Myelodysplastic syndromes (MDS) are clonal hematologic disorders characterized by morphologic abnormalities of myeloid cells and peripheral cytopenias. Although genetic abnormalities underlie the pathogenesis of these disorders and their heterogeneity, current classifications of MDS rely predominantly on morphology. We performed genomic profiling of 3233 patients with MDS or related disorders to delineate molecular subtypes and define their clinical implications. Gene mutations, copy-number alterations, and copy-neutral loss of heterozygosity were derived from targeted sequencing of a 152-gene panel, with abnormalities identified in 91%, 43%, and 11% of patients, respectively. We characterized 16 molecular groups, encompassing 86% of patients, using information from 21 genes, 6 cytogenetic events, and loss of heterozygosity at the TP53 and TET2 loci. Two residual groups defined by negative findings (molecularly not otherwise specified, absence of recurrent drivers) comprised 14% of patients. The groups varied in size from 0.5% to 14% of patients and were associated with distinct clinical phenotypes and outcomes. The median bone marrow (BM) blast percentage across groups ranged from 1.5% to 10%, and the median overall survival ranged from 0.9 to 8.2 years. We validated 5 well-characterized entities, added further evidence to support 3 previously reported subsets, and described 8 novel groups. The prognostic influence of BM blasts depended on the genetic subtypes. Within genetic subgroups, therapy-related MDS and myelodysplastic/myeloproliferative neoplasms had comparable clinical and outcome profiles to primary MDS. In conclusion, genetically-derived subgroups of MDS are clinically relevant and might inform future classification schemas and translational therapeutic research.

Mesenchymal stromal cells (MSCs) are an essential element of the bone marrow (BM) microenvironment, playing a crucial function in regulating hematopoietic stem cell proliferation and differentiation. Recent findings have outlined a putative role for MSCs in hematological malignancy development. So far, conflicting results have been collected concerning MSC abnormalities in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). In particular, a considerable amount of evidence has been accumulated strongly supporting a permissive role of MSCs in malignancy evolution to MDS, while a potentially causative or promoting function performed by MSCs in AML has not yet been fully clarified. Here, we compared MSCs isolated from healthy, MDS, and AML subjects to investigate MSC alterations and to emphasize putative common and/or diverse features. We isolated and expanded MSCs from AML patients (AML-MSCs) and MDS patients (MDS-MSCs), and we analyzed and compared their phenotypic and functional properties with respect to each other and versus healthy donor-derived MSCs (HD-MSCs). We found that stable MSC cultures could be easily established from HD and MDS mononuclear BM-derived cells, while a substantial fraction (25%) of AML patients failed to yield MSCs. Nevertheless, isolated MDS-MSCs and AML-MSCs, as well as HD-MSCs, contained the basic features of MSCs. Indeed, they displayed similar surface marker expression and efficient capacity to differentiate versus osteogenic and adipogenic lineage in vitro. We also proved that MDS-MSCs and AML-MSCs, analyzed by fluorescence in-situ hybridization, did not harbor leukemic cell cytogenetic abnormalities. Moreover, MDS-MSCs and AML-MSCs were similar in terms of ability to sustain AML cell viability and immune-regulatory capacity. However, we were also able to detect some differences between AML-MSCs and MDS-MSCs. Indeed, we found that the frequency of rescued MSCs was lower in the AML group than in the HD and MDS groups, suggesting that a reduced number of MSC precursors could inhabit AML BM. Instead, MDS-MSCs showed the lowest proliferative capacity, reflecting some intrinsic and particular defect. Overall, our results elucidated that MDS-MSCs and AML-MSCs did not show macroscopic and/or tumor-related defects, but both displayed functional features potentially contributing to favor a leukemia-protective milieu.

Mutations in UBA1, which are disease-defining for VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome, have been reported in patients diagnosed with myelodysplastic syndromes (MDS). Here, we define the prevalence and clinical associations of UBA1 mutations in a representative cohort of patients with MDS. Digital droplet polymerase chain reaction profiling of a selected cohort of 375 male patients lacking MDS disease-defining mutations or established World Health Organization (WHO) disease classification identified 28 patients (7%) with UBA1 p.M41T/V/L mutations. Using targeted sequencing of UBA1 in a representative MDS cohort (n = 2027), we identified an additional 27 variants in 26 patients (1%), which we classified as likely/pathogenic (n = 12) and of unknown significance (n = 15). Among the total 40 patients with likely/pathogenic variants (2%), all were male and 63% were classified by WHO 2016 criteria as MDS with multilineage dysplasia or MDS with single-lineage dysplasia. Patients had a median of 1 additional myeloid gene mutation, often in TET2 (n = 12), DNMT3A (n = 10), ASXL1 (n = 3), or SF3B1 (n = 3). Retrospective clinical review, where possible, showed that 82% (28/34) UBA1-mutant cases had VEXAS syndrome–associated diagnoses or inflammatory clinical presentation. The prevalence of UBA1 mutations in patients with MDS argues for systematic screening for UBA1 in the management of MDS.

Background: Therapy-related myeloid neoplasms (t-MN), including myelodysplastic syndromes and acute myeloid leukemia (t-MDS and t-AML) are associated to clinical and biologic unfavorable prognostic features, including high levels of DNA methylation. Methods: We retrospectively evaluated 50 t-MN patients (34 MDS and 16 AML) selected among all patients receiving azacitidine (AZA) at 10 Italian Hematology Centers. Patients had developed a t-MN at a median of 6.5 years (range 1.7- 29) after treatment of the primary tumor (hematological neoplasm, 27 patients; solid tumor, 23 patients). Results: The overall response rate was 42% (complete remission: 10 patients, partial remission: 2 and hematological improvement: 8 patients) and was obtained after a median of 3 cycles (range 1–6). Median overall survival (OS) was 21 months (range 1–53.6+) from AZA start. OS was significantly better in patients with less than 20% blasts, in normal karyotype t-AML and when AZA was used as front-line treatment. This was confirmed by the multivariate analysis. Conclusions: This study reports efficacy of AZA in the largest series of therapy-related MN patients treated with 5-AZA. Our data show that blasts and karyotype maintain their important prognostic role in t-MN also in the azacitidine era.

Erythropoiesis regulation is essential in normal physiology and pathology, particularly in myelodysplastic syndromes (MDS) and β-thalassemia. Several signaling transduction processes, including those regulated by inositides, are implicated in erythropoiesis, and the latest MDS or β-thalassemia preclinical and clinical studies are now based on their regulation. Among others, the main pathways involved are those regulated by transforming growth factor (TGF)-β, which negatively regulates erythrocyte differentiation and maturation, and erythropoietin (EPO), which acts on the early-stage erythropoiesis. Also small mother against decapentaplegic (SMAD) signaling molecules play a role in pathology, and activin receptor ligand traps are being investigated for future clinical applications. Even inositide-dependent signaling, which is important in the regulation of cell proliferation and differentiation, is specifically associated with erythropoiesis, with phospholipase C (PLC) and phosphatidylinositol 3-kinase (PI3K) as key players that are becoming increasingly important as new promising therapeutic targets. Additionally, Roxadustat, a new erythropoiesis stimulating agent targeting hypoxia inducible factor (HIF), is under clinical development. Here, we review the role and function of the above-mentioned signaling pathways, and we describe the state of the art and new perspectives of erythropoiesis regulation in MDS and β-thalassemia.

PI‐PLCβ1 is involved in cell proliferation, differentiation, and myelodysplastic syndrome (MDS) pathogenesis. Moreover, the increased activity of PI‐PLCβ1 reduces the expression of PKC‐α, which, in turn, delays the cell proliferation and is linked to erythropoiesis. Lenalidomide is currently used in low‐risk patients with MDS and del (5q), where it can suppress the del(5q) clone and restore normal erythropoiesis. In this study, we analyzed the effect of lenalidomide on 16 patients with low‐risk del(5q) MDS, as well as del(5q) and non‐del(5q) hematopoietic cell lines, mainly focusing on erythropoiesis, cell cycle, and PI‐PLCβ1/PKC‐a signaling. Overall, 11 patients were evaluated clinically, and 10 (90%) had favorable responses; the remaining case had a stable disease. At a molecular level, both responder patients and del(5q) cells showed a specific induction of erythropoiesis, with a reduced γ /β‐globin ratio, an increase in glycophorin A, and a nuclear translocation of PKC‐α. Moreover, lenalidomide could induce a selective G ₀ /G ₁ arrest of the cell cycle in del(5q) cells, slowing down the rate proliferation in those cells. Altogether, our results could not only better explain the role of PI‐PLC β1/PKC‐α signaling in erythropoiesis but also lead to a better comprehension of the lenalidomide effect on del(5q) MDS and pave the way to innovative, targeted therapies.—Poli, A., Ratti, S., Finelli, C., Mongiorgi, S., Clissa, C., Lonetti, A., Cappellini, A., Catozzi, A., Barraco, M., Suh, P.‐G., Manzoli, L., McCubrey, J. A., Cocco, L., Follo, M. Y. Nuclear translocation of PKC‐α is associated with cell cycle arrest and erythroid differentiation in myelodysplastic syndromes (MDSs). FASEB J. 32, 681–692 (2018). www.fasebj.org