Myelodysplastic Syndromes Cells Research Articles

A new therapeutic perspective: Erastin inhibits tumor progression by driving ferroptosis in myelodysplastic syndromes.

Ferroptosis is a recently identified and evolutionarily conserved form of programmed cell death. This process is initiated by an imbalance in iron metabolism, leading to an overload of ferrous ions. These ions promote lipid peroxidation in the cell membrane through the Fenton reaction. As the cell's antioxidant defenses become overwhelmed, a fatal buildup of reactive oxygen species (ROS) occurs, resulting in the rupture of the plasma membrane. Ferroptosis is implicated in conditions such as ischemia-reperfusion injuries and a range of cancers. In our research, we explored ferroptosis in myelodysplastic syndromes (MDS) by measuring iron levels, transferrin receptor expression, and glutathione peroxidase 4 (GPX4) mRNA. Our findings revealed that MDS patients had significantly higher Fe2+ levels in CD33+ cells and increased transferrin receptor mRNA compared to healthy individuals. GPX4 expression was also higher in MDS but not statistically significant. To investigate potential treatments for myeloid hematological diseases through ferroptosis induction, we treated the myelodysplastic syndrome cell line (SKM-1) and two myeloid leukemia cell lines (KG-1 and K562) with erastin, an iron transfer inducer. We observed that erastin treatment led to glutathione depletion, reduced GPX4 activity, and increased ROS, culminating in cell death by ferroptosis. Furthermore, combining erastin with azacitidine demonstrated a synergistic effect on MDS and leukemia cell lines, suggesting a promising approach for treating these hematological conditions with this drug combination. Our experiments confirm erastin's ability to induce ferroptosis in MDS and highlight its potential synergistic use with azacitidine for treatment.

Elesclomol-copper synergizes with imidazole ketone erastin by promoting cuproptosis and ferroptosis in myelodysplastic syndromes

Myelodysplastic syndromes (MDS) encompass a collection of clonal hematopoietic malignancies distinguished by the depletion of peripheral blood cells. The treatment of MDS is hindered by the advanced age of patients, with a restricted repertoire of drugs currently accessible for therapeutic intervention. In this study, we found that ES-Cu strongly inhibited the viability of MDS cell lines and activated cuproptosis in a copper-dependent manner. Importantly, ferroptosis inducer IKE synergistically enhanced ES-Cu-mediated cytotoxicity both in vitro and in vivo. Of note, the combination of IKE and ES-Cu intensively impaired mitochondrial homeostasis with increased mitochondrial ROS, MMP hyperpolarized, down-regulated iron-sulfur proteins and declined oxygen consumption rate. Additionally, ES-Cu/IKE treatment could enhance the lipoylation-dependent oligomerization of the DLAT. To elucidate the specific order of events in the synergistic cell death, inhibitors of ferroptosis and cuproptosis were utilized to further characterize the basis of cell death. Cell viability assays showed that the glutathione and its precursor N-acetylcysteine could significantly rescue the cell death under either mono or combination treatment, demonstrating that GSH acts at the crossing point in the regulation network of cuproptosis and ferroptosis. Significantly, the reconstitution of xCT expression and knockdown of FDX1 cells have been found to contribute to the tolerance of mono treatment but have little recovery impact on the combined treatment. Collectively, these findings suggest that a synergistic interaction leading to the induction of multiple programmed cell death pathways could be a promising approach to enhance the effectiveness of therapy for MDS.

Systemic inflammatory autoimmune disease before allogeneic hematopoietic stem cell transplantation is a risk factor for death in patients with myelodysplastic syndrome or chronic myelomonocytic leukemia.

Myelodysplastic syndrome (MDS) is well known to be complicated by systemic inflammatory autoimmune disease (SIADs). However, it remains unclear how the prognosis after allogenic hematopoietic stem cell transplantation (allo-HSCT) in patients with MDS is impacted by SIADs that occur before allo-HSCT. Therefore, we hypothesized that SIADs before allo-HSCT may be a risk factor for negative outcomes after allo-HSCT in patients with MDS. We conducted a single-center, retrospective, observational study of sixty-nine patients with MDS or chronic myelomonocytic leukemia who underwent their first allo-HCT. Fourteen of the patients had SIADs before allo-HSCT. In multivariate analysis, the presence of SIADs before allo-HSCT was an independent risk factor for overall survival (HR, 3.36, 95% confidence interval: 1.34-8.42, p = 0.009). Endothelial dysfunction syndrome was identified in five of 14 patients with SIADs who required immunosuppressive therapy or intensive chemotherapy, and notably, all patients with uncontrollable SIADs at allo-HSCT developed serious endothelial dysfunction syndrome and died in the early phase after allo-HSCT. The development of SIADs in the context of MDS is thought to reflect the degree of dysfunction of hematopoietic cells in MDS and suggests a higher risk of disease progression. In addition, MDS patients with SIADs before allo-HSCT are considered to be at higher risk of endothelial dysfunction syndrome because of preexisting vascular endothelial dysfunction due to SIADs. In conclusion, SIADs before allo-HSCT constitute an independent risk factor for death in MDS patients undergoing allo-HSCT.

Comprehensive morphologic characterization of bone marrow biopsy findings in a large cohort of patients with VEXAS syndrome: A single-institution longitudinal study of 111 bone marrow samples from 52 patients.

VEXAS syndrome is an adult-onset autoinflammatory disease caused by a somatic pathogenic mutation in the UBA1 (ubiquitin-like modifier activating enzyme 1) gene. Patients present with rheumatologic manifestations and cytopenias and may have an increased predisposition to myelodysplastic syndrome (MDS) and plasma cell neoplasms. Prior studies have reported on the peripheral blood and bone marrow findings in patients with VEXAS syndrome. Due to the protean clinical presentation and lack of specificity of morphologic features (eg, vacuoles in early erythroid and granulocytic precursors), an optimal screening methodology to identify these patients in a timely fashion is desirable. To further evaluate and describe the salient diagnostic morphologic features in VEXAS syndrome, we carried out a comprehensive study of the largest single-institution cohort to date. Diagnostic and follow-up bone marrow biopsy specimens from 52 male patients with molecularly identified VEXAS syndrome underwent central review. Cytopenias were common in all cases, primarily macrocytic anemia, monocytopenia, and thrombocytopenia. Bone marrow aspirate and biopsy were often hypercellular, with an increased myeloid/erythroid ratio, granulocytic hyperplasia with left shift, erythroid left shift, and megakaryocyte hyperplasia, which exhibited a range of striking morphologic findings. Distinctly vacuolated myeloid and erythroid precursors were seen in more than 95% of cases. Our data reveal potential novel diagnostic features, such as a high incidence of monocytopenia and distinct patterns of atypical megakaryopoiesis, that appear different from dysmegakaryopoiesis typically associated with MDS. In our experience, those findings are suggestive of VEXAS, in the appropriate clinical context.

Ectopic expression of the transcription factor ONECUT3 drives a complex karyotype in myelodysplastic syndromes.

Chromosomal instability is a prominent biological feature of myelodysplastic syndromes (MDS), with over 50% of patients with MDS harboring chromosomal abnormalities or a complex karyotype (CK). Despite this observation, the mechanisms underlying mitotic and chromosomal defects in MDS remain elusive. In this study, we identified ectopic expression of the transcription factor ONECUT3, which is associated with CKs and poorer survival outcomes in MDS. ONECUT3-overexpressing cell models exhibited enrichment of several notable pathways, including signatures of sister chromosome exchange separation and mitotic nuclear division with the upregulation of INCENP and CDCA8 genes. Notably, dysregulation of chromosome passenger complex (CPC) accumulation, besides the cell equator and midbody, during mitotic phases consequently caused cytokinesis failure and defective chromosome segregation. Mechanistically, the homeobox (HOX) domain of ONECUT3, serving as the DNA binding domain, occupied the unique genomic regions of INCENP and CDCA8 and transcriptionally activated these 2 genes. We identified a lead compound, C5484617, that functionally targeted the HOX domain of ONECUT3, inhibiting its transcriptional activity on downstream genes, and synergistically resensitized MDS cells to hypomethylating agents. This study revealed that ONECUT3 promoted chromosomal instability by transcriptional activation of INCENP and CDCA8, suggesting potential prognostic and therapeutic roles for targeting high-risk MDS patients with a CK.

Minor introns impact on hematopoietic malignancies

In the intricate orchestration of the central dogma, pre-mRNA splicing plays a crucial role in the post-transcriptional process that transforms DNA into mature mRNA. Widely acknowledged as a pivotal RNA processing step, it significantly influences gene expression and alters the functionality of gene product proteins. Although U2-dependent spliceosomes efficiently manage the removal of over 99% of introns, a distinct subset of essential genes undergo splicing with a different intron type, denoted as minor introns, using U12-dependent spliceosomes. Mutations in spliceosome component genes are now recognized as prevalent genetic abnormalities in cancer patients, especially those with hematologic malignancies. Despite the relative rarity of minor introns, genes containing them are evolutionarily conserved and play crucial roles in functions such as the RAS-MAPK pathway. Disruptions in U12-type minor intron splicing caused by mutations in snRNA or its regulatory components significantly contribute to cancer progression. Notably, recurrent mutations associated with myelodysplastic syndrome (MDS) in the minor spliceosome component ZRSR2 underscore its significance. Examination of ZRSR2-mutated MDS cells has revealed that only a subset of minor spliceosome-dependent genes, such as LZTR1, consistently exhibit missplicing. Recent technological advancements have uncovered insights into minor introns, raising inquiries beyond current understanding. This review comprehensively explores the importance of minor intron regulation, the molecular implications of minor (U12-type) spliceosomal mutations and cis-regulatory regions, and the evolutionary progress of studies on minor, aiming to provide a sophisticated understanding of their intricate role in cancer biology.

Myelodysplastic syndromes del(5q): Pathogenesis and its therapeutic implications.

Myelodysplastic syndromes (MDS) encompass a heterogeneous set of acquired bone marrow neoplastic disorders characterized by ineffective hematopoiesis within one or more bone marrow lineages. Nearly half of MDS patients carry cytogenetic alterations, with del(5q) being the most prevalent. Since its first description, del(5q) was consistently correlated with a typical clinical phenotype marked by anemia, thrombocytosis, and a low risk of evolving into acute leukemia. Presently, the World Health Organization (WHO) classification of myeloid neoplasms recognizes a specific subtype of MDS known as "myelodysplastic neoplasm with low blast and isolated del(5q)" identified by the sole presence of 5q deletion or in combination with one other abnormality excluding -7/del(7q). Several studies have sought to unravel the biological processes triggered by del(5q) in the development of MDS, revealing the involvement of various genes localized in specific regions of chromosome 5 referred to as common deleted regions (CDR). This intricate biological landscape makes the MDS cells with del(5q) exceptionally sensitive to lenalidomide. Several studies have confirmed the efficacy of lenalidomide in this context. Regrettably, the response to lenalidomide is not conclusive, prompting ongoing research into biological mechanisms that drive patients toward leukemia and strategies to circumvent lenalidomide resistance and disease progression.

MDS-MSC Derived Exosomes Promote MDS Cells Malignant Proliferation through RIG-1/Isgs Pathway Via RN7SL1

The interactions between bone marrow stem cells (BM-MSCs) and myelodysplastic syndromes (MDS) cells perform a crucial role in MDS pathogenesis by secreting cytokines ,growth factors, and exosomes. Although BM-MSCs induced growth of MDS cells has been studied, the role of exosomes from MDS-derived MSC (MDS-MSC-EXO) in this action remains unclear. In our study, we investigate the effect of MDS-MSC-EXO on the viability, proliferation, migration, cellular apoptosis of MDS cell lines, such as MDS-L and SKM-1. Our data identified that MDS-MSC-EXO can promote proliferation, migration, and decrease cell apoptosis. Both LR-MDS-MSC and HR-MDS-MSC-derived exosomes increased MDS cell growth.Exosomes mediate local cell-to-cell communication by transferring mRNAs, miRNAs, and proteins. In our study, we found that MDS-MSC-EXO expressed higher unshielded RN7SL1,an endogenous RNA normally shielded by RNA binding proteins SRP9/14, than that from health controls (HC-MSC-EXO). MDS cloned cells expressed higher retinoic acid inducible gene-I (RIG-I) and interferon-stimulated genes (ISGs), such as ISG15, IFIT1 and IFITM1, which are the receptor and effector molecules of RN7SL1. Both MDS-MSC-EXO and exogenous RN7SL1 can promote MDS cloned cells proliferation and stimulate ISGs expression by activating RIG-1. RIG-I inhibitor can block this effect. We demonstrate that MDS-MSC may promote MDS cloned cell proliferation through RIG-ISGs signal activated by exosomes-derived RN7SL1. Overall, our studies provide new insights that MSC-EXO functions the crucial roles in mediating the progression of MDS via RIG-1/ISGs activation, and demonstrate novel targets of RN7SL1 to the pathogenesis of MDS.This study will provide new ideas for MDS treatment in the aspect of microenvironment.

Distinct Role of STAT3 and STAT5 Factors in MDS and AML

Introduction: Myelodysplastic Syndrome (MDS) refers to a heterogeneous group of clonal hematologic disorders characterized by hematopoietic dysfunctions and impaired differentiation in the hematopoietic stem and progenitor cells compartment. MDS is commonly referred to as a pre-leukemic stage due to the increased risk of progression to Acute Myeloid Leukemia (AML). AML is an aggressive hematologic malignancy characterized by the accumulation of immature myeloblasts in the bone marrow and the peripheral blood. Signal transducers and activators of transcription 3 and 5 (STAT3 and STAT5) are important regulators of several cellular processes including cell proliferation and survival. Abnormal STAT3 and STAT5 signaling has been implicated in various solid and hematologic malignancies, including MDS and AML, which renders them appealing targets for the development of novel therapeutic strategies. Materials: STAT3, STAT5A or STAT5B knock-downs have been generated in two AML (MOLM-13 and Kasumi-1) and two MDS (MDS92 and MDS-L, gift of Prof. Kaoru Tohyama) cell lines, using shRNAs through lenti-viral delivery. The 4 biological replicates with the highest levels of knock-downs for STAT5A, STAT5B or STAT3 have been selected for each cell line and mRNA sequencing has been performed. To define the genome-wide binding of STAT5A, STAT5B and STAT3 in the two conditions, CUT&Tag has been performed in wild type MOLM-13, Kasumi-1 and MDS-L cells. Additionally, ATAC sequencing has been carried out in the same cell lines to investigate the chromatin conformation changes between the two conditions. Furthermore, STAT3, STAT5A and STAT5B knock-downs have been conducted using shRNAs through lenti-viral delivery in CD34+ enriched cells, isolated from bone marrow mononuclear cells of 14 high-risk MDS patients and 14 secondary (MDS-related) AML patients. The samples have been processed for mRNA-Sequencing to model the leukemic transformation in primary human cells. Results: To elucidate the role of STAT3 and STAT5 in MDS to AML transformation, changes in STAT3 and STAT5 target gene networks have been identified. mRNA sequencing data from STAT3, STAT5A or STAT5B knock-downs in MDS and AML cell lines revealed a distinct role of the three factors in each state. The differential binding of each factor, explored through CUT&Tag, and the gene expression profiles in MDS and AML reciprocally reveal a different function of each factor, including pathways involved in cell cycle and apoptosis. Also, the direct and indirect target genes of STAT3, STAT5A and STAT5B in each condition, as well as a possible interplay between STAT3 and STAT5 activation in AML has been uncovered. The exploration of the chromatin accessibility landscapes between the MDS and AML conditions through ATAC sequencing has provided an insight into the chromatin conformation states, unveiling potential leukemia promoting networks, as well as the involvement of STAT3 and STAT5 pathways in the malignant transformation. Furthermore, a potential cross talk of STAT3 and STAT5 factors with the IKAROS family members, has been linked to AML transformation, as significant differential regulation has been detected between MDS and AML conditions after knock-down of STAT5A and STAT5B. The results are currently validated in primary hematopoietic cells of MDS and AML patients after STAT3, STAT5A or STAT5B knock-downs. Conclusions: Our results show a different role between the two STAT5 factors (STAT5A and STAT5B) in MDS and AML conditions and delineate the role of STAT5A, STAT5B and STAT3 factors and their target gene networks in leukemic transformation, thus providing novel targets for therapeutic management of MSD and AML. Funding: This project is funded by the European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 813091 (ARCH)

Trisomy 8 Remodels Chromatin and Activates Transcription of Runx1-Target Genes in Hematopoietic Stem Cell

Myelodysplastic syndrome (MDS) is a poor prognosis cancer that predominantly affects the elderly, arising from hematopoietic stem cell (HSC), resulting in bone marrow failure and predisposition to acute myelogenous leukemia (AML). It has long been known that numerical chromosome anomalies such as trisomy 8 are critical for the development of MDS. Trisomy 8 is known to be associated with relatively poor prognosis, and is a criterion for the diagnosis of MDS in patients. Systemic inflammation has been implicated in the selective advantage for clonal hematopoietic stem cells and the progression of MDS, such as trisomy 8 MDS associated with Behcet's disease. Next generation sequencing studies revealed that trisomy 8 MDS cells highly mutated RUNX1 transcriptional factor (TF) and ASXL1, an epigenetic modifier, both of which regulate HSC function and the differentiation, relative to MDS cells without trisomy 8, suggesting that those mutations may help trisomy 8 to drive the development of MDS. However, previous clinical studies failed to identify genes on the chromosome 8 commonly amplified among trisomy 8 MDS patients, which may account for the pathogenesis of MDS, the molecular mechanism of development of trisomy 8 MDS remains unclear. Thus, we hypothesized that trisomy 8 initiates the transformation by multiple malfunctions of epigenetic and transcriptional regulators in HSC other than a gene dosage effect due to the extra chromosome 8. In order to determine how trisomy 8 affects hematopoiesis, we established a new trisomy 8 model by introducing a human chromosome 8 to mimic trisomy 8 in mouse ES cells. We succeeded in generating trisomy 8 chimeric mice and confirmed the mice showing emergence of hematopoiesis in fetal liver but significantly decreased the chimerism of trisomy 8 cells than the control mice. Wecompetitively transplanted trisomy 8 fetal liver cells at E14.5 into lethally-irradiated wild-type primary recipient mice, and moreover serial transplanted hematopoietic stem and progenitor cells (HSPCs) which isolated from the primary recipient mice. We found that trisomy 8 reduced the self-renewal capacity of HSPC and impeded the differentiation, but it was not sufficient to develop myeloid malignancies in mice. To elucidate the molecular mechanism underlying the impaired hematopoiesis by trisomy 8, we performed RNA sequencing in HSCs isolated from the primary transplanted mice. Gene set enrichment analysis revealed that in comparison with the control HSCs, trisomy 8 HSCs had significantly positive enrichments in type I and type II interferon response genes, but also the canonical target genes of Runx1 and PU.1 TFs were positively enriched in trisomy 8 HSCs. In addition, ATAC-seq analysis revealed that trisomy 8 HSCs changed chromatin accessibility enriched with the binding motif of Runx TF, compared to control HSCs. Given the changes in chromatin accessibility in trisomy 8 HSC, we performed chromatin conformation capture sequencing (Hi-C) in HSPCs and found that trisomy 8 induced changes in chromatin structures such as topologically associating domain (TAD) and compartment A/B in the other chromosomes, which were partly associated with changes in transcription of genes. Since the Runx1 gene is critical for HSC integrity and differentiation, and loss-of-function mutations of RUNX1 were often found in human myeloid malignancies with trisomy 8, we attempted to determine whether Runx1 deletion cancel the impaired self-renewal capacity of trisomy 8 HSC. We found that Runx1 deletion partially canceled the impaired repopulating capacity of trisomy 8 cells in recipient mice, suggesting the selective advantage of loss of function mutation of the RUNX1 gene in trisomy 8 mosaic people and patients with trisomy 8 MDS. Furthermore, as Janus kinase (JAK) and TANK-binding kinase 1 (TBK1) are critical for interferon signalling and activation of their downstream target genes, we revealed that the inhibition of either JAK or TBK function ameliorated trisomy 8-induced chromatin remodelling in HSCs. Overall, trisomy 8 changes chromatin structure and transcription of genes on other chromosomes in HSC accompanied with the malfunction of Runx1 TF, at least in part, due to the activation of interferon signaling, and results in the impaired hematopoiesis. We demonstrated that trisomy 8 creates the basis for transformation of MDS via remodeling chromatin structures in HSC.

Safety and Efficacy of Selinexor Sequential Azacytidine in Newly Diagnosed Patients with Myelodysplastic Syndromes EB1 or EB2: A Single-Center, Single-Arm, Phase Ib/II Trial

Background: Azacytidine is one of the hypomethylating agents (HMAs) for the treatment of higher-risk myelodysplastic syndromes (MDS). However, clinical outcomes of patients treated with azacytidine monotherapy were far from satisfactory with overall response rate (ORR) of 41%-59% and overall survival (OS) of 15-18 months. Selinexor is an oral, first-in-class selective inhibitor of nuclear export compound that inhibits exportin 1 (XPO1). Preclinical studies have shown that inhibition of XPO1 causes nuclear accumulation of p53 and disruption of NF-κB signaling, both relevant targets for myelodysplastic syndromes. Prior study reported an ORR of 26% and mOS of 8.7 months with selinexor in HMAs-refractory MDS patients. We therefore aimed to assess the safety and efficacy of selinexor sequential azacytidine in newly diagnosed patients with MDS-EB1 or EB2. Methods: The effectiveness of simultaneous and sequential treatment of azacytidine and selinexor was measured by cell death rate, cell cycle and colony formation rate in vitro. Western Blot was performed to detect the sublocation of p53. MOLM-13-luciferase xenograft murine model was established to verify the synergetic effect in vivo. Based on the experiments, we conducted a single-center, single-arm, phase Ib/II trial, which included newly diagnosed patients 18 years or older with MDS-EB1 or EB2. Eligible patients received 2-4 weeks long cycles of oral selinexor (40mg qw for 4 weeks; 40mg biw for 2 weeks; 60mg qw for 4 weeks). The primary endpoints were to find the recommended phase 2 dose (RP2D) and maximum tolerated dose (MTD). The secondary endpoint was ORR including complete remission (CR), partial remission (PR), marrow complete remission (mCR), and hematological improvement (HI). This trial was registered with the Chinese Clinical Trial Registry (#ChiCTR2200065167). Results: First we treated three AML cell lines: AML-OCI2, THP1, MV4-11 and three MDS cell lines: SKM-1, MOLM-13, MDSL with azacitidine and selinexor simultaneously or sequentially. Sequential treatment showed stronger synergetic effect (Figure 1A) and reduced colony formation by 90% in SKM-1 and MOLM-13 (Figure 1B). Cell cycle arrest were induced and the percentage of Annexin V cells obviously increased with sequential treatment (Figure 1C). Sequential treatment decreased leukemia burden and prolonged survival of MOLM-13-luciferase xenograft murine model (Figure 1D, E). Mechanistically, pretreatment of azacitidine on SKM-1 and MOLM-13 upregulated the expression level of p53, and selinexor led to massive nuclear accumulation of p53, resulted in better vulnerability lethal to MDS cells (Figure 1F). Secondly, 15 patients were enrolled in our trial with a median age of 57. 26.7% of patients were MDS-EB1, 73.3% were EB2. 93.3% of patients were high or very high risk by the Revised International Prognositic Scoring System (IPSS-R) (see the Table). At Phase Ib, the following selinexor dosages were administered: 40mg qw (n=3), 40mg biw (n=3) and 60mg qw (n=3). One patient in the 60mg qw group discontinued treatment because of nausea, vomiting, fatigue and Herpes simplex virus infection. Subsequently, in order to find RP2D, an extension cohort of 3 additional patients were treated with 40mg qw and 40mg biw, respectively. Therefore, 14 patients were evaluable for efficacy assessment. ORR was 78.6% in 11 patients including CR in 3 patients (21.4%); mCR+HI in 1 patient (7.1%); and mCR in 7 patients (50.0%) (see the swimmer plot). Especially, among the TP53 mutated patients (n=3), one achieved CR, the other two achieved mCR. The common adverse events were neutropenia (93.3%), anemia (86.7%), thrombocytopenia (80.0%), fatigue (53.3%), nausea (26.7%), dizziness (26.7%), vomiting (20.0%) and decreased appetite (20.0%). There were no treatment related deaths. Conclusion: We verified the activity of selinexor sequential azacytidine in MDS both in vitro and in vivo. The treatment showed with encouraging efficacy in newly diagnosed patients with MDS-EB1 or EB2. Adverse events were manageable with supportive care implementation. Enrollment is ongoing to assess the efficacy and safety of this sequential therapy, which may be a new treatment option for EB subjects.

Serum Amyloid A1 (SAA1) Secreted By the Stromal Microenvironment Drives Malignant Clonal Proliferation in Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML)

Drivers of malignant hematopoietic stem cell clonal proliferation from age-related clonal hematopoiesis (ARCH) to Myelodysplastic Syndromes (MDS) and Acute Myeloid Leukemia (AML) are not well understood. However, a pro-inflammatory and immune tolerant aging bone marrow (BM) microenvironment is thought to contribute to disease development. Serum Amyloid A1 (SAA1) has been identified as a novel pro-inflammatory oncoprotein the levels of which progressively increase with MDS and AML progression. SAA1 also selectively promotes growth of patient-derived MDS and AML cells independent of cytogenetic or mutational profile. We aimed to characterize the mechanism by which SAA1 drives oncogenesis within the microenvironment. Herein we further stratified the disease marking properties of SAA1 by clarifying the association of its BM levels with disease progression in MDS and AML. SAA1 levels were significantly elevated in BM aspirates from MDS and AML patients as compared to those from healthy age-matched subjects (p=0.0002 and p=0.0052, respectively). The effects of SAA1 on mutated hematopoietic cells were assessed in a model of age-related clonal hematopoiesis (ARCH). Utilizing a CRISPR-Cas9 system, ASXL1-mutant CD34+ hematopoietic stem cells (HSCs) were generated; the replating capacity of both mutated and healthy CD34+ cells were assessed in the presence or absence of SAA1. Notably, SAA1 increased the replating capability assessed by an increase in the number of colonies formed on serial replating of ASXL1-mutant CD34+ cells, but not those of SAA1-treated healthy CD34+ or vehicle treated ASXL1-mutated CD34+ cells, suggesting that the presence of SAA1 favors stemness of mutated HSCs, and implicating SAA1 as a driver of malignant clonality. To investigate the mechanism of action of SAA1 BM mononuclear cells from AML patients or healthy age-matched subjects were treated with SAA1 or vehicle overnight and single-cell RNA sequencing analysis was performed. Seurat and cell cluster annotation analysis showed that AML samples treated with SAA1 had increased numbers of hematopoietic stem and progenitor cells (HSPCs) and plasmacytoid dendritic cells as compared to matched untreated AML samples. Further, Enrichr analysis showed that pathways upregulated by SAA1 in AML samples included pro-inflammatory molecules, anti-apoptotic, and antiviral defense mechanisms (p<0.0001). Shared genes upregulated in these pathways in HSPCs include interferon-induced signals including JAK2, STAT1, and OAS3, and genes implicated in resistance to chemotherapy, including CD44 and CD36, as well as the anti-phagocytosis signal CD47. Lastly, we initiated the development of a blocking monoclonal antibody against SAA1 as a means of inhibiting its potent oncogenic effects on malignant HSCs and examining its potential therapeutic activity. Utilizing a RAW 264.7 NFκB-luciferase reporter cell line, we screened 51 clones for their ability to selectively block SAA1-mediated NFκB activation. 7 optimal candidates were identified which demonstrated dose-dependent reduction in NFκB activity upon SAA1 challenge. Importantly, these candidate clones were specific for SAA1 and did not affect NFκB activation mediated by lipopolysaccharide (LPS)-mediated activation. We observed up to an 85% reduction of SAA1-mediated activity using supernatants from further subcloning of previously selected candidates. Moreover, cell proliferation of a human AML cell line (OCI-AML3) stimulated by SAA1 was markedly reduced in the presence of candidate antibodies, suggesting anti-proliferative activity. Our findings show that SAA1 levels correlates with disease severity in MDS and AML progression, selectively promotes stemness and proliferative potential of ASXL1-mutant cells, and demonstrates unique immune-related mechanisms selecting for malignant clonality. Further development and purification of a novel inhibitory antibody against SAA1 for future in vivo and ex vivo testing is underway as a promising novel target to reduce SAA1-mediated, niche-driven effects with broad implications in the treatment of hematological malignancies.

Prevalence of Second Cancers in Patients with Polycythemia Vera (PV): A Retrospective Analysis of US Real-World Claims Data

Background:Polycythemia vera (PV) is a chronic myeloproliferative neoplasm (MPN) associated with risk of leukemic or fibrotic progression (Tefferi and Barbui. Am J Hematol. 2017;92;94-108). In addition to the known risk of disease transformation, patients with MPNs have a 60% higher risk of developing second non-hematologic cancers compared to matched controls. Skin cancers are among the most prevalent second cancers, with a reported 2.8-fold increase in risk of non-melanoma skin cancer (NMSC) (Landtblom et al., Leukemia. 2018;32:2203-10). The inherent risk associated with MPNs is exacerbated by cytoreductive treatments, as patients treated with hydroxyurea (HU) were found to have a two-fold higher risk of NMSC compared to patients who were not treated with HU (Barbui et al., Leukemia. 2019;33:1996-2005). Previous studies examined fewer than 2000 patients; the study populations may have included patients with other MPNs and PV. Our objective was to evaluate the frequency of second cancers in patients with a primary PV diagnosis who were treated with HU compared to phlebotomy only using a large US real-world claims database. Methods:This real-world, retrospective cohort study used Optum's deidentified Market Clarity Data consisting of electronic health records (EHR) on 105 million US patients and linked medical and prescription claims between 1/1/2007 to 12/31/2019. More than 82,000 patients with ≥1 PV diagnosis at any time were identified within the extracted dataset. For this analysis, eligible patients were ≥18 years old and required ≥2 PV diagnosis codes with ≥60 days between the first and last diagnosis code in the data period. Patients were indexed at the first diagnosis code of PV preceded by ≥1 year of EHR data (“index”) and required ≥1 year of post-index data or death prior to data period end. Patients with a pre-index diagnosis of secondary polycythemia, leukemia, myelodysplastic syndrome, myelofibrosis, or stem cell transplant were excluded. Second cancers were identified at first occurrence of a diagnosis code for a defined cancer in EHR during specified time periods. Analysis of post-treatment cancer rates used a subset of patients with pre-index medication data and excluded cancer types identified prior to treatment. Results:The total study cohort included 20,089 patients with PV (mean age, 63.0 years; 57.8% male; 88.6% Caucasian). Some patients (15.5%; n=3112) had evidence of a pre-index cancer. Median duration of follow-up was 4.3 years (IQR, 2.4-5.9). Among the total cohort, 35.7% (7181 patients) reported at least one second cancer in the post-index period (“post-index period prevalence”). The rate of second cancers in the post-index period was 109.7 events per 1000 patient years (PY). The most common non-hematologic malignancies were skin cancers (22.1/1000 PY) followed by prostate cancer (8.1/1000 PY) and breast cancer (6.7/1000 PY) (Figure 1). Within the total cohort, 9.1% (1830 patients) reported at least one skin cancer in the post-index period, with 8.3% (1659 patients) experiencing NMSC and 1.4% (286 patients) experiencing melanoma (Figure 1). The post-index period prevalence rate of NMSC was 6 times higher vs. the rate of melanoma (19.8 vs. 3.3/1000 PY). Among NMSCs, the rate of basal cell carcinoma was higher than the rate of squamous cell carcinoma (8.7 vs. 6.9/1000 PY). However, the highest rate was observed for non-specific or unspecified malignant neoplasms (8.9/1000 PY). Within a subset of 17,402 patients with pre-index medication data, 20.2% (n=3512) were exposed to at least 4 weeks of HU (without ruxolitinib) at any time; 19.0% (n=3314) were treated with phlebotomy (PHL) only. The rate of any second malignancy subsequent to treatment was >2.5-times higher in the HU vs. PHL group (140.4 vs. 55.3/1000 PY) (Table 1). Rates of skin cancers were nearly two times higher in the HU vs. PHL group for any skin cancer (31.6 vs. 17.7/1000 PY), NMSC (28.9 vs. 15.7/1000 PY), and melanoma (4.5 vs. 2.9/1000 PY). Conclusion:To our knowledge, this is the largest retrospective study evaluating the frequency of second cancers in a cohort of US patients with PV to-date. We found that patients with PV had high rates of malignancies associated with their primary PV diagnosis. The highest rates were observed for skin cancers, driven by the frequency of NMSC. Patients treated with HU had nearly twice the rate of skin cancers (including NMSC and melanoma) compared to PHL-only treated patients.

Real-World Analysis of Thromboembolic Event Rates in Patients in the United States with Polycythemia Vera

Background:Polycythemia vera (PV) is a myeloproliferative neoplasm associated with an increased risk of thromboembolic events (TEs). PV patients are characterized based on their risk for TEs as low-risk (age <60 without prior history of TE) or high-risk (age ≥60 and/or prior history of TE). Observational studies have shown higher rates of TEs in PV patients compared to matched controls (14.3 vs 4.9/1000 patient years, or PY), with rates as high as 40% in patients with PV (Goyal et al., Blood. 2014;124:4840; Griesshammer et al., Ann Hematol. 2019;98:1071-82; Pemmaraju et al., Leuk Res. 2022;115:106809). We assessed the rate of arterial and venous TEs in a large cohort of low- and high-risk PV patients. Methods:We conducted a real-world, retrospective cohort study using Optum's de-identified Market Clarity Data, which includes electronic health records (EHR) on 105 million US patients between 2007-2019. Eligible patients were ≥18 years [yrs] old and required ≥2 PV diagnosis codes with ≥60 days between the first and last diagnosis code in the data period. Patients were indexed at the first diagnosis code of PV preceded by ≥1 year of EHR data (“index”) and required ≥1 year of post-index data or death prior to data period end (12/31/2019). Patients with a pre-index diagnosis of secondary polycythemia, leukemia, myelodysplastic syndrome, myelofibrosis, or stem cell transplant were excluded. Patients were stratified into three groups based on risk status at index: low-risk (age <60 without prior history of TE), age-based high-risk (age >60 without prior history of TE) and event-based high-risk (prior history of TE). As outcomes, TEs were identified at the first occurrence of a diagnosis code for a TE in EHR during the post-index period. Descriptive statistics were used to characterize the proportion of patients with TEs and event rates per 1000 PY. Kaplan-Meier analysis was used to evaluate the probability of TE-free survival among patients in the three risk groups at various time points. Patients were censored at death or last engagement date. Results:The total study cohort included 20,089 PV patients (mean age, 63.0 yrs; 57.8% male; 88.6% Caucasian). Approximately one-third (34.4%; 6909 patients) were low-risk, 49.4% (9924 patients) were age-based high-risk, and 16.2% (3256 patients) were event-based high-risk. Among 17,402 patients with pre-index medication data, 16.0% (n=2777) received hydroxyurea (HU) pre-index. HU usage was highest in age-based high-risk patients (21.8%) followed by event-based high-risk (20.0%) and low-risk patients (5.8%). Median follow-up period was 4.3 yrs (IQR, 2.4-5.9). Incidence of TEs was assessed within patient groups (Table 1). Overall, 25.1% (5035/20,089) of patients with PV experienced at least one TE during the post-index period. Incidence of TEs was highest among event-based high-risk patients (50.2%; 1634/3256), followed by age-based high-risk (25.0%; 2480/9924) and low-risk patients (13.3%; 921/6909). The rate of post-index TEs was more than triple for event-based high-risk patients vs. age-based high-risk patients (206 vs. 68/1000 PY) and nearly 7 times higher vs. low-risk patients (206 vs. 31/1000 PY). The rate of TEs was more than double for age-based high-risk patients vs. low-risk patients (68 vs. 31/1000 PY). Incidence of arterial and venous TEs was assessed across the total cohort (Table 1). The most common arterial events were stroke (7.1%) and MI/ACS (6.4%). The most common venous events were DVT/deep thrombophlebitis (8.1%) and pulmonary embolism (4.5%). Kaplan-Meier analysis indicated that at ≈2.5 yrs (1000 days) post-index, TE-free survival was ≈90% for low-risk, ≈80% for age-based high-risk, and ≈55% for event-based high-risk patients (Figure 1). At ≈5.5 yrs (2000 days) post-index, TE-free survival had fallen to ≈85% for low-risk, ≈70% for age-based high-risk, and ≈40% for event-based high-risk patients. The downward trend continued until the end of the data period. Conclusion: Findings from this real-world analysis indicate that PV patients experience high rates of arterial and venous TEs. Event-based high-risk patients have a substantially lower probability of TE-free survival compared with age-based high-risk and low-risk patients. While patients with a prior history of TE have the highest thromboembolic risk, all patients with PV are at high risk of TE, indicating an unmet need for reduction of thromboembolic risks.

Prevalence and Outcome of Vexas Syndrome in Unrelated Hematopoietic Stem Cell Transplantation

Introduction Vacuoles, E1 enzyme, X-linked, Autoinflammatory, Somatic (VEXAS) syndrome is a newly identified monogenic disorder defined by somatic UBA1 mutations in hematopoietic progenitor cells, resulting in bone marrow failure (BMF) and systemic inflammation. This syndrome is frequently associated with myelodysplastic syndrome (MDS) or plasma cell dyscrasia. Although high-dose corticosteroids alleviate symptoms and cytopenia, allogeneic hematopoietic stem cell transplantation (HSCT) has been suggested as a potentially curative treatment. This study evaluated the safety and efficacy of unrelated HSCT for treating VEXAS syndrome and its prevalence in patients with hematologic diseases. Methods We conducted retrospective screening for UBA1 mutations in patients with BMF or plasma cell neoplasms who underwent their first HSCT from unrelated donors in Japan between 1995 and 2020. Pre-HSCT blood DNA samples and clinical data were collected from the Japanese Data Center for Hematopoietic Cell Transplantation. Multiplex real-time PCR was used to detect the most common pathogenic UBA1 mutations, including p.M41L (c.121A>C), p.M41V (c.121A>G), and p.M41T (c.122T>C), with detection limits of 0.13%, 0.5%, and 1%, respectively. The presence of these mutations was confirmed using Sanger sequencing and droplet digital PCR. In addition, we examined UBA1 mutations in 2 patients previously diagnosed with MDS but suspected of having VEXAS syndrome based on specific clinical features. Results Among 4340 patients with either BMF or plasma cell neoplasms who underwent unrelated HSCT, 1771 had available pre-HSCT blood DNA samples. The diagnoses of the 1771 patients included MDS (n = 1264), acquired aplastic anemia (n = 375), inherited BMF syndromes (n = 89), plasma cell neoplasms (n = 23), and other acquired BMF syndromes (n = 20). The median patient age at HSCT was 46 (range, 0-75; interquartile range, 20-59) years old, and the male-to-female ratio was 1.6:1. Pathogenic UBA1 mutations were detected in 2 of the 1771 patients (0.11%), both of whom were male MDS patients older than the median age (cases 1 and 2); thus, the prevalence of VEXAS syndrome within this patient subgroup was 0.36%. Both patients received chemotherapy before unrelated donor bone marrow transplantation (BMT), including induction chemotherapy and azacitidine, and achieved complete and partial responses, respectively. In addition, UBA1 mutations were detected in the 2 MDS patients at Kanazawa University Hospital (cases 3 and 4). Case 3 also underwent unrelated BMT after experiencing cytopenia relapse and inflammatory symptoms during corticosteroid reduction. The clinical characteristics and outcomes of the 4 patients with VEXAS syndrome are summarized in Table 1. Variant allele frequencies (VAFs) of UBA1 mutations ranged from 2% to 71%. The 3 patients who underwent unrelated BMT received fludarabine and busulfan-based conditioning regimens and graft-versus-graft disease (GVHD) prophylaxis with tacrolimus + methotrexate or cyclosporine + methotrexate. All 3 patients achieved neutrophil and platelet engraftment. One patient required systemic corticosteroids for acute skin GVHD. One patient died of idiopathic interstitial pneumonia 5 months after BMT. The other 2 patients were alive without relapse, chronic GVHD, or other complications at 18 and 28 months after BMT. Discussion Our results support the curative potential of allogeneic HSCT for VEXAS syndrome without excessive toxicity. The occurrence of transplant-related mortality due to idiopathic interstitial pneumonia may be related to the reported poor outcome of the p.M41V mutation following non-transplant therapy. The low prevalence of VEXAS syndrome in the current cohort may be due to the late age of onset and the rare progression to advanced MDS. The low VAFs of UBA1 mutations in the 2 patients who responded to chemotherapy before HSCT may indicate the efficacy of chemotherapy in selectively eradicating UBA1 mutant clones. Our rapid, sensitive, and cost-effective screening method for VEXAS syndrome can help optimize treatment strategies for this rare disease in future studies.

Selenium Deficiency and Chronic Anemia in an Outpatient Hematology Clinic

Background Selenium (Se) is an essential micronutrient with roles in reproduction, metabolism, immunity, and the cellular cycle, including DNA synthesis and prevention of cell death. Its role in malignancy has also been studied, specifically for esophageal, gastric, lung, prostate cancer. Studies have shown benefit with Se supplementation for those malignancies. There is evidence that low Se levels are independently associated with worse outcomes regarding treatment response and long-term survival for Non-Hodgkin's Lymphoma (NHL) and cutaneous T-cell lymphoma, and additional studies on its association in acute myeloid leukemia (AML), follicular lymphoma (FL), and Hodgkin's lymphoma (HL) have revealed worsening outcomes as well. However, Se was not predicative of treatment response or survival for AML, FL and HL, therefore it's role in the development of hematological malignancies is not clear. Previously, the role of Se in myelodysplastic syndrome (MDS) was investigated, indicating the importance of Se for apoptosis in MDS cell lines. There is evidence that a causal relationship with decreased Se levels and chronic anemia might exist; however, there is a paucity of data regarding its role in the development of chronic anemia and hematological disorders. This descriptive single center study describes Se deficiency in patients with chronic anemia. Objectives Primary endpoint was to determine if there was evidence of Se deficiency in patients with chronic anemia in an academic hematology outpatient clinic. Methods We herein present a descriptive study on 18 patients with hematological disorders and selenium deficiency. Upon initial detection of Se deficiency, levels of hemoglobin (Hb), mean corpuscular volume (MCV), Copper (Cu), Zinc (Zn) and iron (Fe) closest to time of detection were noted if tested. Average values with standard deviation were calculated and association with malignancies were noted. Results Of the 18 patients, 13 had pre-malignant or malignant conditions, including 7/13 who had bone marrow failure/MDS, 5/13 patients with lymphoid disorders, and 1/13 with plasma cell disorders. The 5 remaining patients had chronic gastrointestinal disorders. Averages on detection of Se deficiency were noted to be: Hb (9.9 ± 2.0), MCV (96.0 ± 11.2), Se (100.5 ± 11.6), Cu (95.8 ± 28.8), Zn (70.1 ± 21.5) and Fe (81.6 ± 56.7). No association was noted between Se and other trace elements. 72% (13/18) of patients with Se deficiency also had malignant conditions. Of those with malignancy, 54% were noted to have bone marrow failure. Discussion Currently there are no clinical guidelines that recommend following levels of Se routinely in patients with chronic anemia. Our data might indicate a possible association between chronic anemia and Se deficiency. Given the high number of MDS patients in our study, there might be a link between Se deficiency and MDS. Further investigation is warranted into the role of Se in the development of chronic anemia disorders, as supplementation with Se and routine follow-up may be warranted and can improve clinical outcomes.

Aberrant Splicing of MBD1 Reshapes the Epigenome to Drive Convergent Myeloerythroid Defects in MDS

Splicing defects are a characteristic feature of myelodysplastic syndromes (MDS) and typically associate with specific recurrent splicing factor mutations. However, a subset of transcripts exhibit abnormal splicing regardless of mutational background, occurring even in the absence of splicing-related mutations. These shared splicing events likely include common underlying drivers of MDS hematopoietic defects, yet the functions of the resulting transcripts remain unknown. We identified a long isoform of Methyl-CpG-Binding Domain 1 ( MBD1) as the product of one such mutation-independent splicing event. To understand whether altered MBD1 splicing contributes to hematopoietic dysfunction, we overexpressed isoforms of MBD1 in cord blood CD34+ cells and found that the MDS-associated full-length isoform (MBD1-L), containing MBD1's 3rd CXXC domain, impaired erythroid differentiation by stalling cell cycling and promoting apoptosis. In contrast, the MBD1-ΔCXXC3 isoform (MBD1-S), preferentially produced in healthy cells, did not induce these defects. Recapitulating these findings, the MBD1-L isoform uniquely impaired reconstitution capacity in vivo, particularly in the erythroid and myeloid lineages, and in addition produced an enrichment of the MDS transcriptomic signature on RNA-seq profiling. The unique CXXC3 domain of MBD1-L specifically binds non-methylated CpGs, and exhibits greater target affinity than the shared MBD domain, which is responsible for mCpG binding. Given MBD1's key role in heterochromatin maintenance at mCpG regions, we hypothesized that the unique function of MBD1-L in MDS may be attributable to the refocusing of MBD1-mediated epigenetic repression from canonical, methylated DNA sites to unmethylated sites. Isoform-specific CUT&RUN and multi-omics profiling in cord blood CD34+ cells revealed that the inclusion of the CXXC3 exon triggers a striking redistribution of MBD1 from gene bodies and intergenic regions to hypomethylated promoter CpG islands, resulting in widespread suppression of promoter chromatin accessibility and downregulation of cell-cycle-related transcripts. Characterization of the MBD1 interactome by rapid immunoprecipitation mass spectrometry showed that the MBD1-L isoform preferentially associates with the SETDB1:ATF7IP H3K9 methylator complex, supporting active heterochromatin establishment at MBD1-bound promoters. Among the direct targets uniquely repressed by MBD1-L is BCOR, a recurrent LOF gene in MDS whose loss perturbs hematopoietic differentiation and promotes self-renewal. Downregulation of BCOR by MBD1-L led to the derepression of BCOR-controlled genes, contributing to an enriched stem cell-associated transcriptomic signature resembling BCOR LOF. To investigate whether reversal of MBD1-L splicing can restore hematopoietic output in diseased cells, we delivered lipid-encapsulated splice-switching antisense oligonucleotides targeting the CXXC3 exon into primary human MDS cells, and observed an increase in differentiation in vitro. In addition, depletion of MBD1-L in the MDSL cell line using isoform-specific shRNAs increased cell proliferation, confirming that targeted reduction of MBD1-L inverted the quiescent, differentiation-impaired phenotype imposed by its overexpression. Our results demonstrate that MBD1 isoforms act on separate compartments of genomic sequence to carry out divergent functions in hematopoietic cells, and that disease-associated overproduction of MBD1-L compromises hematopoietic output and lineage differentiation. These findings provide the first evidence that mutation-independent splicing changes can drive hematopoietic dysfunction in MDS.

Selinexor in Combination with Azacitidine (AZA) in Patients with Myelodysplastic Syndrome (MDS): A Real-World Retrospective Study

Background: Patients with higher-risk MDS face limited treatment options and poor outcomes. While HMAs are approved as frontline therapy for higher-risk MDS, primary failure occurs in 50% of HMA-treated patients, and most responders progress within 2 years. There is an unmet need to improve the efficacy of AZA frontline therapy while maintaining a tolerable safety profile. Preclinical studies have demonstrated that the combination of selinexor and azacitidine synergistically inhibits MDS cell proliferation and arrests the cell cycle at the G2/M phase (Yixuan Guo, 2022). The combination of selinexor and azacitidine may be a promising approach for treating MDS. Here, we present this single-center, real-world evidence supporting the use of selinexor in combination with AZA in MDS patients. Methods: Medical records of 19 patients with MDS receiving selinexor in combination with AZA at the First Affiliated Hospital of Soochow University between Jan 2022 and March 2023 were reviewed. Inclusion criteria were a pathologically confirmed diagnosis of MDS, and receipt of ≥1 cycle of azacitidine in combination with selinexor. HMA-naïve patients and those previously treated with HMAs were included. Patients with a diagnosis other than MDS or progression to AML before treatment with selinexor + AZA were excluded. Patients received selinexor 40-60 mg QW orally and AZA 75 mg/m 2 intravenously or subcutaneously on days 1-5 or 7 of each 28-day cycle. Responses were evaluated per the International Working Group criteria. Results: A total of 19 pts (median age, 54 y [range, 13-68 y]) were treated (Table 1). IPSS-R risk was high or very high in 15.8% and 68.4% of pts, respectively. IPSS-M risk was high or very high in 10.5% and 78.9% of pts, respectively. 57.9% of pts were previously treated with HMA. 63.2% had myelofibrosis, 42.1% had hepatosplenomegaly, 31.6% had a TP53 mutation, and 63.2% had poor or very poor-risk cytogenetics. As of Jun 10, 2023 at a median follow-up of 5.9 months (range 1.5-17.4 months), eight pts died (42.1%) due to disease progression. For the total cohort, the overall response rate (ORR) was 73%. In pts with worse survival prognosis, such as relapse and refractory, high and very high in IPSS-R, high and very high in IPSS-M, myelofibrosis and TP53 mut, the ORR was 64%, 81%, 76%, 75% and 83%, respectively (Figure 1). In seven pts with splenomegaly, six pts had reduced spleen volume, and one pt had no significant change by abdominal palpation or imaging examination. The 6-month overall survival (OS) was 74.7% in total pts, and 70.0% in pts previously exposed to HMA (Figure 2a, 2b). Selinexor + AZA therapy improved survival of patients with poor prognosis, such as TP53 mut (0.099), MF (+) (0.613), ≥high in IPSS-R (0.938) (Figure 2c, 2d, 2e). The survival of six pts who received transplantation after combination therapy was significantly prolonged, with 6-month OS was 83.3% (Figure 2f). Conclusion: Selinexor + AZA combination therapy was well tolerated with promising efficacy in patients with untreated or relapse/refractory MDS, including those with TP53 mutation, myelofibrosis, and higher risk in IPSS-R.

The bcl6 corepressor mutation regulates the progression and transformation of myelodysplastic syndromes by repressing the autophagy flux

The occurrence of autophagy dysregulation is vital in the development of myelodysplastic syndrome and its transformation to acute myeloid leukemia. However, the mechanisms are largely unknown. Here, we have investigated the mechanism of the bcl6 corepressor mutation in myelodysplastic syndrome development and its transformation to acute myeloid leukemia. We identified a novel pathway involving histone deacetylase 6 and forkhead box protein O1, which leads to autophagy defects following the bcl6 corepressor mutation. And this further causes apoptosis and cell cycle arrest. The bcl6 corepressor-mutation-repressed autophagy resulted in the accumulation of damaged mitochondria, DNA, and reactive oxygen species in myelodysplastic syndrome cells, which could then lead to genomic instability and spontaneous mutation. Our results suggest that the bcl6 corepressor inactivating mutations exert pro-carcinogenic effects through survival strike, which is only an intermediate process. These findings provide mechanistic insights into the role of the bcl6 corepressor gene in myelodysplastic syndrome.

Iron overload induces dysplastic erythropoiesis and features of myelodysplasia in Nrf2-deficient mice.

Iron overload (IOL) is hypothesized to contribute to dysplastic erythropoiesis. Several conditions, including myelodysplastic syndrome, thalassemia and sickle cell anemia, are characterized by ineffective erythropoiesis and IOL. Iron is pro-oxidant and may participate in the pathophysiology of these conditions by increasing genomic instability and altering the microenvironment. There is, however, lack of in vivo evidence demonstrating a role of IOL and oxidative damage in dysplastic erythropoiesis. NRF2 transcription factor is the master regulator of antioxidant defenses, playing a crucial role in the cellular response to IOL in the liver. Here, we crossed Nrf2-/- with hemochromatosis (Hfe-/-) or hepcidin-null (Hamp1-/-) mice. Double-knockout mice developed features of ineffective erythropoiesis and myelodysplasia including macrocytic anemia, splenomegaly, and accumulation of immature dysplastic bone marrow (BM) cells. BM cells from Nrf2/Hamp1-/- mice showed increased in vitro clonogenic potential and, upon serial transplantation, recipients disclosed cytopenias, despite normal engraftment, suggesting defective differentiation. Unstimulated karyotype analysis showed increased chromosome instability and aneuploidy in Nrf2/Hamp1-/- BM cells. In HFE-related hemochromatosis patients, NRF2 promoter SNP rs35652124 genotype TT (predicted to decrease NRF2 expression) associated with increased MCV, consistent with erythroid dysplasia. Our results suggest that IOL induces ineffective erythropoiesis and dysplastic hematologic features through oxidative damage in Nrf2-deficient cells.