CRISPR/Cas9-mediated editing of ACTB in induced pluripotent stem cells: A model for investigating human ACTB loss-of-function and genetic adaptive responses.

Heterozygous beta-actin (ACTB) indel and nonsense mutations are linked to developmental disorders. We generated two CRISPR/Cas9 human induced pluripotent stem cell (iPSC) lines, WTSIi018-B-19 and WTSIi018-B-20, carrying heterozygous and homozygous indel mutations in ACTB exon 4. Both iPSCs exhibited normal cell morphology, expression of pluripotency markers, and the ability to differentiate into the three primary germ layers. While iPSCs with a heterozygous ACTB mutation maintain genome integrity, homozygous mutants showed a loss of heterozygosity in chromosome three. These mutants provide a powerful model to study the onset, progression, and complex interplay of genetic compensation and phenotypic variation of ACTB-related diseases.

Prime-Editing of human ACTB in induced pluripotent stem cells to model human ACTB Loss-of-Function diseases and compensatory mechanisms.

Beta-actin (ACTB) heterozygous loss-of-function mutations are associated with pleiotropic developmental disorders entailing intellectual disability and frequent organ malformations in affected individuals. We generated two CRISPR/Cas9 prime-edited human induced pluripotent stem cell (iPSC) lines, IUFi004-A-1 and IUFi004-A-2, carrying a heterozygous missense mutation in exon 4 of ACTB. Mutant iPSCs exhibited normal cell morphology and genomic integrity, maintained expression of pluripotency markers, and differentiated into the three primary germ layers. The mutants offer a valuable platform for examining the molecular and functional consequences of ACTB haploinsufficiency, developing effective treatments, and exploring mechanisms underlying phenotypic variability and genetic compensation observed in monogenic diseases.

The Absolute Monocyte Count at Diagnosis Affects Prognosis in Myelodysplastic Syndromes Independently of the IPSS-R Risk Score.

The absolute monocyte count (AMC) is associated with mortality in a variety of medical conditions. Its prognostic impact in myelodysplastic syndromes (MDSs) is less well studied. Therefore, we investigated its potential prognostic value in a cohort from the Düsseldorf MDS registry in relationship to the revised international prognostic scoring system (IPSS-R). An AMC below the population's median (<0.2 × 109/L) was associated with several adverse disease features such as lower haemoglobin levels, lower count of neutrophils and platelets, and a higher percentage of blasts in the bone marrow. MDS patients with an AMC < 0.2 × 109/L had a significantly higher risk of progression into acute myeloid leukemia (AML). In a univariate, proportional hazards model the effect of the AMC as a continuous variable was modelled via p-splines. We found a U-shaped effect with the lowest hazard around 0.3 × 109/L. Accordingly, an AMC within the last quartile of the population (0.4 × 109/L) was associated with a reduced overall survival independently of IPSS-R, but not with the risk of secondary AML. Considering monocytopenia as a risk factor for AML progression in MDS may provide an additional argument for allogeneic transplantation or the use of hypomethylating agents in patients who are not clear candidates for those treatments according to current prognostic scoring systems and/or recommendations. Further studies are needed to assess the prognostic impact of the AMC in the context of prognostic scoring systems, considering the molecular risk profile, and to identify the mechanisms responsible for the higher mortality in MDS patients with a subtle monocytosis.

Biallelic variants in CRIPT cause a Rothmund-Thomson-like syndrome with increased cellular senescence.

PURPOSE: Rothmund-Thomson syndrome (RTS) is characterized by poikiloderma, sparse hair, small stature, skeletal defects, cancer, and cataracts, resembling features of premature aging. RECQL4 and ANAPC1 are the 2 known disease genes associated with RTS in >70% of cases. We describe RTS-like features in 5 individuals with biallelic variants in CRIPT (OMIM 615789). METHODS: Two newly identified and 4 published individuals with CRIPT variants were systematically compared with those with RTS using clinical data, computational analysis of photographs, histologic analysis of skin, and cellular studies on fibroblasts. RESULTS: All CRIPT individuals fulfilled the diagnostic criteria for RTS and additionally had neurodevelopmental delay and seizures. Using computational gestalt analysis, CRIPT individuals showed greatest facial similarity with individuals with RTS. Skin biopsies revealed a high expression of senescence markers (p53/p16/p21) and the senescence-associated ß-galactosidase activity was elevated in CRIPT-deficient fibroblasts. RECQL4- and CRIPT-deficient fibroblasts showed an unremarkable mitotic progression and unremarkable number of mitotic errors and no or only mild sensitivity to genotoxic stress by ionizing radiation, mitomycin C, hydroxyurea, etoposide, and potassium bromate. CONCLUSION: CRIPT causes an RTS-like syndrome associated with neurodevelopmental delay and epilepsy. At the cellular level, RECQL4- and CRIPT-deficient cells display increased senescence, suggesting shared molecular mechanisms leading to the clinical phenotypes.

Remote digital assessment of amyotrophic lateral sclerosis functional rating scale - a multicenter observational study.

OBJECTIVE: Remote self-assessment of the revised amyotrophic lateral sclerosis functional rating scale (ALSFRS-R) using digital data capture was investigated for its feasibility as an add-on to ALSFRS-R assessments during multidisciplinary clinic visits. METHODS: From August 2017 to December 2021, at 12 ALS centers in Germany, an observational study on remote assessment of the ALSFRS-R was performed. In addition to the assessment of ALSFRS-R during clinic visits, patients were offered a digital self-assessment of the ALSFRS-R - either on a computer or on a mobile application ("ALS-App"). RESULTS: An estimated multicenter cohort of 4,670 ALS patients received care at participating ALS centers. Of these patients, 971 remotely submitted the ALSFRS-R, representing 21% of the multicenter cohort. Of those who opted for remote assessment, 53.7% (n = 521) completed a minimum of 4 ALSFRS-R per year with a mean number of 10.9 assessments per year. Different assessment frequencies were found for patients using a computer (7.9 per year, n = 857) and mobile app (14.6 per year, n = 234). Patients doing remote assessments were more likely to be male and less functionally impaired but many patients with severe disability managed to complete it themselves or with a caregiver (35% of remote ALSFRS-R cohort in King's Stage 4). CONCLUSIONS: In a dedicated ALS center setting remote digital self-assessment of ALSFRS-R can provide substantial data which is complementary and potentially an alternative to clinic assessments and could be used for research purposes and person-level patient management. Addressing barriers relating to patient uptake and adherence are key to its success.

Clinical and Cytogenetic Characterization of Early and Late Relapses in Patients Allografted for Myeloid Neoplasms with a Myelodysplastic Component.

An improved understanding of relapse kinetics is required to optimize detection and treatment strategies for the post-transplant relapse of myeloid neoplasms. Therefore, we retrospectively analyzed data from 91 patients allografted for MDS (n = 54), AML-MRC (n = 29) and chronic myelomonocytic leukemia (CMML, n = 8), who relapsed after transplant. Patients with early (<12 months, n = 56) and late relapse (>12 months, n = 35) were compared regarding patient-, disease- and transplant-related factors, including karyotype analyses at diagnosis and relapse. After a median follow-up of 17.4 months after relapse, late relapses showed improved outcomes compared with early relapses (2-yr OS 67% vs. 32%, p = 0.0048). Comparing frequency of distinct patient-, disease- and transplant-related factors among early and late relapses, complex karyotype (p = 0.0004) and unfavorable disease risk at diagnosis (p = 0.0008) as well as clonal evolution at relapse (p = 0.03) were more common in early than in late relapses. Furthermore, patients receiving transplant without prior cytoreduction or in complete remission were more frequently present in the group of late relapses. These data suggest that cytogenetics rather than disease burden at diagnosis and transplant-related factors determine the timepoint of post-transplant relapse and that upfront transplantation may be favored in order to delay relapse.

Generation of an induced pluripotent stem cell line (IUFi002-A) from a Leigh syndrome patient carrying mutations in the NDUFS1 gene.

Human dermal fibroblasts from a Leigh Syndrome (LS) patient harboring the heterozygous NDUFS1 R557X/D618N compound mutation were reprogrammed to generate integration-free induced pluripotent stem cells (iPSCs). The full characterization of IUFi002-A-iPSCs demonstrated that the line is free of exogenous reprogramming genes and maintains the genomic integrity. IUFi002-A-iPSCs' pluripotency was confirmed by the expression of pluripotency markers and embryoid body-based differentiation into cell types representative of each of the three germ layers. The generated iPSC line provides a powerful tool to investigate LS and analyze the molecular mechanisms underlying NDUFS1 mutations-induced pathology.

Use and subjective experience of the impact of motor-assisted movement exercisers in people with amyotrophic lateral sclerosis: a multicenter observational study.

Motor-assisted movement exercisers (MME) are devices that assist with physical therapy in domestic settings for people living with ALS. This observational cross-sectional study assesses the subjective experience of the therapy and analyzes users' likelihood of recommending treatment with MME. The study was implemented in ten ALS centers between February 2019 and October 2020, and was coordinated by the research platform Ambulanzpartner. Participants assessed symptom severity, documented frequency of MME use and rated the subjective benefits of therapy on a numerical scale (NRS, 0 to 10 points, with 10 being the highest). The Net Promotor Score (NPS) determined the likelihood of a participant recommending MME. Data for 144 participants were analyzed. Weekly MME use ranged from 1 to 4 times for 41% of participants, 5 to 7 times for 42%, and over 7 times for 17%. Particularly positive results were recorded in the following domains: amplification of a sense of achievement (67%), diminution of the feeling of having rigid limbs (63%), diminution of the feeling of being immobile (61%), improvement of general wellbeing (55%) and reduction of muscle stiffness (52%). Participants with more pronounced self-reported muscle weakness were more likely to note a beneficial effect on the preservation and improvement of muscle strength during MME treatment (p < 0.05). Overall, the NPS for MME was high (+ 61). High-frequency MME-assisted treatment (defined as a minimum of five sessions a week) was administered in the majority of participants (59%) in addition to physical therapy. Most patients reported having achieved their individual therapeutic objectives, as evidenced by a high level of satisfaction with MME therapy. The results bolster the justification for extended MME treatment as part of a holistic approach to ALS care.

Progenitor cells derived from gene-engineered human induced pluripotent stem cells as synthetic cancer cell alternatives for in vitro pharmacology.

Limitations in genetic stability and recapitulating accurate physiological disease properties challenge the utility of patient-derived (PD) cancer models for reproducible and translational research. A portfolio of isogenic human induced pluripotent stem cells (hiPSCs) with different pan-cancer relevant oncoprotein signatures followed by differentiation into lineage-committed progenitor cells was genetically engineered. Characterization on molecular and biological level validated successful stable genetic alterations in pluripotency state as well as upon differentiation to prove the functionality of our approach. Meanwhile proposing core molecular networks possibly involved in early dysregulation of stem cell homeostasis, the application of our cell systems in comparative substance testing indicates the potential for cancer research such as identification of augmented therapy resistance of stem cells in response to activation of distinct oncogenic signatures.

Generation of an induced pluripotent stem cell line (IUFi001) from a Cockayne syndrome patient carrying a mutation in the ERCC6 gene.

Human fibroblasts from a Cockayne Syndrome (CS) patient carrying the compound heterozygous c.1131 A > T and c.2571C > T within ERCC Excision Repair 6 (ERCC6) were reprogramed to generate integration-free induced pluripotent stem cells (iPSCs). Characterization of IUFi001-iPSCs demonstrated that this iPSC line is free of exogenous reprogrammed genes and maintains the genomic integrity. The pluripotency of IUFi001-iPSCs was confirmed by the expression of the pluripotency-associated markers and by embryoid body-based differentiation into cell types representative of the three germ layers. The generated iPSC line provides a powerful tool to dissect the molecular mechanisms underlying CS caused by mutations within ERCC6.

Influence of platelet count at diagnosis and during the course of disease on prognosis in MDS patients.

Thrombocytopenia at diagnosis and platelet drop within the first 6 months have an adverse effect on prognosis of MDS patients. We therefore were interested in the association and impact on prognosis of morphologic findings of megakaryocytes and platelets with platelet count at diagnosis, bleeding complications, and the drop of platelets during the course of disease. This retrospective analysis was based on 334 MDS patients from the Duesseldorf MDS registry that were followed up for blood counts, bleeding, transfusion dependency, and AML evolution and correlated with morphology of the megakaryocytes and platelets. Thrombocytopenia was found more frequently in higher risk MDS and was associated with hypocellularity of the megakaryocytes in the bone marrow. Signs of bleeding were present at diagnosis in 14% and occurred during the disease in 48% of all MDS patients. Death due to bleeding was ranked third behind infections and AML. A decrement of platelets during the first 6 months was associated with an inferior overall survival of 21 vs. 49 months and with a higher cumulative 2-year AML rate of 22.2% vs. 8.3% (p = 0.001). In a multivariate analysis, besides bone marrow blasts and karyotype, decreasing platelets were also associated with an inferior outcome. Signs of bleeding are present in a relevant number of MDS patients and account for significant morbidity and mortality in MDS. We could demonstrate the prognostic importance of decreasing platelets during the course of disease in all MDS patients, identifying patients at higher risk for death or AML progression.

Academic application of Good Cell Culture Practice for induced pluripotent stem cells.

Human induced pluripotent stem cells (hiPSC) are a promising tool for replacing animal-based experiments. To warrant data reproducibility, quality-controlled research material is recommended. While the need for global harmonization of quality standards for stem cell banking centers, commercial providers, pre-clinical and clinical use of cells is well doc­umented, there are no recommendations available for quality control of hiPSC in an academic research environment to date. To fill this gap, we here give an example of a quality-controlled, two-tiered banking process producing a fully characterized master cell bank (MCB) and partially characterized respective working cell banks (WCB). Characteri­zation includes the study of morphology, mycoplasma contamination, cell line identity, karyotype stability, cell antigen expression and viability, gene expression, pluripotency, and post-thaw recovery. Costs of these procedures are cal­culated. We present the results of the proposed testing panel of two hiPSC MCBs and show that both fulfil the defined specifications regarding the above-mentioned characterization assays during and upon banking. In conclusion, we propose a panel of eight assays, which are practical and useful for an academic research laboratory working with hiPSCs. Meeting these proposed specifications ensures the quality of pluripotent stem cells throughout diverse experi­ments at moderate costs.

Acute myeloid leukemia-induced functional inhibition of healthy CD34+ hematopoietic stem and progenitor cells.

Acute myeloid leukemia (AML) is characterized by an expansion of leukemic cells and a simultaneous reduction of normal hematopoietic precursors in the bone marrow (BM) resulting in hematopoietic insufficiency, but the underlying mechanisms are poorly understood in humans. Assuming that leukemic cells functionally inhibit healthy CD34+ hematopoietic stem and progenitor cells (HSPC) via humoral factors, we exposed healthy BM-derived CD34+ HSPC to cell-free supernatants derived from AML cell lines as well as from 24 newly diagnosed AML patients. Exposure to AML-derived supernatants significantly inhibited proliferation, cell cycling, colony formation, and differentiation of healthy CD34+ HSPC. RNA sequencing of healthy CD34+ HSPC after exposure to leukemic conditions revealed a specific signature of genes related to proliferation, cell-cycle regulation, and differentiation, thereby reflecting their functional inhibition on a molecular level. Experiments with paired patient samples showed that these inhibitory effects are markedly related to the immunomagnetically enriched CD34+ leukemic cell population. Using PCR, ELISA, and RNA sequencing, we detected overexpression of TGFß1 in leukemic cells on the transcriptional and protein level and, correspondingly, a molecular signature related to TGFß1 signaling in healthy CD34+ HSPC. This inhibitory effect of TGFß1 on healthy hematopoiesis was functionally corrobated and could be pharmacologically reverted by SD208, an inhibitor of TGFß receptor 1 signaling. Overall, these data indicate that leukemic cells induce functional inhibition of healthy CD34+ HSPC, at least in part, through TGFß1, suggesting that blockage of this pathway may improve hematopoiesis in AML.

Improving the accuracy of prognostication in chronic myelomonocytic leukemia.

INTRODUCTION: Chronic myelomonocytic leukemia (CMML) is a hematological malignancy that is extremely variable regarding its clinical course. It may present either as a chronic disorder with mild symptoms and low disease burden for several years, thereby justifying a watch-and-wait-strategy, or may soon progress to acute myeloid leukemia (AML) leaving allogeneic stem cell transplantation as the only curative treatment option. AREAS COVERED: Attempts have been made to integrate clinical, cytogenetic, and molecular parameters into scoring systems aiming at providing reliable prognostic information. In this article, we discuss several prognostic parameters and validate prognostic scores in a cohort of 645 patients with CMML. EXPERT OPINION: We show that the CPSS (CMML prognostic scoring system) is a useful prognostic tool. The integration of molecular data into the new CPSS-mol will further improve prognostic accuracy, primarily by identifying an increased proportion of higher-risk patients.

Characterization and application of electrically active neuronal networks established from human induced pluripotent stem cell-derived neural progenitor cells for neurotoxicity evaluation.

Neurotoxicity is mediated by a variety of modes-of-actions leading to disturbance of neuronal function. In order to screen larger numbers of compounds for their neurotoxic potential, in vitro functional neuronal networks (NN) might be helpful tools. We established and characterized human NN (hNN) from hiPSC-derived neural progenitor cells by comparing hNN formation with two different differentiation media: in presence (CINDA) and absence (neural differentiation medium (NDM)) of maturation-supporting factors. As a NN control we included differentiating rat NN (rNN) in the study. Gene/protein expression and electrical activity from in vitro developing NN were assessed at multiple time points. Transcriptomes of 5, 14 and 28 days in vitro CINDA-grown hNN were compared to gene expression profiles of in vivo human developing brains. Molecular expression analyses as well as measures of electrical activity indicate that NN mature into neurons of different subtypes and astrocytes over time. In contrast to rNN, hNN are less electrically active within the same period of differentiation time, yet hNN grown in CINDA medium develop higher firing rates than hNN without supplements. Challenge of NN with neuronal receptor stimulators and inhibitors demonstrate presence of inhibitory, GABAergic neurons, whereas glutamatergic responses are limited. hiPSC-derived GABAergic hNN grown in CINDA medium might be a useful tool as part of an in vitro battery for assessing neurotoxicity.

Impact of somatic mutations in myelodysplastic patients with isolated partial or total loss of chromosome 7.

Monosomy 7 [-7] and/or partial loss of chromosome 7 [del(7q)] are associated with poor and intermediate prognosis, respectively, in myelodysplastic syndromes (MDS), but somatic mutations may also play a key complementary role. We analyzed the impact on the outcomes of deep targeted mutational screening in 280 MDS patients with -7/del(7q) as isolated cytogenetic abnormality (86 with del(7q) and 194 with -7). Patients with del(7q) or -7 had similar demographic and disease-related characteristics. Somatic mutations were detected in 79% (93/117) of patients (82% in -7 and 73% in del(7q) group). Median number of mutations per patient was 2 (range 0-8). There was no difference in mutation frequency between the two groups. Patients harbouring ≥2 mutations had a worse outcome than patients with <2 or no mutations (leukaemic transformation at 24 months, 38% and 20%, respectively, p = 0.044). Untreated patients with del(7q) had better overall survival (OS) compared with -7 (median OS, 34 vs 17 months, p = 0.034). In multivariable analysis, blast count, TP53 mutations and number of mutations were independent predictors of OS, whereas the cytogenetic subgroups did not retain prognostic relevance. This study highlights the importance of mutational analysis in terms of prognosis in MDS patients with isolated -7 or del(7q).

Lymphopenia at diagnosis is highly prevalent in myelodysplastic syndromes and has an independent negative prognostic value in IPSS-R-low-risk patients.

Lymphopenia is associated with an increased mortality in several medical conditions. Its prognostic impact in myelodysplastic syndromes (MDS) is less well studied. Hence, we analyzed 1023 patients from the Düsseldorf MDS-registry with regard to the absolute lymphocyte count (ALC) at diagnosis. An ALC below the median of the population (1.2 × 109/l) was associated with lower counts of neutrophils (median 1.35 vs. 1.92 × 109/l, p < 0.001) and platelets (median 100 vs. 138 × 109/l, p < 0.001) and with a significant lower overall survival in univariate analysis (whole cohort: median 36 vs. 46 months, p = 0.016; 721 patients without hematopoietic stem cell transplantation or induction chemotherapy: median 36 vs. 56 months, p = 0.001). For low-risk MDS according to IPSS-R, an ALC < 1.2 × 109/l was of additional prognostic value in a multivariate Cox regression model together with age (< or ≥65 years) and LDH (< or ≥normal value of 240 U/l; HR 1.46, 95% CI: 1.03-2.08, p = 0.033). These data support the hypothesis of subtle but clinical relevant changes of the adaptive immune system in MDS. Further studies are necessary to identify the ALC cut-off best suitable for prognostication and the mechanisms responsible for the impairment of lymphoid homeostasis in MDS.

Comprehensive analysis of isolated der(1;7)(q10;p10) in a large international homogenous cohort of patients with myelodysplastic syndromes.

The karyotype is a strong independent prognostic factor in myelodysplastic syndromes (MDS). Since the implementation of the new comprehensive cytogenetic scoring system for MDS, chromosome 7 anomalies are no longer generally assigned to poor risk features but are thoroughly separated. However, der(1;7)(q10;p10), hereinafter der(1;7), is merged into the group labeled "any other single" and belongs to the intermediate risk group, just by definition due to lack of adequate clinical data. The aim of our international collaborative was to clarify the "real" prognostic impact of der(1;7) on a homogenous and well-documented data base. We performed detailed analysis of 63 MDS patients with isolated der(1;7) constituting the largest cohort hitherto reported. Furthermore, clinical data are compared with those of patients with isolated del(7q) and isolated monosomy 7. Median overall survival (OS) of patients with der(1;7) is 26 months (hazard ratio (HR) 0.91 for del(7q) vs der(1;7) and 2.53 for monosomy 7 vs der(1;7)). The der(1;7) is associated with profound thrombocytopenia most probably causing the reduced OS which is in striking contrast to the low risk for AML transformation (HR 3.89 for del(7q) vs der(1;7) and 5.88 for monosomy 7 vs der(1;7)). Molecular karyotyping indicates that der(1;7) is generated in a single step during mitosis and that a chromosomal imbalance rather than a single disrupted gene accounts for malignancy. Thus, the current cytogenetic scoring system assigning isolated der(1;7) to the intermediate risk group is now confirmed by a sufficient data set.

Clonal architecture in patients with myelodysplastic syndromes and double or minor complex abnormalities: Detailed analysis of clonal composition, involved abnormalities, and prognostic significance.

The study analyzes the clonal architecture and the abnormalities involved in a series of 191 patients with myelodysplastic syndromes (MDS) and 2-3 clonal abnormalities. All patients were extracted from an international database. The patients were classified into six clonal subtypes (2A-3C) based on the number of abnormalities and the presentation of unrelated clones (UC) and/or a clonal evolution. UC were detected in 23/191 patients (12%). The composition of UC showed great variability. The only recurrent combination of abnormalities was del(5q) and + 8 in 8 of 23 patients (35%). In patients with clonal evolution, the clone size of the primary and secondary clone varied: Patients with -7 and + 8 in the primary clone showed a larger primary and a smaller secondary clone (-7: median 74% vs 10%; +8 73% vs 18%) while patients with del(5q) in the primary clone showed a smaller primary and a larger secondary clone (33% vs 61%). Univariate and multivariate analyses showed no significant differences regarding overall or AML-free survival between the clonal subtypes. Only the subtype 3C (3 abnormalities and clonal evolution) was an independent risk factor for developing AML (Hazard Ratio 5.5 as compared to subtype 2A, P < .05). Finally, our study confirms that the number of abnormalities clearly defines a significant risk factor for overall- as well as AML-free survival. Importantly, in patients with more than one clone, the calculation of the number of abnormalities in the entire sample instead of the number of abnormalities per clone allows a higher prognostic accuracy.

Long-term outcome of high risk patients with myelodysplastic syndromes or secondary acute myeloid leukemia receiving intensive chemotherapy.

Intensive chemotherapy (IC) used to be a common treatment approach for patients with higher-risk myelodysplastic syndromes (MDS) or acute myeloid leukemia after MDS (sAML). We conducted a retrospective analysis of 299 patients, including a matched pair analysis comparing 96 patients receiving IC with 96 patients not undergoing IC, in order to evaluate the impact of IC on overall survival (OS) and to identify factors that influence remission rates and OS. Complete remission (CR) after first induction chemotherapy was reached in 50% of patients. Parameters influencing the probability of achieving CR were blast count in the bone marrow (< 30%), age < 65 years, presence of Auer rods, duration of antecedent MDS shorter than 6 months, and timing of IC in relation to first diagnosis. The difference in survival time was not significantly better for patients receiving IC (median OS 12.7 months vs. 7 months). Parameters favorably influencing survival were the presence of Auer rods, age below 60 years, blast count below 30%, IC given shortly after first diagnosis, and achievement of CR. On multivariate analysis, achieving CR, presence of Auer rods, and percentage of blasts below or above 30% significantly influenced median survival. Relapse occurred in 63% of patients after a median of 9.9 months with a median survival of 7.6 months. Considering the high relapse rate and short survival, we conclude that intensive chemotherapy is not promising for high-risk MDS or sAML.