of JAK2V617F and CALR exon 9 mutations, that could be harnessed as intriguing targets for innovative adoptive immunotherapies. This review focuses on the recent insights in the immunological dysfunctions contributing to the pathogenesis of MPNs and outlines the potential impact of related immunotherapeutic approaches.
... However, Nestin + cells, as welldocumented MSCs in the BM (Mendez-Ferrer et al., 2010), have been shown to be 4-5-fold more abundant in human AML patients, in line with expansion of Nestin + cells in the murine iMLL-AF9 AML model (Hanoun et al., 2014;Forte et al., 2020). This is in striking contrast to the decrease in Nestin + cells in murine models and human MPN (Arranz et al., 2014;Drexler et al., 2019), suggesting that the same group of niche cells can behave differently in various myeloid malignancies and/or stages of leukemic disease. Conditional depletion of Nestin + cells upon AML development in iMLL-AF9 mice lead to a significantly extended mouse survival, suggesting that Nestin + cells promote leukemogenesis in vivo (Forte et al., 2020). ...
The contribution of bone marrow stromal cells to the pathogenesis and therapy response of myeloid malignancies has gained significant attention over the last decade. Evidence suggests that the bone marrow stroma should not be neglected in the design of novel, targeted-therapies. In terms of gene-editing, the focus of gene therapies has mainly been on correcting mutations in hematopoietic cells. Here, we outline why alterations in the stroma should also be taken into consideration in the design of novel therapeutic strategies but also outline the challenges in specifically targeting mesenchymal stromal cells in myeloid malignancies caused by somatic and germline mutations.
... This SDF1 release negatively regulates the proliferation of JAK2V617F HSCs [303], and its expression is inhibited by the action of noradrenaline, which is released by sympathetic nerve fibres, on the β-adrenergic receptors of nestin + MSCs [304,305]. In MPN, LSCs secretes IL-1β, leading to Schwann cell death and subsequent sympathetic neuropathy [256,303,305]. This results in apoptosis of nestin + MSCs [256]. ...
... In MPN, LSCs secretes IL-1β, leading to Schwann cell death and subsequent sympathetic neuropathy [256,303,305]. This results in apoptosis of nestin + MSCs [256]. β-3 sympathomimetic agonist restores sympathetic regulation of nestin + MSCs. ...
... β-3 sympathomimetic agonist restores sympathetic regulation of nestin + MSCs. In JAK2V617F murine models, β-3 sympathomimetic agonist (BRL37344) rescued nestin + MSC, reduced BM fibrosis and diminished MPN HPCs [256,303,305]. Mirabegron, another β-3 sympathomimetic agonist, showed some encouraging results in a phase II clinical trial [256] (Table 3). ...
Myeloproliferative neoplasms (MPNs) are unique hematopoietic stem cell disorders sharing mutations that constitutively activate the signal-transduction pathways involved in haematopoiesis. They are characterized by stem cell-derived clonal myeloproliferation. The key MPNs comprise chronic myeloid leukemia (CML), polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). CML is defined by the presence of the Philadelphia (Ph) chromosome and BCR-ABL1 fusion gene. Despite effective cytoreductive agents and targeted therapy, complete CML/MPN stem cell eradication is rarely achieved. In this review article, we discuss the novel agents and combination therapy that can potentially abnormal hematopoietic stem cells in CML and MPNs and the CML/MPN stem cell-sustaining bone marrow microenvironment.
... Further investigations may reveal novel and useful immunotherapies. In addition, several microenvironmental targets such as IL-1b pathway and IL-1 antagonists or b3-AR agonists for sympathetic neuropathy (such as mirabegron) (260), mitochondrial heterocellular transfer, and inhibition by tigecycline, fatty acid oxidation (FAO) and FAO inhibitor etomoxir that sensitizes AML cells to therapeutic challenge, NO synthase inhibitors (such as cavtratin) (261,262) and endothelium activation (59), are under investigation. ...
Recent studies have provided several insights into acute myeloid leukemia. Studies based on molecular biology have identified eight functional mutations involved in leukemogenesis, including driver and passenger mutations. Insight into Leukemia stem cells (LSCs) and assessment of cell surface markers have enabled characterization of LSCs from hematopoietic stem and progenitor cells. Clonal evolution has been described as having an effect similar to that of microenvironment alterations. Such biological findings have enabled the development of new targeted drugs, including drug inhibitors and monoclonal antibodies with blockage functions. Some recently approved targeted drugs have resulted in new therapeutic strategies that enhance standard intensive chemotherapy regimens as well as supportive care regimens. Besides the progress made in adoptive immunotherapy, since allogenic hematopoietic stem cell transplantation enabled the development of new T-cell transfer therapies, such as chimeric antigen receptor T-cell and transgenic TCR T-cell engineering, new promising strategies that are investigated.
... The drug was well tolerated, with similar to BAT cardiac and bleeding adverse events, but more frequent diarrhea, nausea, and vomiting [19]. Other therapeutic strategies currently under investigation include apoptosis induction of the hematopoietic progenitor or stem cells and reversal of the altered bone marrow microenvironment [20,21]. ...
Primary myelofibrosis (PMF) is a pluripotent hematopoietic stem cell-derived malignancy, included in the heterogeneous group of myeloproliferative neoplasms (MPNs). PMF diagnosis is based on a composite assessment of clinical and laboratory data. The three major diagnostic criteria are: screening for driver mutations, exclusion of other conditions that can cause myelofibrosis, and bone marrow biopsy displaying megakaryocyte changes and fibrosis. PMF treatment options are only partially disease-modifying and consist mainly of symptom control. Recently, a new targeted therapy was introduced for PMF patients, JAK-STAT inhibitors (i.e. ruxolitinib). However, specific subgroups of patients do not benefit from the JAK-STAT inhibitors: (1) those who are carrying JAK2 mutations, but ruxolitinib does not reduce the spleen size; (2) triple negative patients (no JAK2, CALR, or MPL mutations); and (3) those who discontinue JAK-STAT therapy because of side effects. These subgroups are in need of new therapeutic approaches. Mature microRNAs (miRNAs) range from 16 to 28 nucleotides (nt) in length and regulate specific messenger RNAs at the post-transcriptional level. Numerous in vitro and in vivo studies have reported specific miRNAs, as well as complex miRNA networks, to be dysregulated in PMF. Several of these miRNAs were shown to be implicated in essential events of PMF pathophysiology: increase of bone marrow fibrosis, progression to acute myeloid leukemia, resistance to JAK-STAT inhibitors, and activation of differentiation of hematopoietic stem/progenitor cells into megakaryocytes. Hence, we propose miRNAs as a potential minimally invasive diagnostic tool for PMF and as therapeutic targets that could address the unmet medical needs of these patients.
... Defective β3-adrenoceptor signaling was previously identified as the underlying cause of MSC loss in a mouse model of MPN [47]. Follow-up work using the β3-adrenoceptor agonist mirabegron in a phase II study failed to reach the primary end point of JAK2-V617F allele burden reduction but demonstrated improvement of BM fibrosis and increase of MSCs after 24 weeks of mirabegron treatment [73]. ...
The interaction of hematopoietic stem and progenitor cells with their direct neighboring cells in the bone marrow (the so called hematopoietic niche) evolves as a key principle for understanding physiological and malignant hematopoiesis. Significant progress in this matter has recently been achieved making use of emerging high-throughput techniques that allow characterization of the bone marrow microenvironment at single cell resolution. This review aims to discuss these single cell findings in the light of other conventional niche studies that together define the current notion of the niche’s implication in i) normal hematopoiesis, ii) myeloid neoplasms and iii) disease-driving pathways that can be exploited to establish novel therapeutic strategies in the future.
