ALBERT

Description: An extreme heat wave occurred in Russia in the summer of 2010. It had serious impacts on humans and natural ecosystems, it was the strongest recorded globally in recent decades and exceeded in amplitude and spatial extent the previous hottest European summer in 2003. Earlier studies have not succeeded in comparing the magnitude of heat waves across continents and in time. This study introduces a new Heat Wave Magnitude Index (HWMI) that can be compared over space and time. The index is based on the analysis of daily maximum temperature, in order to classify the strongest heat waves that occurred worldwide during the three study periods 1980-1990, 1991-2001 and 2002-2012. In addition, multi-model ensemble outputs from the Intercomparison Project Phase 5 (CMIP5) are used to project future occurrence and severity of heat waves, under different Representative Concentration Pathways (RCP), adopted by the Intergovernmental Panel on Climate Change (IPCC) for its fifth Assessment Report (AR5). Results show that the percentage of global area affected by heat waves has increased in recent decades. Moreover, model predictions reveal an increase in the probability of occurrence of extreme and very extreme heat waves in the coming years: in particular, by the end of this century, under the most severe IPCC AR5 scenario, events of the same severity as that in Russia in the summer of 2010 will become the norm and are projected to occur as often as every two years for regions such as southern Europe, North America, South America, Africa and Indonesia.

Description: Polycythemia vera (PV) presents with iron deficiency at diagnosis, and the mainstay of treatment, i.e. therapeutic phlebotomy, worsens iron deficiency. Iron deficiency is associated with anemia and symptoms of cognitive impairment and fatigue even in the absence of anemia, and patients with low risk PV often suffer from iron deficiency related symptoms How iron deficiency develops in PV patients prior to phlebotomy is not well understood. We previously demonstrated that PV patients exhibit a greater extent of iron deficiency relative to wild type JAK2 patients with other causes of erythrocytosis [Ginzburg Leuk 2018]. We hypothesize that mutated JAK2 leads to aberrant insensitivity of erythropoiesis to iron deficiency. To explore this hypothesis, we first analyzed serum data from iron deficient PV patients (n=14) and blood donors (n=15), normalized for age (51 vs. 42 years, P=0.12) and serum ferritin concentration (22 vs. 22 ng/ml, P=0.95); our data demonstrate that PV patients have significantly lower hepcidin, transferrin saturation, MCV, a higher HCT, and a trend toward higher erythroferrone (ERFE) relative to controls (Fig 1a-1e). Secondly, CD34+ cells were isolated from mononuclear cells and plated with erythropoietin and either 100% or 10% transferrin saturation to mimic iron replete and iron deficient conditions, respectively; differentiation and proliferation were analyzed using flow cytometry. These experiments revealed that although glycophorin A (GPA) and CD36+ cells were decreased in iron deficient relative to iron replete control cells, PV cells continue to proliferate irrespective of iron status (Fig 1f). In addition, only iron deficient control, not PV, cells demonstrated an erythroid lineage specific decrease in proliferation relative to iron replete cells (Fig 1g), demonstrating that iron restriction in PV does not limit erythroid differentiation or proliferation in vitro. Thirdly, we transplanted JAK2 V617F (PV) and wild type (WT) cells into recipient females and placed mice on 35ppm (iron replete (IR)) or 2.5ppm (iron deficient (ID)) diets. IR PV mice exhibited the expected erythrocytosis and decrease in MCV and MCH relative to WT controls (Fig 2a-2d). WT mice on an ID diet exhibited decreased MCV, MCH, and RET-He while PV mice had decreased RBC counts with an increased MCHC (Fig 2a, 2c-2f). These findings demonstrate altered iron regulation in PV erythroblasts with a preference for decreasing RBC count, rather than cellular Hb production, in iron deficiency in vivo. No differences were found in the total number of bone marrow erythroblasts in PV mice relative to WT mice on IR or ID diets. IR and ID PV mice demonstrated splenomegaly relative to WT controls. In addition, IR PV erythroblasts expressed significantly more ERFE relative to WT controls with decreased ERFE expression in ID PV erythroblasts (Fig 2g). Similarly, liver hepcidin expression was lower in IR PV relative to WT controls, but was restored in ID PV mice (Fig 2h), the later likely a response to decreased ERFE expression (Fig 2g); no changes are observed in liver iron concentration in IR relative to ID PV mice. Furthermore, the expected decrease in pSTAT is observed only in ID WT (Fig 2i), not ID PV (Fig 2j) erythroblasts. Lastly, TfR2 protein expression was increased in IR PV relative to WT controls (Fig 2k) and decreased only in ID WT (Fig 2l) but not ID PV (Fig 2m) erythroblasts. Since TfR2 degradation is enhanced during iron deficiency [Khalil JEM 2018] and TfR2 enables iron delivery for mitochondrial heme synthesis [Khalil Blood Adv 2017], persistently increased TfR2 in PV erythroblasts may explain why cellular Hb synthesis (i.e. MCH and RET-He) remains unaffected by iron deficiency in PV mice. Taken together, these findings demonstrate that in vitro iron deficiency fails to limit differentiation and proliferation in PV erythroblasts and enhances STAT5 signaling in ID PV erythroblasts in vivo but results in decreased circulating RBCs in PV mice. In addition, decreased erythroblast ERFE expression in ID PV mice results in increased hepcidin to limit erythroblast iron availability, but persistently increased TfR2 concentration enables mitochondrial iron delivery for Hb synthesis despite cellular iron deficiency. Our studies provide novel mechanistic insights into the dysregulation of iron utilization for erythropoiesis in PV. Disclosures Levine: Prelude Therapeutics: Research Funding; Imago Biosciences: Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Research Funding; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria; Novartis: Consultancy; Gilead: Consultancy; Lilly: Honoraria; Loxo: Membership on an entity's Board of Directors or advisory committees; Qiagen: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Research Funding; Isoplexis: Membership on an entity's Board of Directors or advisory committees. Hoffman:Merus: Research Funding. Ginzburg:La Jolla Pharma: Membership on an entity's Board of Directors or advisory committees.

Description: Background: Activating mutations of NRAS and KRAS genes are common in newly diagnosed acute myeloid leukemia (AML), occurring in 11-16% and 4-5% of patients, respectively. RAS mutations are frequently acquired at time of progression from MDS to AML and are associated with poor survival. Next generation sequencing (NGS) at diagnosis and during complete remission has shown that RAS mutations have high clearance rates with induction chemotherapy. In the CALGB 8525 study, RAS-mutant younger patients (age

Description: Background Measurable residual disease (MRD) is associated with inferior outcomes in patients with acute myeloid leukemia (AML). MRD monitoring enhances risk stratification and may guide therapeutic intervention. Post-induction MRD is frequently cleared with further therapy and the clearance may lead to better outcomes. In contrast, persistent MRD is associated with poor outcomes. At present it is not possible to predict which patients are likely to clear MRD with further therapy. Here we report a simple, objective, widely applicable and quantitative MFC approach using the ratio of blast/PDC to predict persistent MRD and poor outcomes in AML. Patients and Methods A cohort of 136 adult patients with a confirmed diagnosis of AML by WHO criteria who underwent standard induction therapy at a single center between 4/2014 and 9/2017 was initially included. 69 patients achieved complete morphologic remission (36 MRD-neg. and 33 MRD-pos.). MRD status was assessed by MFC using a different from normal (DfN) approach. PDC were quantified as the percent of total WBC by flow cytometry based on low side scatter, moderate CD45, CD303, bright CD123 and HLA-DR expression. Results The proportion of PDC was markedly decreased in patients with AML (≥20% blasts) (N=136) with a median of 0.016% (interquartile range IQR: 0.0019%-0.071%, Figure 1A), more than 10-fold lower than observed in normal controls (median 0.23%, IQR 0.17%-0.34%) (N=20). While there was no difference between MRD-neg. and normal control groups (median 0.31%, IQR: 0.17%-0.49%; vs. 0.28%, IQR: 0.17%-0.34%), MRD-pos. group had significantly reduced PDC proportion compared to the control (median 0.074%, IQR: 0.022%-0.33%, Wilcoxon rank sum, p=0.019). In an attempt to achieve better separation and to eliminate possible effects of hemodilution, the ratio of blast/PDC was calculated by using the proportions of blasts and PDCs out of total WBCs as quantitated by flow cytometry. A cut-off threshold of the blast/PDC ratio of 10 was chosen to separate each group (Figure 1B). Importantly, a ratio cut-off of 10 had a corresponding specificity of 97.4% for predicting MRD positivity status. MRD positivity was significantly associated with inferior overall survival (OS) and relapse-free survival (RFS) in our study cohort (OS HR 4.11 (95% CI: 1.30-13.03), p=0.016; RFS HR 4.20 (95% CI: 1.49-11.82), p=0.007, Figure 1C and D). The 2-year cumulative incidence of relapse in the MRD-neg. group compared to MRD-pos. group was 10% (95% CI: 2-24%) vs. 37% (95% CI: 18-56%, p=0.014). Importantly, blast/PDC ratio ≥10 was also strongly associated with inferior OS and RFS (OS HR 3.12 (95% CI: 1.13-8.60), p= 0.028; RFS HR 4.05 (95% CI: 1.63-10.11), p=0.003, Figure 1E and F), which is similar in magnitude to MRD positivity. Furthermore, MRD-pos. patients with blast/PDC ratio

Description: Secondary acute myeloid leukemias (AMLs) evolving from an antecedent myeloproliferative neoplasm (MPN) are characterized by a unique set of cytogenetic and molecular features distinct from de novo AML. Given the high frequency of poor-risk cytogenetic and molecular features, malignant clones are frequently insensitive to traditional AML chemotherapeutic agents. Allogeneic stem cell transplant, the only treatment modality shown to have any beneficial long-term outcome, is often not possible given the advanced age of patients at time of diagnosis and frequent presence of competing comorbidities. Even in this setting, relapse rates remain high. As a result, outcomes are generally poor and there remains a significant unmet need for novel therapeutic strategies. Although advances in cancer genomics have dramatically enhanced our understanding of the molecular events governing clonal evolution in MPNs, the cell-intrinsic and -extrinsic mechanisms driving leukemic transformation at this level remain poorly understood. Here, we review known risk factors for the development of leukemic transformation in MPNs, recent progress made in our understanding of the molecular features associated with leukemic transformation, current treatment strategies, and emerging therapeutic options for this high-risk myeloid malignancy.

Description: Fanconi anaemia (FA) is a multigenic chromosomal (chr) instability syndrome characterized by a wide array of congenital defects and early predisposition to bone marrow (BM) failure and solid tumors. Most frequent hematologic complications are aplastic anemia (AA) or myelodysplasia (MDS) with a high AML transformation rate. Genomic instability resulting from impaired DNA crosslink repair in FA may lead to unbalanced chr aberrations that can be detected in clonally expanding BM stem cells. Most frequent abnormalities include gain1q, del 7 and gain3q, the latter two are associated with a poor prognosis. Due to low resolution of metaphase cytogenetics (MC) and its dependence upon in-vitro cell growth, detection of chr lesions is often limited or non-informative. High-density SNP arrays (SNPA) can be used for a very precise identification of unbalanced genomic lesions and copy-neutral LOH. We hypothesize that using SNPA previously cryptic clonal genomic aberrations can be detected in some FA patients and their characterization may help predict propensity to transformation and point towards pathogenetic lesions. We applied Affymetrix 250K SNP-A to investigate patients with FA (N=21: 15 FANCA, 4 FANCG and 2 FANCC; avg age: 12y) and as controls healthy BM transplant donors (n=50). For comparison, non-FA patients with AA (n=68), MDS (n=98) and secondary AML (n=30) were analyzed. Lesions spanning more than 100 SNPs were defined as significant. Copy number polymorphisms detected in controls and public data bases were not considered pathologic. Globally, MC was abnormal in 48% of FA patients while SNP-karyotyping revealed chr lesions in 85% of patients. The additional lesions detected by SNPA included monosomy 7, deletions (1,5,6,11,12,22,18,20) and gains (1,3,4,21). For most FA patients comparative genomic hybridization and FISH were performed and confirmed genomic gains and losses detected by SNPA. In addition, using SNPA, copy number-neutral LOH, segmental uniparental disomy (UPD) was identified in 43% of FA patients on various chr (2,3,4,6,8,9,11,13,16,18). By comparison tin AA, MDS and AML, UPD was present in 31%, 21% and 23%, respectively. Next, patients with known recurrent defects were analyzed with regard to the presence of additional UPD, which was detected in 50% of patients with gain on chr.3q vs. in 38% of patients without gain3q, 50% of patients with vs. 36% without gain1q, and in 42% of patients with vs. 28% without del 7, respectively. On 3 FA patients with advanced MDS (RAEBt; n=2) and AA (n=1) SNPA was performed on archived samples collected prior to progression into AML. In all these patients clonal chr aberrations were detected, including various previously cryptic lesions (gain4q, gain21q, UPD on 8q and 9p). Our study represents the first application of high-density SNPA for karyotyping of patients with FA and demonstrates that this method may uncover cryptic clonal chr lesions, including copy number neutral LOH due to UPD. This type of lesion is a novel finding in FA. Further studies are required to determine the functional significance of these lesions and their correlation with leukemia or solid tumors.

Description: Interstitial deletion of chromosome 5 has been frequently reported in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) pointing towards the pathogenic role of this region in phenotype and clonal evolution. To investigate the frequency of 5q LOH and better delineate the commonly deleted region (CDR) we applied metaphase cytogenetics (MC) and performed array-based interphase cytogenetics on DNA using high-density 250K SNP-array (SNP-A) in malignant myeloid disorders. SNP-A analysis was performed using Affymetrix 250K SNP-array on a subset of 250 patients and 118 controls. SNP-A data were analyzed using CNAG v.3.0. Using SNP-A-based karyotyping 5q abnormalities were identified in 42/250 patients vs. 29/250 by MC (17% vs. 11%, primarily as a result of resolving uninformative MC). By SNP-A, previously cryptic lesions other than 5q deletions were identified in 71% of the patients (30/42) and included e.g., upd(1)(p35.1pter), upd(17)(p13.1pter), del(7)(q21.3q36.2). In 2 patients somatic UPD5q was found. In 6 other patients 5q micro-deletions of chromosome 5 were detected likely representing CNVs. Subsequently, we undertook an SNP-A-based definition of the CDR covering approximately 1.92MB and positioned within previously reported regions. This region spans approximately from base pair 137,528,564-139,451,907 and several candidate tumor suppressor genes including those already reported e.g., CTNNA1 and EGR1 as well as CDC25, ETF1 to name a few. As mutations in any of the genes in CDR on 5q were not found, the malignant phenotype can result either from haploinsufficiency or hemizygozity for a pathogenic allele occurring otherwise in heterozygous or major allele in homozygous constellation. Previously, underexpression of RPS14 was demonstrated to be responsible for dysplastic features in 5q- syndrome, but haploinsufficiency of this gene does not explain the clonal evolution and malignant properties of clones with del5q. Utilizing another advantage of SNP-A, the ability to assign genotypes, we also performed analysis of the allele distribution within newly defined CRD on 5q31. We genotyped 987 SNPs located in the common area of LOH (5q31) to determine whether for certain loci allelic distribution in the hemizygous deletion mutant is skewed to one or the other allele (when the original diploid constellation is heterozygous). For the purpose of this study we hypothesized that informative alleles must display a very low frequency in homozygous constellation in controls. To test if the distribution of remaining alleles (and the inherent change of genotype) in the deleted region on 5q is random or skewed towards one allelic variant we applied a multinomial test statistics. Because of the discrete nature of the problem, p-values were calculated by selecting the minimum of the left or right tail of the event spectrum. This value was computed without approximation by using the exact form of the binomial distribution. We used the smaller of the two p-values from the two binomial models as our reported p-value. SNPs present in ARHGAP26 locus appeared to be most significantly skewed (p