Acute Myeloid Leukemia Transformation Research Articles

Myelodysplastic Syndrome (MDS) in a Child who Transformed to Acute Myeloid Leukaemia with Myelodysplasia-Related Changes (AML-MRC)

Background: Myelodysplastic syndromes (MDS) are a group of clonal hematopoietic stem cell diseases, accounting for < 5% of all hematologic malignancies in children.  MDS is characterized by cytopenia, dysplasia in one or more of the major myeloid lineages, ineffective hematopoiesis, and recurrent genetic abnormalities. Approximately 30% of patients with MDS show progression to AML within a few months up to several years. This is an MDS case report in children who have undergone Acute Myeloid Leukaemia transformation with Myelodysplasia-Related Changes (AML-MRC).Case Report: A 16-year-old boy came to the Emergency Department of Sanglah General Hospital with a complaint of pain and swelling in both knees and joints since last month. Fatigue and fever happened a week before admission. Previously, he had been hospitalized twice with the same symptoms. Physical examination revealed pale conjunctiva with pancytopenia in the laboratory result. A blood smear showed normochromic normocytic anemia, neutropenia, and thrombocytopenia. Bone marrow aspiration morphology showed hypercellularity with blast presentation > 10% and multilineage dysplasia > 50% supporting MDS diagnosed transformation to Acute Myeloid Leukaemia with Myelodysplasia-Related Changes (AML-MRC). Leukemia phenotyping showed positive results on CD 33, CD 36 and HLA-DR presenting Monocytic Lineage.                                                                                                            Conclusion: High degree dysplasia and blast percentage in blood and bone marrow increase tendency to transform into acute myeloid leukemia (AML). The diagnose of MDS requires analysis of blood and bone marrow morphology and cytogenetic analysis as a recommendation by WHO classification.

PF551 CD10 EXPRESSION ON MYELOID PRECURSOR CELLS (CD34 + CD117 +) AS A PREDICTIVE FACTOR FOR PROGRESSION OF PRIMARY MYELODISPLASTIC SYNDROMES

Background: Prognosis assessment of the course of the disease is a clinically important stage in the initial diagnosis of myelodysplastic syndrome (MDS). Analysis of bone marrow cells by flow cytometry allows to identify specific aberrations of both immature and mature cells of various hematopoietic lines. Considering the fact that the definition of MDS includes a diverse group of myeloid neoplasms, the presence of multiple aberrations may be important for diagnostics and risk stratification in patients with MDS. Thus, the assessment of specific immunophenotypic aberrations in MDS blast cells can be a practical tool for predicting the course of the disease and the risk of transformation into acute myeloid leukemia (AML). Aims: To identify the immunophenotypic predictors of AML transformation in myeloid progenitor cells (CD34 + CD117 +) in the bone marrow of adults with primary MDS. Methods: The study included 116 adult patients at the State Medical Center «Minsk Scientific-Practical Center of Surgery, Transplantology and Hematology» from February 2012 to December 2017 with primary MDS diagnosis. Immunophenotyping was performed before the start of a specific MDS treatment. The median age was 59.1 (interq. range 29:79), among them 62 were males (53%) (median of age 58.7 (interq. range 29.2:78.4) and 54 were females (47%) (median of age 59.1 (interq. range 38:79). Expression of antigens in the bone marrow blast cells was determined using eight-color flow cytometry on a FACSCanto II instrument (Becton Dickinson, USA). The cytogenetics was performed by a standard G-banding. Results: A comparative analysis of the time from the initial diagnosis to the disease transformation into AML was performed based on the pattern of expression of immunophenotypic markers on a subpopulation of CD34 + CD117 + myeloid progenitor cells. A marker of asynchronous expression of CD10 was shown as the most significant predictor of disease transformation into AML, and its’ expression does not depend on the MDS variant, according to the WHO classification, and on the presence of cytogenetic aberrations determining unfavorable prognosis, such as anomalies of chromosome 7 and complex karyotype. The presence of aberrant expression of CD10 ≥ 10% on myeloid progenitor cells (CD34 + CD117 +) in MDS patients was found to be associated with a rapid progression of the disease and significantly increased the risk (p = 0.0023) of transformation into AML. It was shown that in the group of patients, who had the expression of CD10 on their bone marrow blast cells, the median time to transformation into AML was 6 months, which is significantly (p < 0.001) shorter than in the group of patients where this aberration was absent, where the median time to transformation into AML was 48 months (Figure 1).Summary/Conclusion: The presence of aberrant expression of CD10 on myeloid progenitor cells (CD34 + CD117 +) is an immunophenotypic predictor of the increased risk of rapid transformation into AML in adult patients with MDS.

Myelodysplastic syndrome from theoretical review to clinical application view.

Myelodysplastic syndromes (MDS), called ineffective hematopoiesis is indicated by bone marrow failure and tendency to acute myeloid leukemia transformation. Since the disease is more common in elderly with non- hematology co-morbidities, the research for less toxic and curative novel agents is essential. More than 12 years without new Food and Drug Administration approved drugs in MDS management through the whole course, only 5 drugs. We summarized the basic data in diagnosis, treatment guidelines and future direction.

Prognostic Impact of Activating Mutations of the NRAS/KRAS Genes in Patients with Primary and Post-Essential Thrombocythemia (ET)/Polycythemia Vera (PV) Myelofibrosis (MF)

Introduction: MF is a Philadelphia-negative myeloproliferative neoplasm (Ph-negative MPN) associated with driver mutations in the JAK-STAT pathway (e.g. JAK2, CALR, MPL) and other mutations in genes that lead to epigenetic changes and altered RNA splicing (e.g. TET2, SRSF2, ASXL1, EZH2). The RAS-signaling pathway is frequently altered in acute myeloid leukemia (AML) and other myeloid malignancies, but few studies have evaluated the prevalence of such mutations in patients with MF. We sought to describe the frequency and clinical features of RAS mutations in patients with MF.Methods: We analyzed next-generation sequencing data from 723 patients with a diagnosis of primary MF (N=520), post-PV MF (N=119) and post-ET MF (N=84). Sequencing was performed with either paired tumor-normal whole exome sequencing (WES; N=56) or selected gene panel for genes associated with myeloid malignancies (N=667). The following 16 genes were analyzed in all 723 patients and were considered as the common denominator for analysis: ASXL1, CALR, DNMT3A, EZH2, FLT3, IDH1, IDH2, JAK2, KIT, KRAS, MPL, NRAS, RUNX1, TET2, TP53, WT1. RAS mutations were considered as oncogenic mutations in NRAS and/or KRAS. Molecular high risk (MHR) mutations were considered as mutations in any one of the 4 genes: ASXL1, EZH2, IDH1, IDH2 (SRSF2 mutations were not included since they were not evaluated in all cases). Odds ratio (OR) and P-values were estimated using Fisher's exact test in pairwise comparisons among genetic features, and P-values were adjusted for multiple comparisons using the Benjamini-Hochberg procedure, with significant Q-values considered as those <0.15. Overall survival (OS) was estimated using the Kaplan-Meier method, and defined as the time from the date of sample collection for mutation analysis until death from any cause, with patients alive at last follow-up censored. Cumulative incidence of AML transformation (CI-AML) was defined from the time of mutation analysis until evolution to AML, considering death without AML transformation as a competing risk. A Cox proportional hazards model was fitted to determine the hazard ratio (HR) of independent covariates associated with the dependent variable OS.Results: Mutations in RAS genes were found in 44 patients (6.1%; 95% confidence interval [CI] 4.5-8.1%). There were 32 cases with NRAS mutations (4.4%; 95% CI 3-6.2%), 15 cases with KRAS mutations (2%; 95% CI 1.2-3.4%), and there were 3 cases with mutations in both genes (0.4%; 95% CI 0.1-1.3%). JAK-STAT driver genes found in the cohort included JAK2 (N=486), CALR (N=94), MPL (N=35) and Triple-negative status (N=108), and there was no difference in the distribution of JAK-STAT driver genes among patients with NRAS/KRAS mutations (p=0.96). Patients with NRAS/KRAS mutations had a higher number of non-driver mutations (median 3 vs 1; p=8.5e-20), higher white blood cell counts (median 15.1x109/L vs 10.5x109/L, p=0.02) and more frequently harbored ASXL1 mutations (OR 3.00, q=0.01), EZH2 mutations (OR 3.40, q=0.11) and MHR mutations (OR 3.13, q=0.004). There was a negative association of NRAS/KRAS mutations with del(20q) changes in karyotype (OR <0.001, q=0.06). After a median follow-up of 30 months and 439 events, OS was significantly reduced in patients harboring NRAS/KRAS mutations (median 19.5 months vs. 44.6 months, HR 2.95, p-value=5.99e-06; figure 1). The presence of NRAS/KRAS mutations was also associated with a higher CI-AML (4 years 18.4% vs. 14%, p-value=0.03). In a multivariate Cox model, NRAS/KRAS mutations were associated with worse OS after adjusting for other prognostic variables in MF (Table 1), including Dynamic International Prognostic Score System (DIPSS), karyotype and presence of MHR mutations. In a subset analysis of 396 patients with Int-2/High risk DIPSS, there were 202 patients who were treated with ruxolitinib. NRAS/KRAS mutated patients who received ruxolitinib (N=24) had superior OS than patients who did not receive the drug, but still inferior when compared to patients with wild-type NRAS/KRAS (Figure 2).Conclusion: Patients with a diagnosis of MF who harbor NRAS/KRAS mutations comprise a high-risk subgroup with poor outcomes. NRAS/KRAS mutational status should be evaluated in future prognostic models in MF. The efficacy of JAK2 inhibitors needs to be further studied in this subgroup of patients, as well as the possibility of targeting the RAS pathway with MEK inhibitors. [Display omitted] DisclosuresRampal:Incyte: Honoraria, Research Funding; Jazz: Consultancy, Honoraria; Celgene: Honoraria; Stemline: Research Funding; Constellation: Research Funding. Verstovsek:Italfarmaco: Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy; Celgene: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau.

Pan-Myeloid Leukemia Analysis: Machine Learning-Based Approach to Predict Phenotype and Clinical Outcomes Using Mutation Data

Pan-Myeloid Leukemia Analysis: Machine Learning-Based Approach to Predict Phenotype and Clinical Outcomes Using Mutation Data

A Novel DNA Methyltransferase Inhibitor Is Effective in an In Vivo Model of Myelodysplastic Syndrome

The myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell disorders characterized by ineffective hematopoiesis, peripheral blood cytopenias, dysplasia and a propensity for transformation to acute myeloid leukemia (AML). MDS is frequently associated with epigenetic gene silencing via methylation of cytosine residues in gene regulatory regions, and DNA methyl-transferase 1 (DNMT1) inhibitors, such as 5'azacytidine and 5-aza-2'-deoxycytidine (decitabine, DAC), are two of the three agents that are FDA approved for treatment of MDS. Although these drugs are not curative, they induce hematological improvement or improved survival in a significant fraction of MDS patients. Two novel, thiol-substituted 2'-deoxycytidine (dCyd) analogs designated T-dCyd (4'-thio-2'-deoxycytidine) and Aza-T-dCyd (5-aza-4'-thio-2'-deoxycytidine) were synthesized and shown to be potent DNMT1 inhibitors in vitro. We evaluated these drugs in vivo using the NUP98-HOXD13 (NHD13) mouse model for MDS.To mimic human MDS hematopoiesis, in which a portion of the hematopoietic output is provided by the MDS clone, and a portion provided by normal, non-MDS cells, we transplanted wild-type (WT) mice with a mixture of WT murine hematopoietic cells and NHD13 (MDS) hematopoietic cells. This bone marrow transplant (BMT) produces chimaeric recipients with bone marrow comprised of hematopoietic cells derived from both the MDS clone as well as normal hematopoietic precursors. WT and MDS cells in the mice can be distinguished by differential CD45 alleles (CD45.1 and CD45.2, respectively), which enables analysis and purification of the MDS and WT cells; this feat is not easily achieved with human MDS patient samples, which lack cell surface antigens specific for the MDS clone. At 8 weeks post-transplant; engraftment of MDS cells was documented by the presence of CD45.2+ cells in the peripheral blood, and the starting CBCs showed signs consistent with MDS including peripheral blood cytopenia and macrocytosis. Mice were randomly assigned to one of the three groups. 1) PBS, 2) T-dCyd, 3) Aza-T-dCyd. T-Cyd was dosed at 4 mg/kg/d intraperitoneally (IP) on weekdays for 2 weeks (10 doses), followed by three weeks rest; this constituted one cycle of therapy. Aza-T-dCyd was administered on the same schedule at 4 mg/kg/d IP. Flow cytometry and CBC were assessed on day 21 of each cycle, and treatment continued for up to one year, or until mice were humanely euthanized due to tachypnea, lethargy, or other signs of AML. Between four and six mice were treated per group, and the entire experiment was repeated three times and results pooled for T-dCyd, once for Aza-T-dCyd.The T-dCyd treated chimaeric mice showed significantly enhanced overall survival associated with hematological improvement including hemoglobin concentration, platelet and absolute neutrophil count compared to PBS treated mice (median survival 45.4 vs 28 weeks, p=0.0187). In addition to a survival advantage, AML onset was significantly delayed in the T-dCyd treated mice (median time to AML transformation 35 weeks for PBS vs unreached for T-dCyd, p=0.0111), although there was no significant change in MDS (CD45.2) engraftment between the T-dCyd and PBS treated mice. For Aza-T-dCyd group, we did not detect a survival benefit nor hematologic improvement, although we suspect this may have been secondary to unexpected toxicity at the selected dose. In sum, these results demonstrate the utility of chimaeric WT/MDS mice as a pre-clinical model for human MDS, and show that treatment with T-dCyd, a new DNMT1 inhibitor, leads to a survival advantage, hematologic improvement, and delayed transformation to AML. DisclosuresAplan:NIH Office of Technolgy Transfer: Employment, Patents & Royalties: NUP98-HOXD13 mice.

A Leukemic Stem Cell Signature-Based Scoring System for Predicting Prognosis in Myelodysplastic Syndrome Patients

Leukemic stem cells (LSCs) possess biological properties shared with normal hematopoietic stem cells. They are responsible for chemoresistance and relapse in acute myeloid leukemia (AML). Although myelodysplastic syndrome (MDS) has traditionally been regarded as a “hematopoietic stem cell disorder”, the clinical and biological impact of LSCs on MDS patients are not well defined. To address this question, we used the Affymetrix HTA 2.0 microarray platform to profile 16 out of the 17 recently reported stemness genes (one of them, the ARHGAP22 gene was not included in our array) in our 160 adult primary MDS patients. The diagnoses were based on the 2016 World Health Organization (WHO) classification. Patients with antecedent chemotherapy or hematologic malignancies were excluded. Forty-one (25.6%) patients had MDS with single lineage dysplasia (MDS-SLD), 20 (12.5%) had MDS with ring sideroblasts (MDS-RS), 31 (19.4%) had MDS with multilineage dysplasia (MDS-MLD), 32 (20%) had MDS with excess blasts-1 (MDS-EB1) and 36 (22.5%), MDS-EB2. The risk distribution of the cohort was very-high risk, 15.3%; high risk, 21.3%; intermediate risk, 24.7%; low risk 34.7%; and very-low risk 4% according to the revised international prognosis scoring system (IPSS-R). We identified 4 genes (LAPTM4B, NGFRAP1, NYNRIN, and EMP1) whose expression levels were significantly correlated with overall survival (OS). An LSC4 score (0.731 x LAPTM4B - 1.259 x NGFRAP1 + 0.304 x NYNRIN + 0.231 x EMP1) was constructed based on the weighted sums derived from Cox regression analysis. Higher LSC4 scores were associated with higher IPSS-R scores, complex cytogenetics, and higher incidences of mutations of RUNX1, ASXL1, TP53, SRSF2 and ZRSR2. High-score patients had significantly higher 3-year AML transformation rate (36.3% vs 11.3%, P<0.001), shorter leukemia-free survival (LFS, median, 33.6 months vs not reached, P< 0.001, Figure 1A), and shorter OS (median, 15.9 months vs 77.3 months, P<0.001, Figure 1B). In subgroup analysis, LSC4 score could also well stratify MDS patients with normal karyotype into different risk groups (p<0.001). The prognostic significance of LSC4 scores for LFS, AML transformation rate, and OS still remained valid in both IPSS-R lower-risk (very low, low, and intermediate risk) subgroup (Figures 2A and 2B) and higher-risk subgroup (high and very high risk). In multivariate analysis, higher LSC4 score was an independent adverse risk factor for OS, irrespective of age, IPSS-R, and mutation status of ASXL1, TP53, EZH2, SRSF2, and ZRSR2 (Table). We also well validated the prognostic significance of the LSC4 scores in an independent cohort of 30 MDS patients (Figures 3A and 3B). Interestingly, patients with higher LSC4 scores had a better OS if they received allogeneic transplantation than those who did not. It implied that transplantation might ameliorate the poor survival impact of high LSC4 scores. In conclusion, through comprehensive analysis, we have created a simple and powerful LSC4 scoring system, which serves as an independent prognostic factor for OS and LFS in MDS patients. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

The Role of FLT3-ITD Mutation on de Novo MDS in Chinese Population

BackgroundFLT3 mutations have been well-studied in acute myeloid leukemia (AML), and the detection of the FLT3 gene has become a clinical routine. However, it has not been fully analyzed in other hematologic malignancies, such as myelodysplastic syndromes (MDS). Materials and MethodsBetween 2010 and 2016, 304 adult patients with de novo MDS had the FLT3 sequence tested on their bone marrow sample. With 279 patients who had follow-up information, we also analyzed the impact of clinical and laboratory characteristics as well as FLT3 mutation status and treatment on prognosis. ResultsWe found that the transformation rate was 3 (42.9%) of 7 patients in the FLT3-ITD-positive group, compared with 31 (10.4%) of 297 among FLT3-ITD-negative patients (P = .033). The median progression-free survival of the FLT3-ITD mutated and wild-type groups were 43 days and 363.5 days, respectively (P < .0001). The median overall survival (OS) of the 2 groups were 218 days and 410.5 days, respectively (P < .0001). We also found that 5 factors had independent prognostic impact on OS: white blood cell counts, bone marrow blast percentage, cytogenetics, transplantation status, and FLT3-ITD mutation. Furthermore, compared with the transformation group, the non-progression group was younger (P = .034), with a lower platelet count (P = .022), a lower bone marrow blast percentage (P = .001), a lower FLT3-ITD incidence (P = .007), and a longer OS (P < .0001). ConclusionsWhen observed at the MDS stage, patients harboring FLT3-ITD mutations had higher AML transformation rate, quicker disease progression, and shorter survival than wild-type patients. Nevertheless, once the disease progressed to leukemia, the impact of FLT3-ITD mutations on prognosis was slight. In addition, the prognosis of secondary AML was very poor whether there was an FLT3-ITD mutation or not.

Shwachman-Diamond Syndrome: Molecular Mechanisms and Current Perspectives.

Shwachman-Diamond syndrome (SDS) is a rare inherited disease mainly caused by mutations in the Shwachman-Bodian-Diamond Syndrome (SBDS) gene. However, it has recently been reported that other genes, including DnaJ heat shock protein family (Hsp40) member C21 (DNAJC21), elongation factor-like 1 (EFL1) and signal recognition particle 54 (SRP54) are also associated with an SDS-like phenotype. Interestingly, SBDS, DNAJC21, EFL1 and SRP54 are involved in ribosome biogenesis: SBDS, through direct interaction with EFL1, promotes the release of the eukaryotic initiation factor 6 (eIF6) during ribosome maturation, DNAJC21 stabilizes the 80S ribosome, and SRP54 facilitates protein trafficking. These findings strengthen the postulate that SDS is a ribosomopathy. SDS is a multiple-organ disease mainly characterized by bone marrow failure, bone malformations, pancreatic insufficiency and cognitive disorders. Almost 15-20% of patients with SDS present myelodysplastic syndrome with a high risk of acute myeloid leukemia (AML) transformation. Unfortunately, besides bone marrow transplantation, no gene-based therapy for SDS has yet been developed. This review aims to recapitulate the recent findings on the molecular mechanisms of SDS underlying bone marrow failure, hematopoiesis and AML development and to draw a realistic picture of current perspectives.

The Src family kinase Fgr is a transforming oncoprotein that functions independently of SH3-SH2 domain regulation.

Fgr is a member of the Src family of nonreceptor tyrosine kinases, which are overexpressed and constitutively active in many human cancers. Fgr expression is restricted to myeloid hematopoietic cells and is markedly increased in a subset of bone marrow samples from patients with acute myeloid leukemia (AML). Here, we investigated the oncogenic potential of Fgr using Rat-2 fibroblasts that do not express the kinase. Expression of either wild-type or regulatory tail-mutant constructs of Fgr promoted cellular transformation (inferred from colony formation in soft agar), which was accompanied by phosphorylation of the Fgr activation loop, suggesting that the kinase domain of Fgr functions independently of regulation by its noncatalytic SH3-SH2 region. Unlike other family members, recombinant Fgr was not activated by SH3-SH2 domain ligands. However, hydrogen-deuterium exchange mass spectrometry data suggested that the regulatory SH3 and SH2 domains packed against the back of the kinase domain in a Src-like manner. Sequence alignment showed that the activation loop of Fgr was distinct from that of all other Src family members, with proline rather than alanine at the +2 position relative to the activation loop tyrosine. Substitution of the activation loop of Fgr with the sequence from Src partially inhibited kinase activity and suppressed colony formation. Last, Fgr expression enhanced the sensitivity of human myeloid progenitor cells to the cytokine GM-CSF. Because its kinase domain is not sensitive to SH3-SH2-mediated control, simple overexpression of Fgr without mutation may contribute to oncogenic transformation in AML and other blood cancers.

Necroinflammation emerges as a key regulator of hematopoiesis in health and disease

The hematopoietic system represents an organ system with an exceptional capacity for the production of mature blood cells from a small and mostly quiescent pool of hematopoietic stem cells (HSCs). This extraordinary capacity includes self-renewal but also the propensity to rapidly respond to extrinsic needs, such as acute infections, severe inflammation, and wound healing. In recent years, it became clear that inflammatory signals such as cytokines, chemokine and danger signals from pathogens (PAMPs) or dying cells (DAMPs) impact on HSCs, shaping their proliferation status, lineage bias, and repopulating ability and subsequently increasing the output of mature effector cells. However, inflammatory danger signals negatively impact on the capacity of HSCs to self-renew and to maintain their stem cell capabilities. This is evidenced in conditions of chronic inflammation where bone marrow failure may originate from HSC exhaustion. Even in hematopoietic cancers, inflammatory signals shape the phenotype of the malignant clone as exemplified by necrosome-dependent inflammation elicited during malignant transformation in acute myeloid leukemia. Accordingly, understanding the contribution of inflammatory signals, and specifically necroinflammation, to HSC integrity, HSC long-term functionality, and malignant transformation has attracted substantial research and clinical interest. In this review, we highlight recent developments and open questions at the interplay between inflammation, regulated necrosis, and HSC biology in the context of blood cell development, acute and chronic inflammation, and hematopoietic cancer.

Differing clinical features between Japanese and Caucasian patients with myelodysplastic syndromes: Analysis from the International Working Group for Prognosis of MDS

Clinical features of myelodysplastic syndromes (MDS) could be influenced by many factors, such as disease intrinsic factors (e.g., morphologic, cytogenetic, molecular), extrinsic factors (e.g, management, environment), and ethnicity. Several previous studies have suggested such differences between Asian and European/USA countries. In this study, to elucidate potential differences in primary untreated MDS between Japanese (JPN) and Caucasians (CAUC), we analyzed the data from a large international database collected by the International Working Group for Prognosis of MDS (300 and 5838 patients, respectively). JPN MDS were significantly younger with more severe cytopenias, and cytogenetic differences: less del(5q) and more +1/+1q, -1/del(1p), der(1;7), -9/del(9q), del(16q), and del(20q). Although differences in time to acute myeloid leukemia transformation did not occur, a significantly better survival in JPN was demonstrated, even after the adjustment for age and FAB subtypes, especially in lower, but not in higher prognostic risk categories. Certain clinical factors (cytopenias, blast percentage, cytogenetic risk) had different impact on survival and time to transformation to leukemia between the two groups. Although possible confounding events (e.g., environment, diet, and access to care) could not be excluded, our results indicated the existence of clinically relevant ethnic differences regarding survival in MDS between JPN and CAUC patients. The good performance of the IPSS-R in both CAUC and JP patients underlines that its common risk model is adequate for CAUC and JP.

Distinct Genomic Associations to Predict Acute Myeloid Leukemia (AML) Progression from Myelodysplastic Syndromes (MDS)

Approximately 25% of MDS patients (pts) progress to AML; 15% of those with lower-risk, and up to 40-50% of those with higher-risk disease. Predicting those who are likely to progress to AML early in their disease course could directly impact treatment decisions: lower-risk pts with higher risk of AML transformation may be offered transplant, while higher-risk pts may be considered for AML-like therapy. In this study, we developed a genomic model that evaluates mutational patterns and their association with AML progression. Development of this model mimics Netflix or Amazon9s recommender system in which customers who bought products A and B, are likely to buy C: pts who have a mutation in genes A and B, are then likely to progress to AML. Clinical and genomic data ofMDS pts diagnosed between 1/1996 and 9/2016 were analyzed. A panel of 60 gene mutations obtained by next generation targeted deep sequencing was included. Association rules using Apriori algorithm was used to study the relationship between multiple genes/cytogenetic abnormalities and AML progression in an unbiased approach. Association rules are a machine learning method that uncovers relationships between variables in a given dataset. Rules with highest confidence (>90% that an association exists) and highest lift (strength of the association) were chosen. Univariate and multivariate analyses were used to evaluate the impact of mutations on AML progression, adjusting for covariates (age, IPSS-R). A total of 527 pts with lower-/higher-risk MDS were analyzed, of those 105 (20%) progressed to AML, with a median time to progression (TTP) of 13 months (interquartile range, 6.3-29.8). Median age at diagnosis was 67 years (range, 19-99) and 38% were female. Risk categories by IPSS-R included: 78 (15%) very low, 200 (38%) low, 95 (18%) intermediate, 98 (18.5%) high, and 56 (10.5%) very high; IPSS-R cytogenetic risk groups were 15 (3%) very good, 331 (63%) good, 87 (16%) intermediate, 37 (7%) poor, and 57 (11%) very poor. The most commonly mutated genes were SF3B1 (14%), ASXL1 (13%), TET2 (12%), SRSF2 (11%), DNMT3A (10%), STAG2 (9%), and TP53 (8%). Per IPSS-R, 14% of pts with very low/low, 21% with intermediate and 30% with high/very high categories progressed to AML. Univariate analyses identified the following cytogenetic and mutational abnormalities to be associated with AML progression: complex karyotype (CK, P =.019), ASXL1 ( P =.023), DNMT3A ( P =.011), FLT3 ( P =.031), PHF6 ( P =.018), PTPN11 ( P =.037) , RUNX1 ( P =.001) , STAG2 ( P =.023) , TET2 ( P =.047) , U2AF1 ( P =.038), and ≥3 gene mutations( P =.002), whereas SF3B1 was associated with lower risk of AML progression ( P =.025). In multivariate Cox regression analyses, including significant mutations, age ( P =.5) and IPSS-R scores ( P RUNX1 (HR2.8 ; 95% CI 1.6-5.3; P= .001), and ≥3 gene mutations (HR=1.65; 95% CI 1.1-2.5; P= .021) were independent factors that associated with AML progression, while SF3B1 (HR=.417; 95% CI .205-.851, P =.016) was associated with lower risk of progression. Association rules identified the following combinations of cytogenetic/genomic abnormalities for AML progression: (ASXL1, RUNX1, STAG2/BCOR), (ASXL1, STAG2, ZRSR2/SRSF2), (ASXL1, TET2, Trisomy8 ), (RUNX1, STAG2, Trisomy 8 ), ( CK , TP53, TET2) . These combinations were associated with a 5 month reduction in time to AML progression.[Figure 1] Using genomic data to reliably identify patients at highest risk for progression to AML early in their disease course can dramatically alter treatment recommendations. Independent factors such as RUNX1 and ≥3 gene mutations predicted for AML progression, while SF3B1 was associated with a reduced risk. Certain cytogenetic/genomic abnormalities were associated with shorter time to AML progression. This data suggests that MDS pts with ≥3 genomic abnormalities or certain genomic combinations could be offered more aggressive therapy early in their disease course including those with lower risk disease. Disclosures Sekeres: Celgene: Membership on an entity9s Board of Directors or advisory committees. Gerds: Incyte: Consultancy; CTI BioPharma: Consultancy. Advani: Takeda/ Millenium: Research Funding; Pfizer: Consultancy. Maciejewski: Alexion Pharmaceuticals, Inc.: Consultancy, Membership on an entity9s Board of Directors or advisory committees, Other: Speaker Fees; Apellis Pharmaceuticals: Consultancy; Ra Pharma: Consultancy.

Validation of the 2017 revision of the WHO chronic myelomonocytic leukemia categories

AbstractThe 2017 revision of the World Health Organization (WHO) classification includes substantial changes to the subclassification of chronic myelomonocytic leukemia (CMML): (1) a 3-tiered blast-based scheme including a novel “CMML-0” category replacing a 2-tiered system in place since 2001 and (2) 2 CMML subtypes, myelodysplastic (MDS-CMML) and myeloproliferative (MP-CMML), based on a white blood cell count cutoff of 13 × 109/L. The clinical utility of this subclassification scheme, particularly the expansion of blast-based subgroups, has not been validated. In this study, a large single-institution CMML patient cohort (n = 629) was used to assess the prognostic impact of the newly proposed categories. Patients were risk stratified according to the CMML-specific Prognostic Scoring System (CPSS) and the MD Anderson Prognostic Scoring System. MP-CMML patients had significantly shorter overall survival (OS; P < .0001; hazard ratio: 0.53, 95% confidence interval: 0.42-0.65) and median duration to acute myeloid leukemia (AML) transformation (P < .0001; 15.2 vs 22.0 months) compared with MDS-CMML patients. The CMML-0 group included 36.4% patients with higher risk CPSS categories and 11.2% of patients with high-risk cytogenetics. Among treatment-naïve patients (n = 499), there was a marginal difference in OS between the CMML-0 and CMML-12017 subgroups (P = .0552). The WHO 2017 blast-based categories were not associated with AML-free survival. Incorporation of the WHO 2017 blast-based subgroups in a modified CPSS scheme had a neutral effect and did not improve its prognostic strength. Our data support the inclusion of MP-CMML and MDS-CMML subtypes in the WHO 2017 revision. Although of some utility in MP-CMML, the 3-tiered blast-based system is not well supported in this study.

USP22 deficiency leads to myeloid leukemia upon oncogenic Kras activation through a PU.1-dependent mechanism

AbstractRas mutations are commonly observed in juvenile myelomonocytic leukemia (JMML) and chronic myelomonocytic leukemia (CMML). JMML and CMML transform into acute myeloid leukemia (AML) in about 10% and 50% of patients, respectively. However, how additional events cooperate with Ras to promote this transformation are largely unknown. We show that absence of the ubiquitin-specific peptidase 22 (USP22), a component of the Spt-Ada-GCN5-acetyltransferase chromatin-remodeling complex that is linked to cancer progression, unexpectedly promotes AML transformation in mice expressing oncogenic KrasG12D/+. USP22 deficiency in KrasG12D/+ mice resulted in shorter survival compared with control mice. This was due to a block in myeloid cell differentiation leading to the generation of AML. This effect was cell autonomous because mice transplanted with USP22-deficient KrasG12D/+ cells developed an aggressive disease and died rapidly. The transcriptome profile of USP22-deficient KrasG12D/+ progenitors resembled leukemic stem cells and was highly correlated with genes associated with poor prognosis in AML. We show that USP22 functions as a PU.1 deubiquitylase by positively regulating its protein stability and promoting the expression of PU.1 target genes. Reconstitution of PU.1 overexpression in USP22-deficient KrasG12D/+ progenitors rescued their differentiation. Our findings uncovered an unexpected role for USP22 in Ras-induced leukemogenesis and provide further insights into the function of USP22 in carcinogenesis.

An increased percentage of myeloid CD34+ bone marrow cells stratifies intermediate IPSS-R myelodysplastic syndrome patients into prognostically significant groups.

The Revised International Prognostic Scoring System (IPSS-R) for myelodysplastic syndromes (MDS) has established an intermediate category where a disease-modifying intervention is a matter of debate. Flow cytometry allows us to determine a fraction of immature myeloid cells in a semiautomated procedure. The aim of this study, mirroring IPSS-R study inclusion criteria, was to test whether bone marrow (BM) CD34+My percentage has independent prognostic value in the MDS setting. BM CD34+My cells were quantified, at diagnosis, selecting CD34+/CD45+/CD11b±/CD13+. Patients were excluded when receiving treatment for altering the natural course of the disease and when IPSS-R could not be calculated due to the lack of metaphases. Finally, Cox analyses were performed, on a series of 260 patients, for overall survival (OS) and time to acute myeloid leukemia (AML) transformation. By analyzing ROC curves, the most accurate prognostic variable, regarding blasts by cytology and CD34+ by cytometry, was the percentage of blasts by microscopy. The percentage of CD34+My in BM showed an AUC of 0.767 and 0.576 for time to AML transformation and OS, respectively. When performing a multivariate regression including the IPSS-R and the percentage of BM CD34+My cells >1%, both factors predicted for a shorter time to AML transformation. In addition, CD34+My percentage successfully stratified the intermediate IPSS-R category into two prognostic groups with a relative risk of 5.73 (95% CI [1.2-27.8]; P=.03). We found that BM CD34+My percentage has an independent value concerning the IPSS-R, especially relevant for the prediction of transformation to AML and within the intermediate group.

MULTICOLOR FLOW CYTOMETRY AS A DIAGNOSTIC TOOL IN MYELODYSPLASTIC SYNDROMES

Myelodysplastic syndromes (MDS) are heterogeneous group of clonal hematological neoplasms characterized by cytopenias, sighs of dysplasia and high risk of transformation in acute myeloid leukemia. Diagnostics of MDS requires a comprehensive approach, but even when performing cytological, cytogenetic, molecular and histological studies, it is not always possible to establish a diagnosis. Multicolor flow cytometry (MFC) is a high-tech method, increasingly used in the diagnosis of various hematologic diseases. Over the last years, MFC is standardized for assessing of dysplasia in all cell compartments of the bone marrow. Many research groups have demonstrated high sensitivity and specificity of this diagnostic approach, which indicates the necessity of integration of flow cytometric study into diagnostic protocols of MDS.

Atypical chronic myeloid leukaemia: A case of an orphan disease-A multicenter report by the Polish Adult Leukemia Group.

Atypical chronic myeloid leukaemia (aCML) belongs to myelodysplastic/myeloproliferative neoplasms. Because of its rarity and changing diagnostic criteria throughout subsequent classifications, data on aCML are very scarce. Therefore, we at the Polish Adult Leukemia Group performed a nationwide survey on aCML. Eleven biggest Polish centres participated in the study. Altogether, 45 patients were reported, among whom only 18 patients (40%) fulfilled diagnostic criteria. Among misdiagnosed patients, myelodysplastic/myeloproliferative syndrome unclassifiable and chronic myelomonocytic leukaemia were the most frequent diagnoses. Thirteen patients were male, median age 64.6years (range 40.4-80.9). The median parameters at diagnosis were as follows: white blood cell count 97×109 /L (23.8-342) with immature progenitors amounting at 27.5% (12-72), haemoglobin 8.6g/dL (3.9-14.9), and platelet count 66×109 /L (34-833). Cytoreductive treatment was used in all patients, and 2 patients underwent allogeneic hematopoietic stem cell transplantation. The median overall survival was 14.1months (95% CI, 7.2), with median acute myeloid leukaemia-free survival of 13.3months (95% CI, 3.6-22.6). Cumulative incidence of acute myeloid leukaemia transformation after 1year in aCML group was 12.5% (95% CI, 0%-29.6%). To conclude, aCML harbours a poor prognosis. Treatment options are limited, with allogeneic hematopoietic stem cell transplantation being the only curative method at present, although only a minority of patients are transplant eligible. Educational measures are needed to improve the quality of diagnoses.

DNA Methylation in Cancer and Development: An Evolving Paradigm

DNA methylation is an epigenetic mark which, in mammals, occurs primarily on position 5 of cytosines, especially at those found in the context of CpG dinucleotides. This CpG methylation is known to play a major role in gene regulation. Cytosine methylation is regulated by the DNA methyltransferases, responsible for adding the methyl group to unmethylated CpGs, and the TET dioxygenases, involved in the DNA demethylation pathway. Initially, DNA methylation was believed to be important mainly for gene silencing through promoter DNA methylation, especially at CpG-rich promoters containing CpG islands. However, our understanding of the role that DNA methylation plays in gene regulation during normal development and how this process becomes deregulated in cancer, has evolved in recent years. Moreover, the discovery of frequent mutations in DNMT3A and TET2 both in clonal hematopoiesis of indeterminate significance as well as in many hematological malignancies has brought new interest into understanding what role DNA methylation plays in normal HSC function as well as how it contributes to malignant transformation. In this session, we will review the current understanding in the field of DNA methylation and gene regulation, and present data on DNA methylation in normal HSCs as well as the role that this epigenetic mark plays during leukemic transformation in acute myeloid leukemia. Disclosures No relevant conflicts of interest to declare.

Non-driver mutations in patients with JAK2V617F-mutated polycythemia vera or essential thrombocythemia with long-term molecular follow-up

JAK2V617F monitoring and NGS of non-driver genes was performed in 100 patients with polycythemia vera (PV) or essential thrombocythemia (ET) with long molecular follow-up. Patients who did not progress to myelofibrosis (MF) or acute myeloid leukemia (AML) after more than 10years (n = 50) showed a low frequency of mutations at first sample (18%) and an incidence rate of 1.7 new mutations × 100person-years. Mutations were detected at first sample in 83% of PV/ET patients who later progressed to AML (n = 12) with these patients having a rate of 25.6 mutations × 100 person-years. Presence of mutations at diagnosis was the unique risk factor for acquiring a new genetic event (HR 2.7, 95% CI 1.1-6.8, p = 0.03) after correction for age, PV diagnosis, and total duration of hydroxyurea (HU) exposure. Patients with additional mutation at first sample showed a higher probability of developing cytopenia under HU therapy and a higher risk of AML (HR 12.2, 95% CI 2.6-57.1, p = 0.001) with mutations in ASXL1 (p < 0.0001), TP53 (p = 0.01), SRSF2 (p < 0.0001), IDH1/2 (p < 0.0001), and RUNX1 (p < 0.0001) being associated with a higher probability of AML. Myelofibrotic transformation was more frequent in patients with additional mutations, especially in SF3B1 (p = 0.02) and IDH1/2 (p < 0.0001) although a persistently high or a progressive increase of the JAK2V617F allele burden while receiving cytoreduction was the strongest predictor of MF transformation (HR 10.8, 95% CI 2.4-49.1, p = 0.002). In conclusion, NGS may be useful to identify a minority of PV and ET patients with high genetic instability and increased risk of AML transformation.