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.

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.

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.

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.

Wilms' Tumor 1 Gene Expression Using a Standardized European LeukemiaNet-Certified Assay Compared to Other Methods for Detection of Minimal Residual Disease in Myelodysplastic Syndrome and Acute Myelogenous Leukemia after Allogeneic Blood Stem Cell Transplantation.

Overexpression of the Wilms' tumor 1 (WT1) gene is informative in many patients with acute myelogenous leukemia (AML) and myelodysplastic syndromes (MDS) and is measurable in peripheral blood (PB). Despite these advantages, WT1 has not broadly been established as a marker for minimal residual disease (MRD) monitoring after allogeneic hematopoietic stem cell transplantation (allo-HSCT) due to limited patient numbers, differing sample sources, and nonstandardized in-house methods. To estimate the value of WT1 as an MRD marker, we serially quantified PB WT1 expression using a standardized European LeukemiaNet-certified assay in 59 patients with AML and MDS after allo-HSCT. We compared its performance with routine methods such as chimerism, XY-fluorescence in situ hybridization (FISH), disease-specific cytogenetic, and molecular analyses, which were accessible in 100%, 34%, 68%, and 37%, respectively. Twenty-four patients (41%) relapsed within a median of 126 days after allo-HSCT, and 20 of them showed at least 1 elevated WT1 value above the validated cutoff. The other 35 patients (59%) remained in complete remission, and only 1 patient had a transient increase in WT1 expression. This reflects a sensitivity of 83% and a specificity of 97% for WT1 and appears to be favorable compared with the sensitivities and specificities observed for chimerism (33% and 91%), XY-FISH (67% and 73%), cytogenetic (33% and 77%), and molecular (78% and 85%) analyses. Further supporting its predictive impact, elevated WT1 expression prompted an earlier BM biopsy and consecutively the diagnosis of relapse in 62% of patients. The results of this real-life experience imply that PB WT1 expression is measurable by a standardized assay and predicts imminent relapse after allo-HSCT with high sensitivity and specificity in most patients with AML and MDS.

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.

Cytogenetic clonal evolution in myelodysplastic syndromes is associated with inferior prognosis.

BACKGROUND: The karyotype of bone marrow cells at the time of diagnosis is a strong prognostic parameter for overall survival as well as acute myeloid leukemia (AML) progression in patients with myelodysplastic syndromes (MDS). However, to the authors' knowledge, few data exist regarding the prognostic impact of cytogenetic clonal evolution during the course of MDS. METHODS: The authors evaluated follow-up karyotype analyses in 549 patients from the Dusseldorf MDS Registry. RESULTS: Clonal evolution was detectable in 24% of the entire cohort and in 18% of 294 patients receiving best supportive care. The authors noted a clear adverse effect of clonal evolution on the risk of leukemic transformation (hazard ratio, 2.233; P = .036) and overall survival (hazard ratio, 3.677; P<.001). The authors also analyzed the prognostic influence of subclones detectable at the time of diagnosis. Again, such a finding was associated with a significantly shorter overall survival and a higher 5-year-probability of acute myeloid leukemia progression (30% vs 22%). CONCLUSIONS: The results of the current study support the belief that follow-up karyotype analyses should be performed, especially in patients with lower-risk and intermediate-risk MDS, to identify those patients who are at higher risk of disease progression and therefore might benefit from earlier or more intensive treatment. Cancer 2017;123:4608-4616. © 2017 American Cancer Society.

Causes of death in 2877 patients with myelodysplastic syndromes.

Patients with myelodysplastic syndromes face a poor prognosis. The exact causes of death have not been described properly in the past. We performed a retrospective analysis of causes of death using data of 3792 patients in the Düsseldorf registry who have been followed up for a median time of 21 months. Medical files as well as death certificates were screened and primary care physicians were contacted. Death after AML evolution, infection, and bleeding was considered to be clearly disease-related. Further categories of causes of death were heart failure, other possibly disease-related reasons, such as hemochromatosis, disease-independent reasons as well as cases with unclear causes of death. Median age at the time of diagnosis was 71 years. At the time of analysis, 2877 patients (75.9 %) had deceased. In 1212 cases (42.1 %), the exact cause of death could not be ascertained. From 1665 patients with a clearly documented cause of death, 1388 patients (83.4 %) succumbed directly disease-related (AML (46.6 %), infection (27.0 %), bleeding (9.8 %)), whereas 277 patients (16.6 %) died for reasons not directly related with myelodysplastic syndromes (MDS), including 132 patients with cardiac failure, 77 non-disease-related reasons, 23 patients with solid tumors, and 45 patients with possibly disease-related causes like hemochromatosis. Correlation with IPSS, IPSS-R, and WPSS categories showed a proportional increase of disease-related causes of death with increasing IPSS/IPSS-R/WPSS risk category. Likewise, therapy-related MDS were associated with a higher percentage of disease-related causes of death than primary MDS. This reflects the increasing influence of the underlying disease on the cause of death with increasing aggressiveness of the disease.

Frequency of del(12p) is commonly underestimated in myelodysplastic syndromes: Results from a German diagnostic study in comparison with an international control group.

In myelodysplastic syndromes (MDS), deletion of the short arm of chromosome 12 (del(12p)) is usually a small abnormality, rarely detected as a single aberration by chromosome banding analysis (CBA) of bone marrow metaphases. Del(12p) has been described in 0.6 to 5% of MDS patients at initial diagnosis and is associated with a good to intermediate prognosis as a sole anomaly according to current scoring systems. Here, we present the results of a systematic del(12p) testing in a German prospective diagnostic study (clinicaltrials.gov: NCT01355913) on 367 MDS patients in whom CD34+ peripheral blood cells were analysed for the presence of del(12p) by sequential fluorescence in situ hybridization (FISH) analyses. A cohort of 2,902 previously published MDS patients diagnosed by CBA served as control. We demonstrate that, using a sensitive FISH technique, 12p deletion occurs significantly more frequently in MDS than previously described (7.6% by CD34+ PB-FISH vs. 1.6% by CBA, P < 0.001) and is often associated with other aberrations (93% by CD34+ PB-FISH vs. 60% by CBA). Additionally, the detection rate can be increased by repeated analyses in a patient over time which is important for the patient´s prognosis to distinguish a sole anomaly from double or complex aberrations. To our knowledge, this is the first study to screen for 12p deletions with a suitable probe for ETV6/TEL in 12p13. Our data suggest that the supplement of a probe for the detection of a 12p deletion to common FISH probe panels helps to avoid missing a del(12p), especially as part of more complex aberrations.

Development and validation of a prognostic scoring system for patients with chronic myelomonocytic leukemia.

The natural course of chronic myelomonocytic leukemia (CMML) is highly variable but a widely accepted prognostic scoring system for patients with CMML is not available. The main aim of this study was to develop a new CMML-specific prognostic scoring system (CPSS) in a large series of 558 patients with CMML (training cohort, Spanish Group of Myelodysplastic Syndromes) and to validate it in an independent series of 274 patients (validation cohort, Heinrich Heine University Hospital, Düsseldorf, Germany, and San Matteo Hospital, Pavia, Italy). The most relevant variables for overall survival (OS) and evolution to acute myeloblastic leukemia (AML) were FAB and WHO CMML subtypes, CMML-specific cytogenetic risk classification, and red blood cell (RBC) transfusion dependency. CPSS was able to segregate patients into 4 clearly different risk groups for OS (P < .001) and risk of AML evolution (P < .001) and its predictive capability was confirmed in the validation cohort. An alternative CPSS with hemoglobin instead of RBC transfusion dependency offered almost identical prognostic capability. This study confirms the prognostic impact of FAB and WHO subtypes, recognizes the importance of RBC transfusion dependency and cytogenetics, and offers a simple and powerful CPSS for accurately assessing prognosis and planning therapy in patients with CMML.

Validation of cytogenetic risk groups according to International Prognostic Scoring Systems by peripheral blood CD34+FISH: results from a German diagnostic study in comparison with an international control group.

International Prognostic Scoring Systems are used to determine the individual risk profile of myelodysplastic syndrome patients. For the assessment of International Prognostic Scoring Systems, an adequate chromosome banding analysis of the bone marrow is essential. Cytogenetic information is not available for a substantial number of patients (5%-20%) with dry marrow or an insufficient number of metaphase cells. For these patients, a valid risk classification is impossible. In the study presented here, the International Prognostic Scoring Systems were validated based on fluorescence in situ hybridization analyses using extended probe panels applied to cluster of differentiation 34 positive (CD34(+)) peripheral blood cells of 328 MDS patients of our prospective multicenter German diagnostic study and compared to chromosome banding results of 2902 previously published patients with myelodysplastic syndromes. For cytogenetic risk classification by fluorescence in situ hybridization analyses of CD34(+) peripheral blood cells, the groups differed significantly for overall and leukemia-free survival by uni- and multivariate analyses without discrepancies between treated and untreated patients. Including cytogenetic data of fluorescence in situ hybridization analyses of peripheral CD34(+) blood cells (instead of bone marrow banding analysis) into the complete International Prognostic Scoring System assessment, the prognostic risk groups separated significantly for overall and leukemia-free survival. Our data show that a reliable stratification to the risk groups of the International Prognostic Scoring Systems is possible from peripheral blood in patients with missing chromosome banding analysis by using a comprehensive probe panel (clinicaltrials.gov identifier:01355913).

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.

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.

User expectations and experiences of an assistive robotic arm in amyotrophic lateral sclerosis: a multicenter observational study.

OBJECTIVE: Robotic arms are innovative assistive devices for ALS patients with progressive motor deficits of arms and hands. The objective was to explore the patients´ expectations towards a robotic arm system and to assess the actual experiences after the provision of the device. METHODS: A prospective observational study was conducted at 9 ALS centers in Germany. ALS-related functional deficits were assessed using the ALS-Functional Rating Scale-revised (ALSFRS-R). Motor deficit of the upper limbs was determined using a subscore of three arm-related items of the ALSFRS-R (items 4-6; range 0-12 points). User expectations before provision (expectation group, n = 85) and user experiences after provision (experience group, n = 14) with the device (JACO Assistive Robotic Device, Kinova, Boisbriand, QC, Canada) were assessed. RESULTS: In the total cohort, mean ALSFRS-R subscore for arm function was 1.7 (SD: 2.0, 0-9) demonstrating a severe functional deficit of the upper limbs. In the expectation group (n = 85), the following use cases of the robotic arm have been prioritized: handling objects (89%), close-body movements (88%), pressing buttons (87%), serving drinks (86%), and opening cabinets and doors (85%). In the experience group (n = 14), handling objects (79%), serving drinks (79%), near-body movements (71%), pushing buttons (71%), serving food (64%), and opening doors (64%) were the most frequent used cases. Most patients used the device daily (71.4%, n = 10), and 28.6% (n = 4) several times a week. All patients of the experience group found the device helpful, felt safe while using the device, and were satisfied with its reliability. NPS of the assistive robotic arm revealed 64% "promoters" (strong recommendation), 29% "indifferents" (uncertain recommendation) and 7% "detractors" (no recommendation). Total NPS was + 57 demonstrating strong patient satisfaction. CONCLUSIONS: Initiation of procurement with a robotic assistive arm was confined to patients with severe functional deficit of the upper limbs. User experience underlined the wide spectrum of use cases of assistive robotic arms in ALS. The positive user experience together with high satisfaction underscore that robotic arm systems serve as a valuable treatment option in ALS patients with severe motor deficits of the arms.

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.

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.

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.

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.

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.