Publications by authors named "Dirk Reinhardt"

177 Publications

Advisory opinion of the AWMF Ad hoc Commission In-vitro Diagnostic Medical Devices regarding in-vitro diagnostic medical devices manufactured and used only within health institutions established in the Union according to Regulation (EU) 2017/746 (IVDR).

Ger Med Sci 2021 1;19:Doc08. Epub 2021 Jun 1.

Deutsche Gesellschaft für Pharmazeutische Medizin (DGPharMed).

In view of the approaching application date of Regulation (EU) 2017/746 ("IVDR") and the resulting EU-wide, harmonized requirements for in-vitro diagnostic medical devices (IVD) manufactured and used within European health institutions, the Ad hoc Commission IVD of the German Association of the Scientific Medical Societies (AWMF) takes a national position on the details of the requirements and conditions related to the use of these IVD products. The Ad hoc Commission IVD emphasizes the relevance of examination procedures developed in medical laboratories, especially in the field of orphan diseases and new diagnostic markers. The IVDR sets an adequate regulatory framework for IVD manufactured and used within health institutions as long as these requirements are fulfilled with reliability and in accordance with the current state of the art in medical laboratory sciences. At the same time, the IVDR requirements have to be regarded under a pragmatic view and in accordance with the quality management systems approved within the different EU Member States. On the one hand, the mandatory requirements of the RiLiBÄK play an essential role in Germany. On the other hand, elements of voluntarily applicable international standards may support the fulfilment of product requirements for safety and performance according to Annex I of the IVDR. Both the complexity and possible solutions for the implementation of the IVDR requirements are discussed on the basis of examples such as the required documentation, performance evaluation and software validation. The Ad hoc Commission IVD recommends that, while aiming at a preferably EU-wide harmonized interpretation of the IVDR requirements, the flexibility in medical laboratory diagnostics necessary for patient care, including the use of IVD from in-house production, should be emphasized.
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http://dx.doi.org/10.3205/000295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8204380PMC
June 2021

A homozygous nonsense mutation early in exon 5 of BRCA2 is associated with very severe Fanconi anemia.

Eur J Med Genet 2021 Aug 10;64(8):104260. Epub 2021 Jun 10.

Department of Pediatrics III, University Hospital Essen, University of Duisburg-Essen, 45122 Essen, Germany; Department of Otorhinolaryngology & Head/Neck Surgery, University Hospital Düsseldorf, Heinrich Heine University, 40225 Düsseldorf, Germany. Electronic address:

Fanconi anemia (FA) due to biallelic mutations in the BRCA2 gene is very rare and associated with an extremely high risk of early-onset of aggressive childhood malignancies, predominantly brain tumors, leukemia, and nephroblastoma. Here, we present a consanguineous family with three affected children of the D1 subtype of FA and describe the clinical consequences of the earliest known biallelic nonsense/stop-gain germ-line mutation in BRCA2, exon 5 c.469A>T, that leads to a premature stop of translation, p.Lys157*. The three patients were born with severe intrauterine growth restrictions and different degrees of congenital malformations. Altogether, they developed eight distinct malignancies and died within their first three years of life. Treatment with a reduced chemotherapy regimen was only performed in patient 2 for his first tumor, a nephroblastoma, which the patient tolerated well for eight months, until he developed myelodysplastic syndrome (MDS) and then acute myeloid leukemia (AML). Finally, the third patient experienced a hepatoblastoma, an unclassified brain tumor and MDS in parallel and died in her second year of life. Our report re-emphasizes the aggressiveness and fatality of the FA-D1 children with biallelic BRCA2 nonsense mutations, that are both located before exon 11, which contains binding domains for the RAD51 recombinase.
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http://dx.doi.org/10.1016/j.ejmg.2021.104260DOI Listing
August 2021

Far from Health: The Bone Marrow Microenvironment in AML, A Leukemia Supportive Shelter.

Children (Basel) 2021 May 8;8(5). Epub 2021 May 8.

Department of Pediatric Hematology and Oncology, Clinic of Pediatrics III, Essen University Hospital, 45147 Essen, Germany.

Acute myeloid leukemia (AML) is the second most common leukemia among children. Although significant progress in AML therapy has been achieved, treatment failure is still associated with poor prognosis, emphasizing the need for novel, innovative therapeutic approaches. To address this major obstacle, extensive knowledge about leukemogenesis and the complex interplay between leukemic cells and their microenvironment is required. The tremendous role of this bone marrow microenvironment in providing a supportive and protective shelter for leukemic cells, leading to disease development, progression, and relapse, has been emphasized by recent research. It has been revealed that the interplay between leukemic cells and surrounding cellular as well as non-cellular components is critical in the process of leukemogenesis. In this review, we provide a comprehensive overview of recently gained knowledge about the importance of the microenvironment in AML whilst focusing on promising future therapeutic targets. In this context, we describe ongoing clinical trials and future challenges for the development of targeted therapies for AML.
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http://dx.doi.org/10.3390/children8050371DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8150304PMC
May 2021

Survival Following Relapse in Children with Acute Myeloid Leukemia: A Report from AML-BFM and COG.

Cancers (Basel) 2021 May 12;13(10). Epub 2021 May 12.

Department of Pediatric Hematology-Oncology, Pediatrics III, University Hospital of Essen, 45147 Essen, Germany.

Post-relapse therapy remains critical for survival in children with acute myeloid leukemia (AML). We evaluated survival, response and prognostic variables following relapse in independent cooperative group studies conducted by COG and the population-based AML-BFM study group. BFM included 197 patients who relapsed after closure of the last I-BFM relapse trial until 2017, while COG included 852 patients who relapsed on the last Phase 3 trials (AAML0531, AAML1031). Overall survival at 5 years (OS) was 42 ± 4% (BFM) and 35 ± 2% (COG). Initial high-risk features (BFM 32 ± 6%, COG 26 ± 4%) and short time to relapse (BFM 29 ± 4%, COG 25 ± 2%) predicted diminished survival. In the BFM dataset, there was no difference in OS for patients who had a complete remission with full hematopoietic recovery (CR) following post-relapse re-induction compared to those with partial neutrophil and platelet recovery (CRp and CRi) only (52 ± 7% vs. 63 ± 10%, = 0.39). Among 90 patients alive at last follow-up, 87 had received a post-relapse hematopoietic stem cell transplant (HSCT). OS for patients with post-relapse HSCT was 54 ± 4%. In conclusion, initial high-risk features and early relapse remain prognostic. Response assessment with full hematopoietic recovery following initial relapse therapy does not predict survival. These data indicate the need for post-relapse risk stratification in future studies of relapse therapies.
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http://dx.doi.org/10.3390/cancers13102336DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8151466PMC
May 2021

A Study of Regulatory Challenges of Pediatric Oncology Phase I/II Trial Submissions and Guidance on Protocol Development.

Clin Pharmacol Ther 2021 May 29. Epub 2021 May 29.

Hopp Children's Cancer Center Heidelberg (KiTZ), Heidelberg, Germany.

The purpose of this study was to identify key deficiencies in pediatric oncology early phase clinical trial protocols in Germany and to provide guidance for efficient trial protocol development. A systematic review of the response letters of German competent authorities (CAs) and Ethics Committees to phase I/II pediatric oncology trial submissions in the period from 2014 to 2019 was performed. Documents were requested from all five Society for Paediatric Oncology and Haematology in Germany (GPOH) phase I/II trial networks plus all nine German Innovative Therapies for Children with Consortium Cancer (ITCC) centers. A blinded dataset containing aggregated data from 33 studies was analyzed for validation. All deficiencies were reviewed, listed, and weighted using a structured matrix according to frequency, category, significance, and feasibility. In total, documents of 17 trials from 6 different sites were collected. Two hundred fifty deficiencies identified by the CAs were identified and categorized into eight categories. "Toxicity and safety" was the most prominent category (27.6%), followed by "Manufacturing and Import" (18%). The majority of deficiencies were categorized as minor and potential measures as easy to address, but an important group of major and difficult to implement deficiencies was also identified. The blinded validation dataset confirmed these findings. The majority of the EC deficiencies could be resolved by changing the wording in the patient-facing documents. In conclusion, this study was able to detect a pattern of key deficiencies. Most of the shortcomings can be anticipated by minor changes in the protocol and increased awareness can prevent time-consuming revisions, withdrawals, or even rejections. A corresponding guideline describing key regulatory aspects is provided.
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http://dx.doi.org/10.1002/cpt.2319DOI Listing
May 2021

Eye Tumors in Childhood as First Sign of Tumor Predisposition Syndromes: Insights from an Observational Study Conducted in Germany and Austria.

Cancers (Basel) 2021 Apr 14;13(8). Epub 2021 Apr 14.

Institute of Human Genetics, Medical Faculty, University Duisburg-Essen, 45122 Essen, Germany.

Retinoblastoma and other eye tumors in childhood are rare diseases. Many eye tumors are the first signs of a genetic tumor predisposition syndrome and the affected children carry a higher risk of developing other cancers later in life. Clinical and genetic data of all children with eye tumors diagnosed between 2013-2018 in Germany and Austria were collected in a multicenter prospective observational study. In five years, 300 children were recruited into the study: 287 with retinoblastoma, 7 uveal melanoma, 3 ciliary body medulloepithelioma, 2 retinal astrocytoma, 1 meningioma of the optic nerve extending into the eye. Heritable retinoblastoma was diagnosed in 44% of children with retinoblastoma. One child with meningioma of the optic nerve extending into the eye was diagnosed with neurofibromatosis 2. No pathogenic constitutional variant in was detected in a child with medulloepithelioma while two children did not receive genetic analysis. Because of the known association with tumor predisposition syndromes, genetic counseling should be offered to all children with eye tumors. Children with a genetic predisposition to cancer should receive a tailored surveillance including detailed history, physical examinations and, if indicated, imaging to screen for other cancer. Early detection of cancers may reduce mortality.
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http://dx.doi.org/10.3390/cancers13081876DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8070790PMC
April 2021

Redirecting the Immune Microenvironment in Acute Myeloid Leukemia.

Cancers (Basel) 2021 Mar 20;13(6). Epub 2021 Mar 20.

Department of Pediatric Hematology and Oncology, Clinic of Pediatrics III, University Hospital Essen, 45147 Essen, Germany.

Acute myeloid leukemia is a life-threatening malignant disorder arising in a complex and dysregulated microenvironment that, in part, promotes the leukemogenesis. Treatment of relapsed and refractory AML, despite the current overall success rates in management of pediatric AML, remains a challenge with limited options considering the heavy but unsuccessful pretreatments in these patients. For relapsed/refractory (R/R) patients, hematopoietic stem cell transplantation (HSCT) following ablative chemotherapy presents the only opportunity to cure AML. Even though in some cases immune-mediated graft-versus-leukemia (GvL) effect has been proven to efficiently eradicate leukemic blasts, the immune- and chemotherapy-related toxicities and adverse effects considerably restrict the feasibility and therapeutic power. Thus, immunotherapy presents a potent tool against acute leukemia but needs to be engineered to function more specifically and with decreased toxicity. To identify innovative immunotherapeutic approaches, sound knowledge concerning immune-evasive strategies of AML blasts and the clinical impact of an immune-privileged microenvironment is indispensable. Based on our knowledge to date, several promising immunotherapies are under clinical evaluation and further innovative approaches are on their way. In this review, we first focus on immunological dysregulations contributing to leukemogenesis and progression in AML. Second, we highlight the most promising therapeutic targets for redirecting the leukemic immunosuppressive microenvironment into a highly immunogenic environment again capable of anti-leukemic immune surveillance.
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http://dx.doi.org/10.3390/cancers13061423DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8003817PMC
March 2021

Second Relapse of Pediatric Patients with Acute Myeloid Leukemia: A Report on Current Treatment Strategies and Outcome of the AML-BFM Study Group.

Cancers (Basel) 2021 Feb 14;13(4). Epub 2021 Feb 14.

Department of Pediatric Hematology-Oncology, Pediatrics III, University Hospital of Essen, 45147 Essen, Germany.

Successful management of relapse is critical to improve outcomes of children with acute myeloid leukemia (AML). We evaluated response, survival and prognostic factors after a second relapse of AML. Among 1222 pediatric patients of the population-based AML-Berlin-Frankfurt-Munster (BFM) study group (2004 until 2017), 73 patients met the quality parameters for inclusion in this study. Central review of source documentation warranted the accuracy of reported data. Treatment approaches included palliation in 17 patients (23%), intensive therapy with curative intent ( 46, 63%) and other regimens ( 10). Twenty-five patients (35%) received hematopoietic stem cell transplantation (HSCT), 21 of whom (88%) had a prior HSCT. Survival was poor, with a five-year probability of overall survival (pOS) of 15 ± 4% and 31 ± 9% following HSCT ( 25). Early second relapse (within one year after first relapse) was associated with dismal outcome (pOS 2 ± 2%, 44 vs. 33 ± 9%, 29; < 0.0001). A third complete remission (CR) is required for survival: 31% ( 14) of patients with intensive treatment achieved a third CR with a pOS of 36 ± 13%, while 28 patients (62%) were non-responders (pOS 7 ± 5%). In conclusion, survival is poor but possible, particularly after a late second relapse and an intensive chemotherapy followed by HSCT. This analysis provides a baseline for future treatment planning.
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http://dx.doi.org/10.3390/cancers13040789DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7918758PMC
February 2021

BCR-ABL1 positive AML or CML in blast crisis? A pediatric case report with inv(3) and t(9;22) in the initial clone.

Cancer Genet 2021 Jun 19;254-255:70-74. Epub 2021 Feb 19.

Department of Human Genetics, Hannover Medical School, Carl-Neuberg-Str.1, 30625 Hannover, Germany. Electronic address:

The co-occurrence of an inversion inv(3)(q21q26)/GATA2-MECOM and a Philadelphia translocation t(9;22)(q34;q11)/BCR-ABL1 in the context of chronic myeloid leukemia (CML) in blast crisis or acute myeloid leukemia (AML) has only rarely been described. To our knowledge, this co-occurrence has been reported in six pediatric patients with CML but not in pediatric patients with AML. Here, we report on a 7-year-old girl, who, presented with a t(9;22) and inv(3) in 14 of 15 metaphases and an additional monosomy 7 was detected in 5 of these metaphases (ISCN: 46,​XX,​inv(3)(q21q26),​t(9;22)(q34q11)[9]/45,​idem,​-7[5]/46,​XX[1]). The p190 BCR-ABL1 fusion transcript was detected by multiplex PCR and targeted RNA sequencing. Due to these results, a clear distinction between a CML in blast crisis and a BCR-ABL1 positive AML was not possible. The patient was treated according to the treatment recommendations of the AML-BFM study group and additionally received tyrosine kinase inhibitor therapy (Dasatinib). The treatment with Dasatinib was successful in eliminating the inv(3)/t(9;22) clone, but the ancestral inv(3) clone persisted. Based upon these findings we diagnosed an AML with inv(3) and a secondary acquisition of t(9;22). This treatment as well as an allogenic transplantation has led to a complete remission of the disease up to this date (21 months post diagnosis).
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http://dx.doi.org/10.1016/j.cancergen.2021.02.007DOI Listing
June 2021

Paediatric Strategy Forum for medicinal product development for acute myeloid leukaemia in children and adolescents: ACCELERATE in collaboration with the European Medicines Agency with participation of the Food and Drug Administration.

Eur J Cancer 2020 09 17;136:116-129. Epub 2020 Jul 17.

FORMA Therapeutics, USA.

Purpose: The current standard-of-care for front-line therapy for acute myeloid leukaemia (AML) results in short-term and long-term toxicity, but still approximately 40% of children relapse. Therefore, there is a major need to accelerate the evaluation of innovative medicines, yet drug development continues to be adult-focused. Furthermore, the large number of competing agents in rare patient populations requires coordinated prioritisation, within the global regulatory framework and cooperative group initiatives.

Methods: The fourth multi-stakeholder Paediatric Strategy Forum focused on AML in children and adolescents.

Results: CD123 is a high priority target and the paediatric development should be accelerated as a proof-of-concept. Efforts must be coordinated, however, as there are a limited number of studies that can be delivered. Studies of FLT3 inhibitors in agreed paediatric investigation plans present challenges to be completed because they require enrolment of a larger number of patients than actually exist. A consensus was developed by industry and academia of optimised clinical trials. For AML with rare mutations that are more frequent in adolescents than in children, adult trials should enrol adolescents and when scientifically justified, efficacy data could be extrapolated. Methodologies and definitions of minimal residual disease need to be standardised internationally and validated as a new response criterion. Industry supported, academic sponsored platform trials could identify products to be further developed. The Leukaemia and Lymphoma Society PedAL/EUpAL initiative has the potential to be a major advance in the field.

Conclusion: These initiatives continue to accelerate drug development for children with AML and ultimately improve clinical outcomes.
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http://dx.doi.org/10.1016/j.ejca.2020.04.038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7789799PMC
September 2020

Exposure of Patient-Derived Mesenchymal Stromal Cells to TGFB1 Supports Fibrosis Induction in a Pediatric Acute Megakaryoblastic Leukemia Model.

Mol Cancer Res 2020 10 8;18(10):1603-1612. Epub 2020 Jul 8.

Department of Pediatric Hematology and Oncology, University Children's Hospital Essen, Essen, Germany.

Bone marrow fibrosis (BMF) is a rare complication in acute leukemia. In pediatrics, it predominantly occurs in acute megakaryoblastic leukemia (AMKL) and especially in patients with trisomy 21, called myeloid leukemia in Down syndrome (ML-DS). Defects in mesenchymal stromal cells (MSC) and cytokines specifically released by the myeloid blasts are thought to be the main drivers of fibrosis in the bone marrow niche (BMN). To model the BMN of pediatric patients with AMKL in mice, we first established MSCs from pediatric patients with AMKL ( = 5) and ML-DS ( = 9). Healthy donor control MSCs ( = 6) were generated from unaffected children and adolescents ≤18 years of age. Steady-state analyses of the MSCs revealed that patient-derived MSCs exhibited decreased adipogenic differentiation potential and enrichment of proliferation-associated genes. Importantly, TGFB1 exposure promoted early profibrotic changes in all three MSC entities. To study BMF induction for longer periods of time, we created an humanized artificial BMN subcutaneously in immunodeficient NOD.Cg-Prkdc Il2rg/SzJ mice, using a mixture of MSCs, human umbilical vein endothelial cell, and Matrigel. Injection of AMKL blasts as producers of TGFB1 into this BMN after 8 weeks induced fibrosis grade I/II in a dose-dependent fashion over a time period of 4 weeks. Thus, our study developed a humanized mouse model that will be instrumental to specifically examine leukemogenesis and therapeutic targets for AMKL blasts in future. IMPLICATIONS: TGFB1 supports fibrosis induction in a pediatric AMKL model generated with patient-derived MSCs. VISUAL OVERVIEW: http://mcr.aacrjournals.org/content/molcanres/18/10/1603/F1.large.jpg.
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http://dx.doi.org/10.1158/1541-7786.MCR-20-0091DOI Listing
October 2020

Veno-Venous Extracorporeal Membrane Oxygenation in Adult Patients with Sickle Cell Disease and Acute Chest Syndrome: a Single-Center Experience.

Hemoglobin 2020 Mar 14;44(2):71-77. Epub 2020 Apr 14.

Department of Bone Marrow Transplantation, West German Cancer Center, University of Duisburg-Essen, University Hospital Essen, Essen, Germany.

Acute chest syndrome (ACS) in adult patients with sickle cell disease represents a leading cause of death. It is characterized by a new density on chest X-ray accompanied by fever and/or respiratory symptoms. Currently, 49 adult patients with sickle cell disease are registered at our department. By now, 12 patients (24.5%) suffered from ACS and two patients showed multiple/recurrent (>2) episodes. Death in one patient was related to acute respiratory failure secondary to ACS. In three patients with ACS, invasive mechanical ventilation and subsequent veno-venous extracorporeal membrane oxygenation (VV-ECMO) was mandatory. Veno-venous ECMO was applied within 24 hours upon arrival to the intensive care unit (ICU). All patients were treated aggressively for ACS including exchange transfusions [packed red blood cell (pRBC) units 5-16] maintaining a Hb S threshold of <30.0% in addition to broad-spectrum antibiotics, resulting in a successful outcome following decannulation from VV-ECMO (49 hours, 251 hours, 30 min., and 98 hours, respectively). Limited information is presently available on the use of VV-ECMO in adult patients with sickle cell disease in the context of acute respiratory failure secondary to ACS. The adequate timing of the decision to place ECMO in critically ill adults with sickle cell disease, incapable of being treated by conventional mechanical ventilation secondary to very severe vaso-occlusive crisis (VOC), might further reduce mortality rates while treating the underlying condition.
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http://dx.doi.org/10.1080/03630269.2020.1745827DOI Listing
March 2020

Measurable residual disease assessment by qPCR in peripheral blood is an informative tool for disease surveillance in childhood acute myeloid leukaemia.

Br J Haematol 2020 07 16;190(2):198-208. Epub 2020 Mar 16.

Department of Paediatrics and Adolescent Medicine, Aarhus University Hospital, Aarhus, Denmark.

Serial assessments of measurable (or minimal) residual disease (MRD) by qPCR may identify nascent relapse in children with acute myeloid leukaemia (AML) and enable pre-emptive therapy. We investigated the kinetics and prognostic impact of recurrent fusion transcripts (RUNX1-RUNX1T1, CBFB-MYH11, KMT2A-MLLT3 or KMT2A-ELL) in 774 post-induction samples from bone marrow (BM, 347) and peripheral blood (PB, 427) from 75 children with AML. BM MRD persistence during consolidation did not increase the risk of relapse, and MRD at therapy completion did not correlate to outcome (HR = 0·64/MRD log reduction (CI: 0·32-1·26), P = 0·19). In contrast, 8/8 patients with detectable MRD in PB after first consolidation relapsed. Persistence (n = 4) and shifting from negative to positive (n = 10) in PB during follow-up predicted relapse in 14/14 patients. All 253 PB samples collected during follow-up from 36 patients in continuous complete remission were MRD negative. In core-binding factor AML, persistent low-level MRD positivity in BM during follow-up was frequent but an increment to above 5 × 10 heralded subsequent haematological relapse in 12/12 patients. We demonstrate that MRD monitoring in PB after induction therapy is highly informative and propose an MRD increment above 5 × 10 in PB and BM as a definition of molecular relapse since it always leads to haematological relapse.
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http://dx.doi.org/10.1111/bjh.16560DOI Listing
July 2020

Phospho-Profiling Linking Biology and Clinics in Pediatric Acute Myeloid Leukemia.

Hemasphere 2020 Feb 16;4(1):e312. Epub 2019 Dec 16.

CCRI, Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.

Aberrant activation of key signaling-molecules is a hallmark of acute myeloid leukemia (AML) and may have prognostic and therapeutic implications. AML summarizes several disease entities with a variety of genetic subtypes. A comprehensive model spanning from signal activation patterns in major genetic subtypes of pediatric AML (pedAML) to outcome prediction and pre-clinical response to signaling inhibitors has not yet been provided. We established a high-throughput flow-cytometry based method to assess activation of hallmark phospho-proteins (phospho-flow) in 166 bone-marrow derived pedAML samples under basal and cytokine stimulated conditions. We correlated levels of activated phospho-proteins at diagnosis with relapse incidence in intermediate (IR) and high risk (HR) subtypes. In parallel, we screened a set of signaling inhibitors for their efficacy against primary AML blasts in a flow-cytometry based ex vivo cytotoxicity assay and validated the results in a murine xenograft model. Certain phospho-signal patterns differ between genetic subtypes of pedAML. Some are consistently seen through all AML subtypes such as pSTAT5. In IR/HR subtypes high levels of GM-CSF stimulated pSTAT5 and low levels of unstimulated pJNK correlated with increased relapse risk overall. Combination of GM-CSF/pSTAT5 and basal/pJNK separated three risk groups among IR/HR subtypes. Out of 10 tested signaling inhibitors, midostaurin most effectively affected AML blasts and simultaneously blocked phosphorylation of multiple proteins, including STAT5. In a mouse xenograft model of -rearranged pedAML, midostaurin significantly prolonged disease latency. Our study demonstrates the applicability of phospho-flow for relapse-risk assessment in pedAML, whereas functional phenotype-driven ex vivo testing of signaling inhibitors may allow individualized therapy.
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http://dx.doi.org/10.1097/HS9.0000000000000312DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7000467PMC
February 2020

Insights into the limitations of transient expression systems for the functional study of p53 acetylation site and oncogenic mutants.

Biochem Biophys Res Commun 2020 04 12;524(4):990-995. Epub 2020 Feb 12.

Department of Pediatric Hematology/Oncology, Hannover Medical School, Carl-Neuberg-Straße 1, 30625, Hannover, Germany. Electronic address:

Tumor suppressor protein p53 protects cells against malignant transformation mostly through transcriptional activation. Lysine acetylation is required to mediate activation of p53. The protein displays eight lysine residues and their evolutionary conservation argues for an essential role. The aim of this study was to investigate the significance of individual acetylation sites in mediating p53 functions. Differences in intracellular localization, protein expression levels, and transcriptional activity were investigated by overexpressing acetylation-deficient p53 variants in the colon carcinoma-derived p53 knock-out cell line HCT 116 p53. We found that not all lysine residues are equally capable of promoting p53's functions. Individual amino acid mutations or combinations thereof led to altered p53 expression levels, intracellular distribution, or transcriptional transactivation capacity, as compared to the wild-type protein. However, we observed that the choice of protein tag and expression vector could significantly alter obtained results on certain aspects of p53 function.
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http://dx.doi.org/10.1016/j.bbrc.2020.02.002DOI Listing
April 2020

Outcome of children relapsing after first allogeneic haematopoietic stem cell transplantation for acute myeloid leukaemia: a retrospective I-BFM analysis of 333 children.

Br J Haematol 2020 05 3;189(4):745-750. Epub 2020 Feb 3.

Division of Pediatric Hematology and Oncology, Department of Pediatrics, Universities of Medical University Hannover, Hannover, Germany.

Outcome of 333 children with acute myeloid leukaemia relapsing after a first allogeneic haematopoietic stem cell transplantation was analyzed. Four-year probability of overall survival (4y-pOS) was 14%. 4y-pOS for 122 children receiving a second haematopoietic stem cell transplantation was 31% and 3% for those that did not (P = <0·0001). Achievement of a subsequent remission impacted survival (P = <0·0001). For patients receiving a second transplant survival with or without achieving a subsequent remission was comparable. Graft source (bone marrow vs. peripheral blood stem cells, P = 0·046) and donor choice (matched family vs. matched unrelated donor, P = 0·029) positively impacted survival after relapse. Disease recurrence and non-relapse mortality at four years reached 45% and 22%.
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http://dx.doi.org/10.1111/bjh.16441DOI Listing
May 2020

Social inequalities in the participation and activity of children and adolescents with leukemia, brain tumors, and sarcomas (SUPATEEN): a protocol for a multicenter longitudinal prospective observational study.

BMC Pediatr 2020 01 31;20(1):48. Epub 2020 Jan 31.

Institute of Medical Sociology (IMS), Martin Luther University Halle-Wittenberg, Magdeburger Str. 8, 06112, Halle (Saale), Germany.

Background: About 2000 children and adolescents under the age of 18 are diagnosed with cancer each year in Germany. Because of current medical treatment methods, a high survival rate can be reached for many types of the disease. Nevertheless, patients face a number of long-term effects related to the treatment. As a result, physical and psychological consequences have increasingly become the focus of research in recent years. Social dimensions of health have received little attention in health services research in oncology so far. Yet, there are no robust results that allow an estimation of whether and to what extent the disease and treatment impair the participation of children and adolescents and which factors mediate this effect. Social participation is of great importance especially because interactions with peers and experiences in different areas of life are essential for the development of children and adolescents.

Methods: Data are collected in a longitudinal, prospective, observational multicenter study. For this purpose, all patients and their parents who are being treated for cancer in one of the participating clinics throughout Germany will be interviewed within the first month after diagnosis (t1), after completion of intensive treatment (t2) and half a year after the end of intensive treatment (t3) using standardized questionnaires. Analysis will be done by descriptive and multivariate methods.

Discussion: The results can be used to identify children and adolescents in high-risk situations at an early stage in order to be able to initiate interventions tailored to the needs. Such tailored interventions will finally reduce the risk of impairments in the participation of children and adolescents and increase quality of life.

Trial Registration: ClinicalTrials.gov: NCT04101123.
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http://dx.doi.org/10.1186/s12887-020-1943-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6995139PMC
January 2020

Early deaths from childhood cancer in Germany 1980-2016.

Cancer Epidemiol 2020 04 16;65:101669. Epub 2020 Jan 16.

German Childhood Cancer Registry, Institute for Medical Biostatistics, Epidemiology and Informatics (IMBEI), University Medical Centre of the Johannes Gutenberg University Mainz, 55101, Mainz, Germany. Electronic address:

Background: Even though the survival of childhood cancer has improved over the last decades, there are still children dying shortly after diagnosis. The aim of the study is to add to understanding of the reasons for deaths shortly after date of diagnosis.

Methods: Using data of the population-based German Childhood Cancer Registry (cancer below 15 years of age diagnosed between 1980 and 2016), we compared characteristics of 671 children with cancer who died within 30 days of diagnosis to 53,649 patients with childhood cancer who survived longer. In addition to a descriptive analysis, we used logistic regression with multivariable fractional polynomials to describe the relationship between early death with age at diagnosis and year of diagnosis.

Results: The number of early death cases and the risk of early death have decreased considerably since 1980 (2.6% of study population in 1980-89 to 0.6% in 2010-16). Children under one year of age were at highest early death risk (odds ratio = 4.10, 95% confidence interval: 3.32-5.05 compared to 7-year-old patients). These results are similar to results from other studies. Moreover, children with acute myeloid leukemia and hepatic tumors had a higher early death risk, children with acute lymphoid leukemia a lower risk compared to patients with central nervous system tumors used as a reference group.

Conclusion: Even though the risk for early death has declined overall with advances in diagnosis and therapy, special attention needs to be paid to infants and children with AML and hepatic tumors, who are especially at risk.
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http://dx.doi.org/10.1016/j.canep.2020.101669DOI Listing
April 2020

Evaluation of dsDNA from extracellular vesicles (EVs) in pediatric AML diagnostics.

Ann Hematol 2020 Mar 13;99(3):459-475. Epub 2020 Jan 13.

Department of Pediatric Hematology and Oncology, University Children's Hospital, University Hospital of Essen, Hufelandstrasse 55, 45147, Essen, Germany.

Acute myeloid leukemia (AML) is a heterogeneous malignant disease characterized by a collection of genetic and epigenetic changes. As a consequence, AML can evolve towards more aggressive subtypes during treatment, which require additional therapies to prevent future relapse. As we have previously detected double-stranded DNA (dsDNA) in tumor-derived extracellular vesicles (EVs), in this current study we attempted to evaluate the potential diagnostic applications of AML EV-dsDNA derived from primary bone marrow and peripheral blood plasma samples. EVs from plasma of 29 pediatric AML patients (at initial diagnosis or during treatment) were isolated by ultracentrifugation, after which dsDNA was extracted from obtained EVs and analyzed for leukemia-specific mutations using next generation sequencing (NGS) and GeneScan-based fragment-length analysis. In 18 out of 20 patients, dsDNA harvested from EVs mirrored the (leukemia-specific) mutations found in the genomic DNA obtained from primary leukemia cells. In the nanoparticle tracking analysis (NTA), a decrease in EV numbers was observed in patients after treatment compared with initial diagnosis. Following treatment, in 75 samples out of the 79, these mutations were no longer detectable in EV-dsDNA. In light of our results, we propose the use of leukemia-derived EV-dsDNA as an additional measure for mutational status and, potentially, treatment response in pediatric AML.
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http://dx.doi.org/10.1007/s00277-019-03866-wDOI Listing
March 2020

Hematopoietic stem cell transplantation for children with acute myeloid leukemia-results of the AML SCT-BFM 2007 trial.

Leukemia 2020 02 2;34(2):613-624. Epub 2019 Oct 2.

Department of Pediatrics, Division of Pediatric Hematology and Oncology, University of Düsseldorf, Düsseldorf, Germany.

AML SCT-BFM 2007 was the first hematopoietic stem cell transplantation (HCT) trial in Germany to comply with the European Clinical Trials Directive, and aimed to standardize pediatric HCT for acute myeloid leukemia (AML) across centers in Germany, Austria, and the Czech Republic. Children with high-risk features and a good early response achieving a complete first remission (CR-1) and those in CR-2 after a first relapse were stratified to receive HCT from a matched donor after myeloablative conditioning consisting of busulfan, cyclophosphamide, and melphalan. Four-year EFS and OS were 61 and 70%. Cumulative incidence of relapse (CIR) was 22%. TRM was 15% and correlated with age reaching 9% (SE 3%) in children younger than 12 years and 31% (SE 9%) in older children and adolescents. Children with poorly responding primary disease or relapse were allocated to receive early HCT after a cytoreductive regimen with fludarabine, amsacrine, and cytarabine, followed by reduced intensity conditioning and prophylactic donor lymphocyte infusions. Four-year EFS and OS were 49 and 53%. CIR was 38% and TRM 11%. For patients with primary poor response disease, early use of RIC HCT followed by prophylactic DLI can induce long-term remissions in more than 50% (EFS 46% (SE 9%)).
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http://dx.doi.org/10.1038/s41375-019-0584-8DOI Listing
February 2020

Mutated , and Fusion in Various Combinations Define a Poor Prognostic Group in Pediatric Acute Myeloid Leukemia.

J Oncol 2019 30;2019:1609128. Epub 2019 Jul 30.

Department of Pediatrics III, University Children's Hospital Essen, University of Duisburg-Essen, 45122 Essen, Germany.

Acute myeloid leukemia is a life-threatening malignancy in children and adolescents treated predominantly by risk-adapted intensive chemotherapy that is partly supported by allogeneic stem cell transplantation. Mutations in the gene and fusion are predictors of poor survival outcome/prognosis that frequently occur in combination with internal tandem duplications of the juxta-membrane domain of ( To re-evaluate the effect of these factors in contemporary protocols, 353 patients (<18 years) treated in Germany with AML-BFM treatment protocols between 2004 and 2017 were included. Presence of mutated and in blasts (n=19) resulted in low 3-year event-free survival of 29% and overall survival of 33% compared to rates of 45-63% and 67-87% in patients with only one (only only mutation; n=29) or none of these mutations (n=272). Including and high allelic ratio (AR) of (AR ≥0.4) in the analysis revealed very poor outcomes for patients with co-occurrence of all three factors or any of double combinations. All these patients (n=15) experienced events and the probability of overall survival was low (27%). We conclude that co-occurrence of mutation, and with an AR ≥0.4 as triple or double mutations still predicts dismal response to contemporary first- and second-line treatment for pediatric acute myeloid leukemia.
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http://dx.doi.org/10.1155/2019/1609128DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6699323PMC
July 2019

Genotyping circulating tumor DNA of pediatric Hodgkin lymphoma.

Leukemia 2020 01 20;34(1):151-166. Epub 2019 Aug 20.

General Pediatrics, Oncology and Hematology, Vestische Kinder und Jugendklinik Datteln, Witten/Herdecke University, Witten, Germany.

We used hybrid capture-targeted next-generation sequencing of circulating cell-free DNA (ccfDNA) of pediatric Hodgkin lymphoma (PHL) patients to determine pathogenic mechanisms and assess the clinical utility of this method. Hodgkin-Reed/Sternberg (HRS) cell-derived single nucleotide variants, insertions/deletions, translocations and VH-DH-JH rearrangements were detected in pretherapy ccfDNA of 72 of 96 patients. Number of variants per patient ranged from 1 to 21 with allele frequencies from 0.6 to 42%. Nine translocation breakpoints were detected. Genes involved in JAK/STAT, NFkB and PI3K signaling and antigen presentation were most frequently affected. SOCS1 variants, mainly deletions, were found in most circulating tumor (ct) DNAs, and seven of the nine translocation breakpoints involved SOCS1. Analysis of VH-DH-JH rearrangements revealed an origin of PHL HRS cells from partially selected germinal center B cells. Amounts of pretherapy ctDNA were correlated with metabolic tumor volumes. Furthermore, in all ccfDNA samples of 43 patients with early response assessment quantitative qPET < 3, indicative of a favorable clinical course, ctDNA was not detectable. In contrast, in five of six patients with qPET > 3, indicative of an unfavorable clinical course, ctDNA remained detectable. ccfDNA analysis of PHL is thus a suitable approach to determine pathogenic mechanisms and monitor therapy response.
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http://dx.doi.org/10.1038/s41375-019-0541-6DOI Listing
January 2020

Mechanisms of Progression of Myeloid Preleukemia to Transformed Myeloid Leukemia in Children with Down Syndrome.

Cancer Cell 2019 08 11;36(2):123-138.e10. Epub 2019 Jul 11.

MRC MHU, BRC Hematology Theme, Oxford Biomedical Research Centre, Oxford Centre for Haematology, WIMM, Radcliffe Department of Medicine, University of Oxford, Oxford OX3 9DU, UK; Department of Paediatrics, University of Oxford, Oxford OX3 9DS, UK.

Myeloid leukemia in Down syndrome (ML-DS) clonally evolves from transient abnormal myelopoiesis (TAM), a preleukemic condition in DS newborns. To define mechanisms of leukemic transformation, we combined exome and targeted resequencing of 111 TAM and 141 ML-DS samples with functional analyses. TAM requires trisomy 21 and truncating mutations in GATA1; additional TAM variants are usually not pathogenic. By contrast, in ML-DS, clonal and subclonal variants are functionally required. We identified a recurrent and oncogenic hotspot gain-of-function mutation in myeloid cytokine receptor CSF2RB. By a multiplex CRISPR/Cas9 screen in an in vivo murine TAM model, we tested loss-of-function of 22 recurrently mutated ML-DS genes. Loss of 18 different genes produced leukemias that phenotypically, genetically, and transcriptionally mirrored ML-DS.
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http://dx.doi.org/10.1016/j.ccell.2019.06.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6863161PMC
August 2019

Phase I dose-escalation study of volasertib in pediatric patients with acute leukemia or advanced solid tumors.

Pediatr Blood Cancer 2019 10 5;66(10):e27900. Epub 2019 Jul 5.

Department of Paediatrics, University Hospital Essen, Essen, Germany.

Background: Volasertib induces mitotic arrest and apoptosis by targeting Polo-like kinases. In this phase I dose-escalation study, the maximum tolerated dose (MTD), pharmacokinetics (PK), and preliminary efficacy of volasertib were determined in pediatric patients.

Methods: Patients aged 2 to <18 years with relapsed/refractory acute leukemia/advanced solid tumors (ST) without available effective treatments were enrolled-cohort C1 (aged 2 to <12 years); cohort C2 (aged 12 to <18 years). The patients received volasertib intravenously (starting dose: 200 mg/m body surface area on day 1, every 14 days). The primary endpoint was the pediatric MTD for further development.

Results: Twenty-two patients received treatment (C1: leukemia, n = 4; ST, n = 8; C2: leukemia, n = 3; ST, n = 7). No dose-limiting toxicities (DLTs) occurred up to 300 mg/m volasertib in C1; two patients in C2, at 250 mg/m volasertib, had DLTs in cycle 1, one of which led to death; therefore, the MTD of volasertib in C2 was 200 mg/m . The most common grade 3/4 adverse events (all patients) were febrile neutropenia, thrombocytopenia, and neutropenia (41% each). Stable disease (SD) was the best objective response (leukemia, n = 5; ST, n = 2); the duration of SD was short in all patients, except in one with an ST. PK profiles were generally comparable across dose groups and were consistent with those in adults.

Conclusion: The pediatric MTD/dose for further development was identified. There were no unexpected safety or PK findings; limited antitumor/antileukemic activity was demonstrated.
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http://dx.doi.org/10.1002/pbc.27900DOI Listing
October 2019

Cardio-toxicity in childhood cancer survivors "Cure is not enough".

J Thorac Dis 2018 Dec;10(Suppl 35):S4344-S4350

Clinic of Pediatrics III, University Hospital Essen, D-45122 Essen, Germany.

The number of pediatric cancer survivors is growing, and they are getting older. Therapy-induced cardiotoxicity therefore is debated as an ongoing problem. Recognition of the side effects in the use of anthracyclines and radiation as well as the patients' clinical condition and comorbidities leads back as far as the beginning of systematic cancer treatment in children in the 1980s. Since, numerous case reports, meta-analyses and retrospective surveys were published worldwide. However, randomized clinical trials with standardized protocols yet fail to be designed. This article gives an overview of the recent reports and emphasizes on the heterogeneity of the different approaches. A standardized work-up which may identify the patient at risk-including the patient's history and condition, individual genetic dispositions, dosage and method of drug application, consideration of co-medication, radiation therapy and dose, standardized imaging methods-is the main proposition of our report. The fusion of already established sources, e.g., data of different registries or study centers, might help to create preventive strategies for and a better understanding of patients with therapy induced cardiomyopathy.
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http://dx.doi.org/10.21037/jtd.2018.11.28DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6328387PMC
December 2018

Detection of AML-specific mutations in pediatric patient plasma using extracellular vesicle-derived RNA.

Ann Hematol 2019 03 23;98(3):595-603. Epub 2019 Jan 23.

Department of Pediatric Hematology and Oncology, University Hospital Essen, Hufelandstraße 55, D-45147, Essen, Germany.

Despite high remission rates, almost 25% of patients with AML will suffer relapse 3-5 years after diagnosis. Therefore, in addition to existing diagnostic and MRD detection tools, there is still a need for the development of novel approaches that can provide information on the state of the disease. Extracellular vesicles (EVs), containing genetic material reflecting the status of the parental cell, have gained interest in recent years as potential diagnostic biomarkers in cancer. Therefore, isolation and characterization of blood and bone marrow plasma-derived EVs from pediatric AML patients could be an additional approach in AML diagnostics and disease monitoring. In this study, we attempt to establish a plasma EV-RNA-based method to detect leukemia-specific FLT3-ITD and NPM1 mutations using established leukemia cell lines and primary pediatric AML plasma samples. We were successfully able to detect FLT3-ITD and NPM1 mutations in the EV-RNA using GeneScan-based fragment-length analysis and real-time PCR assays, respectively, in samples before therapy. This was corresponding to the gDNA mutational analysis from leukemic blasts, and supports the potential of using EV-RNA as a diagnostic biomarker in pediatric AML.
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http://dx.doi.org/10.1007/s00277-019-03608-yDOI Listing
March 2019
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