Publications by authors named "Brigitte Schlegelberger"

267 Publications

Classification of fluorescent R-Band metaphase chromosomes using a convolutional neural network is precise and fast in generating karyograms of hematologic neoplastic cells.

Cancer Genet 2021 Nov 20;260-261:23-29. Epub 2021 Nov 20.

Department of Human Genetics, Hannover Medical School, Hannover 30625, Germany.

Karyotype analysis has a great impact on the diagnosis, treatment and prognosis in hematologic neoplasms. The identification and characterization of chromosomes is a challenging process and needs experienced personal. Artificial intelligence provides novel support tools. However, their safe and reliable application in diagnostics needs to be evaluated. Here, we present a novel laboratory approach to identify chromosomes in cancer cells using a convolutional neural network (CNN). The CNN identified the correct chromosome class for 98.8% of chromosomes, which led to a time saving of 42% for the karyotyping workflow. These results demonstrate that the CNN has potential application value in chromosome classification of hematologic neoplasms. This study contributes to the development of an automatic karyotyping platform.
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http://dx.doi.org/10.1016/j.cancergen.2021.11.005DOI Listing
November 2021

Transcriptional and Mutational Profiling of B-Other Acute Lymphoblastic Leukemia for Improved Diagnostics.

Cancers (Basel) 2021 Nov 12;13(22). Epub 2021 Nov 12.

Institute of Human Genetics, Hannover Medical School (MHH), 30625 Hannover, Germany.

B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most common cancer in children, and significant progress has been made in diagnostics and the treatment of this disease based on the subtypes of BCP-ALL. However, in a large proportion of cases (B-other), recurrent BCP-ALL-associated genomic alterations remain unidentifiable by current diagnostic procedures. In this study, we performed RNA sequencing and analyzed gene fusions, expression profiles, and mutations in diagnostic samples of 185 children with BCP-ALL. Gene expression clustering showed that a subset of B-other samples partially clusters with some of the known subgroups, particularly -positive. Mutation analysis coupled with gene expression profiling revealed the presence of distinctive BCP-ALL subgroups, characterized by the presence of mutations in known ALL driver genes, e.g., and . Moreover, we identified novel fusion partners of lymphoid lineage transcriptional factors , and . In addition, we report on low blast count detection thresholds and show that the use of EDTA tubes for sample collection does not have adverse effects on sequencing and downstream analysis. Taken together, our findings demonstrate the applicability of whole-transcriptome sequencing for personalized diagnostics in pediatric ALL, including tentative classification of the B-other cases that are difficult to diagnose using conventional methods.
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http://dx.doi.org/10.3390/cancers13225653DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8616234PMC
November 2021

The Clinical Utility of Optical Genome Mapping for the Assessment of Genomic Aberrations in Acute Lymphoblastic Leukemia.

Cancers (Basel) 2021 Aug 30;13(17). Epub 2021 Aug 30.

Department of Human Genetics, Hannover Medical School, 30625 Hannover, Germany.

Acute lymphoblastic leukemia (ALL) is the most prevalent type of cancer occurring in children. ALL is characterized by structural and numeric genomic aberrations that strongly correlate with prognosis and clinical outcome. Usually, a combination of cyto- and molecular genetic methods (karyotyping, array-CGH, FISH, RT-PCR, RNA-Seq) is needed to identify all aberrations relevant for risk stratification. We investigated the feasibility of optical genome mapping (OGM), a DNA-based method, to detect these aberrations in an all-in-one approach. As proof of principle, twelve pediatric ALL samples were analyzed by OGM, and results were validated by comparing OGM data to results obtained from routine diagnostics. All genomic aberrations including translocations (e.g., dic(9;12)), aneuploidies (e.g., high hyperdiploidy) and copy number variations (e.g., , ) known from other techniques were also detected by OGM. Moreover, OGM was superior to well-established techniques for resolution of the more complex structure of a translocation t(12;21) and had a higher sensitivity for detection of copy number alterations. Importantly, a new and unknown gene fusion of and due to a translocation t(9;11) was detected. We demonstrate the feasibility of OGM to detect well-established as well as new putative prognostic markers in an all-in-one approach in ALL. We hope that these limited results will be confirmed with testing of more samples in the future.
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http://dx.doi.org/10.3390/cancers13174388DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8431583PMC
August 2021

Cryptic TCF3 fusions in childhood leukemia: Detection by RNA sequencing.

Genes Chromosomes Cancer 2022 Jan 13;61(1):22-26. Epub 2021 Sep 13.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

Acute lymphoblastic leukemia (ALL) is the most frequent malignancy in childhood and adolescence. In more than 60% of cases of this heterogeneous disease, a genetic marker is identified via cytogenetic or molecular analyses. TCF3 gene fusions occur in 5%-11% of ALL patients. In < 1%, the TCF3 alteration in ALL leads to a TCF3-HLF fusion gene. Even though this is a very rare event, the detection of a TCF3-HLF fusion gene is associated with a very poor prognosis with incurable relapses in almost all patients. The frequent TCF3-PBX1 fusion gene, which is detectable in 5%-10% of childhood B-cell precursor ALLs and ~3.8% of adult B-cell precursor ALLs, is associated with a rather good prognosis, that is, an observed event-free 5-year survival of approximately 85%. Thus, the distinction of the different partner genes fused to TCF3 is essential for risk assessment. To verify RNA sequencing as a tool for detection of known and unknown fusion genes, we screened 200 cases of pediatric B-cell precursor ALL with "targeted" RNA sequencing in a pilot project in comparison to classical cytogenetic analyses (chromosome R-banding analysis), fluorescence in situ hybridization, and PCR. We observed a TCF3 fusion gene in 6.5% (13/200) of the patients. Ten (5%) patients displayed a TCF3-PBX1 fusion gene, two (1%) patients a TCF3-FLI1 fusion gene, and one (0.5%) patient a TCF3-HLF fusion gene. For the TCF3 fusions, we obtained discrepant results with the different methods, which are described in the article. Taken together, translocations leading to TCF3 fusion genes might appear cryptic and may remain undetected by a single method.
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http://dx.doi.org/10.1002/gcc.22998DOI Listing
January 2022

MicroRNA-192-5p inhibits migration of triple negative breast cancer cells and directly regulates Rho GTPase activating protein 19.

Genes Chromosomes Cancer 2021 Nov 6;60(11):733-742. Epub 2021 Aug 6.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

Among the different breast cancer subtypes, triple-negative breast cancer (TNBC) is associated with a poor prognosis, low survival rates, and high expression of histone deacetylases. Treatment with histone deacetylase inhibitor trichostatin A (TSA) leads to an increased expression of potential tumor-suppressive miRNAs. Characterization of these miRNAs can help to find new molecular targets for treatment of TNBC. We identified differentially expressed miRNAs by microarray analyses after treatment with TSA in the TNBC cell lines HCC38, HCC1395, and HCC1935. The gene locus of hsa-miRNA-192-5p (miR-192) and hsa-miR-194-2 (miR-194-2) with its host gene, long noncoding RNA miR-194-2HG, has been linked to inhibition of migration in different tumor types. Therefore, we examined tumor-relevant functional effects using WST-1-based proliferation, capsase-3/7-based apoptosis, and trans-well migration assays after transfection with miRNA mimics or specific siRNAs. We demonstrated the tumor-suppressive capacity of miR-192 in TNBC cells, which was exerted through inhibition of proliferation, induction of apoptosis, and reduction of migration. Gene expression and bioinformatics analyses of TNBC cell lines transfected with miR-192 mimics, identified a number of genes involved in migration including the Rho GTPase Activating Protein ARHGAP19. Through RNA immunoprecipitation we demonstrated the direct binding of miR-192 and ARHGAP19. Downregulation of ARHGAP19 expression by either miR-192 or siRNA inhibited migration of TNBC cells significantly. Our findings demonstrate that overexpression of epigenetically deregulated miR-192 decreases proliferation, promotes apoptosis, and inhibits migration of TNBC cell lines.
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http://dx.doi.org/10.1002/gcc.22982DOI Listing
November 2021

Review of guidelines for the identification and clinical care of patients with genetic predisposition for hematological malignancies.

Fam Cancer 2021 Oct 31;20(4):295-303. Epub 2021 May 31.

Division of Pediatric Hematology and Oncology, Medical Center, Department of Pediatrics and Adolescent Medicine, Faculty of Medicine, University of Freiburg, Freiburg, Germany.

Since WHO has recognized myeloid neoplasms with germline predisposition as a new entity in 2016, it has become increasingly clear that diagnosing familial leukemia has critical implications for both the patient and his/her family, and that interdisciplinary teams of hematologists and clinical geneticists should provide care for this specific patient group. Here, we summarize consensus criteria for the identification and screening of patients with genetic predisposition for hematologic malignancies, as provided by different working groups, e.g. by the Nordic MDS group and the AACR. In addition to typical clinical features, results from targeted deep sequencing may point to a genetic predisposition. We review strategies to distinguish somatic and germline variants and discuss recommendations for genetic analyses aiming to identify the underlying genetic variant that should follow established quality criteria to detect both SNVs and CNVs and to determine the pathogenicity of genetic variants. To enhance the knowledge about hematologic neoplasms with germline predisposition we recommend archiving clinical and genetic data and archiving them in international registries.
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http://dx.doi.org/10.1007/s10689-021-00263-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8484082PMC
October 2021

Students' attitudes towards somatic genome editing versus genome editing of the germline using an example of familial leukemia.

J Community Genet 2021 Jul 8;12(3):397-406. Epub 2021 May 8.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

Although the discussion on possibilities and pitfalls of genome editing is ever present, limited qualitative data on the attitudes of students, who will come into contact with this technology within a social and professional context, is available. The attitude of 97 medical students and 103 students of other subjects from Hannover and Oldenburg, Germany, was analyzed in winter 2017/18. For this purpose, two dilemmas on somatic and germline genome editing concerning familial leukemia were developed. After reading the dilemmas, the students filled out a paper-and-pencil test with five open questions. The qualitative evaluation of the answers was carried by a deductive-inductive procedure of content analysis. There was a high approval for the use of somatic genome editing. When it came to germline genome editing, concerns were raised regarding enhancement, interventions in nature, and loss of uniqueness. The students recognized that somatic genome editing and germline genome editing prove different ethical challenges and need to be judged separately. Many students expressed not feeling fully informed. The results of this project show the importance of educating the public about the possibilities, limitations, and risks of somatic and germline genome editing. We recommend that this should already be addressed in schools in order to optimally prepare students and adults for participation in public discourse. Especially for patients affected by genetic diseases, it is of great importance that the treating physicians and geneticists are sufficiently informed about the method of genome editing to ensure good counseling.
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http://dx.doi.org/10.1007/s12687-021-00528-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8241980PMC
July 2021

Psychological Distress and Coping Ability of Women at High Risk of Hereditary Breast and Ovarian Cancer before Undergoing Genetic Counseling-An Exploratory Study from Germany.

Int J Environ Res Public Health 2021 04 19;18(8). Epub 2021 Apr 19.

Department of Human Genetics, Hanover Medical School, 30625 Hannover, Germany.

Carriers of pathogenic variants causing hereditary breast and ovarian cancer (HBOC) are confronted with a high risk to develop malignancies early in life. The present study aimed to determine the type of psychological distress and coping ability in women with a suspicion of HBOC. In particular, we were interested if the self-assessed genetic risk had an influence on health concerns and coping ability. Using a questionnaire established by the German HBOC Consortium, we investigated 255 women with breast cancer and 161 healthy women before they were seen for genetic counseling. The group of healthy women was divided into groups of high and low self-assessed risk. In our study, healthy women with a high self-assessed risk stated the highest stress level and worries about their health and future. A quarter of the women requested psychological support. Overall, only few women (4-11%) stated that they did not feel able to cope with the genetic test result. More women (11-23%, highest values in the low-risk group) worried about the coping ability of relatives. The results of our exploratory study demonstrate that the women, who presented at the Department of Human Genetics, Hanover Medical School, Germany were aware of their genetic risk and had severe concerns about their future health, but still felt able to cope with the genetic test result.
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http://dx.doi.org/10.3390/ijerph18084338DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8073852PMC
April 2021

Plasma Metabolome Signature Indicative of Germline Status Independent of Cancer Incidence.

Front Oncol 2021 7;11:627217. Epub 2021 Apr 7.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

Individuals carrying a pathogenic germline variant in the breast cancer predisposition gene (g+) are prone to developing breast cancer. Apart from its well-known role in DNA repair, BRCA1 has been shown to powerfully impact cellular metabolism. While, in general, metabolic reprogramming was named a hallmark of cancer, disrupted metabolism has also been suggested to drive cancer cell evolution and malignant transformation by critically altering microenvironmental tissue integrity. Systemic metabolic effects induced by germline variants in cancer predisposition genes have been demonstrated before. Whether or not systemic metabolic alterations exist in g+ individuals independent of cancer incidence has not been investigated yet. We therefore profiled the plasma metabolome of 72 g+ women and 72 age-matched female controls, none of whom (carriers and non-carriers) had a prior cancer diagnosis and all of whom were cancer-free during the follow-up period. We detected one single metabolite, pyruvate, and two metabolite ratios involving pyruvate, lactate, and a metabolite of yet unknown structure, significantly altered between the two cohorts. A machine learning signature of metabolite ratios was able to correctly distinguish between g+ and controls in ~82%. The results of this study point to innate systemic metabolic differences in g+ women independent of cancer incidence and raise the question as to whether or not constitutional alterations in energy metabolism may be involved in the etiology of -associated breast cancer.
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http://dx.doi.org/10.3389/fonc.2021.627217DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8058469PMC
April 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 06 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

Unbalanced translocation der(5;17) resulting in a TP53 loss as recurrent aberration in myelodysplastic syndrome and acute myeloid leukemia with complex karyotype.

Genes Chromosomes Cancer 2021 Jun 19;60(6):452-457. Epub 2021 Feb 19.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

A complex karyotype, detected in myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML), is associated with a reduced median survival. The most frequent chromosomal aberrations in complex karyotypes are deletions of 5q and 17p harboring the tumor suppressor gene TP53. The unbalanced translocation der(5;17) involving chromosome 5q and 17p is a recurrent aberration in MDS/AML, resulting in TP53 loss. We analyzed the karyotypes of 178 patients with an unbalanced translocation der(5;17) using fluorescence R-/G-banding analysis. Whenever possible, fluorescence in situ hybridization (FISH) (n = 138/141), multicolor FISH (n = 8), telomere length measurement (n = 9), targeted DNA sequencing (n = 13), array-CGH (n = 7) and targeted RNA sequencing (n = 2) were conducted. The der(5;17) aberration was accompanied with loss of genetic material in 7q (53%), -7 (27%), gain of 21q (29%), +8 (17%) and - 18 (16%) and all analyzed patients (n = 13) showed a (likely) pathogenic variant inTP53. The der(5;17) cohort showed significantly shortened telomeres in comparison to the healthy age-matched controls (P < .05), but there was no significant telomere shortening in comparison to MDS/AML patients with a complex karyotype without der(5;17). No fusion genes resulted from the unbalanced translocation. This study demonstrates that the unbalanced translocation der(5;17) is associated with a biallelic inactivation of TP53 due to a deletion of TP53 in one allele and a pathogenic variant of the second TP53 allele. Since the breakpoints are located within (near-) heterochromatic regions, alterations of DNA methylation or histone modifications may be involved in the generation of der(5;17).
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http://dx.doi.org/10.1002/gcc.22938DOI Listing
June 2021

Knockout of the HMG domain of the porcine SRY gene causes sex reversal in gene-edited pigs.

Proc Natl Acad Sci U S A 2021 01;118(2)

Institute of Farm Animal Genetics, Friedrich-Loeffler-Institut, Mariensee, 31535 Neustadt am Rübenberge, Germany;

The sex-determining region on the Y chromosome (SRY) is thought to be the central genetic element of male sex development in mammals. Pathogenic modifications within the SRY gene are associated with a male-to-female sex reversal syndrome in humans and other mammalian species, including rabbits and mice. However, the underlying mechanisms are largely unknown. To understand the biological function of the SRY gene, a site-directed mutational analysis is required to investigate associated phenotypic changes at the molecular, cellular, and morphological level. Here, we successfully generated a knockout of the porcine SRY gene by microinjection of two CRISPR-Cas ribonucleoproteins, targeting the centrally located "high mobility group" (HMG), followed by a frameshift mutation of the downstream SRY sequence. This resulted in the development of genetically male (XY) pigs with complete external and internal female genitalia, which, however, were significantly smaller than in 9-mo-old age-matched control females. Quantitative digital PCR analysis revealed a duplication of the SRY locus in Landrace pigs similar to the known palindromic duplication in Duroc breeds. Our study demonstrates the central role of the HMG domain in the SRY gene in male porcine sex determination. This proof-of-principle study could assist in solving the problem of sex preference in agriculture to improve animal welfare. Moreover, it establishes a large animal model that is more comparable to humans with regard to genetics, physiology, and anatomy, which is pivotal for longitudinal studies to unravel mammalian sex determination and relevant for the development of new interventions for human sex development disorders.
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http://dx.doi.org/10.1073/pnas.2008743118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7812820PMC
January 2021

Establishment and Characterization of a Sclerosing Spindle Cell Rhabdomyosarcoma Cell Line with a Complex Genomic Profile.

Cells 2020 12 11;9(12). Epub 2020 Dec 11.

Department of Orthopaedic Surgery, Eberhard Karls University Tuebingen, 72072 Tuebingen, Germany.

Sclerosing spindle cell rhabdomyosarcoma (SSRMS) is a rare rhabdomyosarcomas (RMS) subtype. Especially cases bearing a myogenic differentiation 1 () mutation are characterized by a high recurrence and metastasis rate, often leading to a fatal outcome. SSRMS cell lines are valuable in vitro models for studying disease mechanisms and for the preclinical evaluation of new therapeutic approaches. In this study, a cell line established from a primary SSRMS tumor of a 24-year-old female after multimodal chemotherapeutic pretreatment has been characterized in detail, including immunohistochemistry, growth characteristics, cytogenetic analysis, mutation analysis, evaluation of stem cell marker expression, differentiation potential, and tumorigenicity in mice. The cell line which was designated SRH exhibited a complex genomic profile, including several translocations and deletions. Array-comparative genomic hybridization (CGH) revealed an overall predominating loss of gene loci. The mesenchymal tumor origin was underlined by the expression of mesenchymal markers and potential to undergo adipogenic and osteogenic differentiation. Despite myogenic marker expression, terminal myogenic differentiation was inhibited, which might be elicited by the hotspot mutation. In vivo tumorigenicity could be confirmed after subcutaneous injection into NOD/SCID/γ mice. Summarized, the SRH cell line is the first adult SSRMS cell line available for preclinical research on this rare RMS subtype.
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http://dx.doi.org/10.3390/cells9122668DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7763666PMC
December 2020

Dihydropyrimidine Dehydrogenase Testing prior to Treatment with 5-Fluorouracil, Capecitabine, and Tegafur: A Consensus Paper.

Oncol Res Treat 2020 23;43(11):628-636. Epub 2020 Oct 23.

Klinik für Innere Medizin I, Universitätsklinikum Ulm, Ulm, Germany.

Background: 5-Fluorouracil (FU) is one of the most commonly used cytostatic drugs in the systemic treatment of cancer. Treatment with FU may cause severe or life-threatening side effects and the treatment-related mortality rate is 0.2-1.0%.

Summary: Among other risk factors associated with increased toxicity, a genetic deficiency in dihydropyrimidine dehydrogenase (DPD), an enzyme responsible for the metabolism of FU, is well known. This is due to variants in the DPD gene (DPYD). Up to 9% of European patients carry a DPD gene variant that decreases enzyme activity, and DPD is completely lacking in approximately 0.5% of patients. Here we describe the clinical and genetic background and summarize recommendations for the genetic testing and tailoring of treatment with 5-FU derivatives. The statement was developed as a consensus statement organized by the German Society for Hematology and Medical Oncology in cooperation with 13 medical associations from Austria, Germany, and Switzerland. Key Messages: (i) Patients should be tested for the 4 most common genetic DPYD variants before treatment with drugs containing FU. (ii) Testing forms the basis for a differentiated, risk-adapted algorithm with recommendations for treatment with FU-containing drugs. (iii) Testing may optionally be supplemented by therapeutic drug monitoring.
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http://dx.doi.org/10.1159/000510258DOI Listing
March 2021

IDH1/2 mutations in acute myeloid leukemia patients and risk of coronary artery disease and cardiac dysfunction-a retrospective propensity score analysis.

Leukemia 2021 05 18;35(5):1301-1316. Epub 2020 Sep 18.

Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Carl-Neuberg Strasse 1, 30625, Hannover, Germany.

Clonal hematopoiesis of indeterminate potential (CHIP) is linked to leukemia gene mutations and associates with an increased risk for coronary artery disease and poor prognosis in ischemic cardiomyopathy. Two recurrently mutated genes in CHIP and adult acute myeloid leukemia (AML) encode for isocitrate dehydrogenases 1 and 2 (IDH1 and IDH2). Global expression of mutant IDH2 in transgenic mice-induced dilated cardiomyopathy and muscular dystrophy. In this retrospective observational study, we investigated whether mutant IDH1/2 predisposes to cardiovascular disease in AML patients. Among 363 AML patients, IDH1 and IDH2 mutations were detected in 26 (7.2%) and 39 patients (10.7%), respectively. Mutant IDH1 patients exhibited a significantly higher prevalence of coronary artery disease (26.1% vs. 6.4%, p = 0.002). Applying inverse probability-weighting analysis, patients with IDH1/2 mutations had a higher risk for a declining cardiac function during AML treatment compared to IDH1/2 wild type patients [left ventricular ejection fraction pretreatment compared to 10 months after diagnosis: 59.2% to 41.9% (p < 0.001) vs 58.5% to 55.4% (p = 0.27), respectively]. Mechanistically, RNA sequencing and immunostaining in hiPS-derived cardiomyocytes indicated that the oncometabolite R-2HG exacerbated doxorubicin mediated cardiotoxicity. Evaluation of IDH1/2 mutation status may therefore help identifying AML patients at risk for cardiovascular complications during cytotoxic treatment.
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http://dx.doi.org/10.1038/s41375-020-01043-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8102189PMC
May 2021

De novo missense variants in the RAP1B gene identified in two patients with syndromic thrombocytopenia.

Clin Genet 2020 10 21;98(4):374-378. Epub 2020 Jul 21.

Department of Human Genetics, Hannover Medical School, Hanover, Germany.

We present two independent cases of syndromic thrombocytopenia with multiple malformations, microcephaly, learning difficulties, dysmorphism and other features. Exome sequencing identified two novel de novo heterozygous variants in these patients, c.35G>T p.(Gly12Val) and c.178G>C p.(Gly60Arg), in the RAP1B gene (NM_001010942.2). These variants have not been described previously as germline variants, however functional studies in literature strongly suggest a clinical implication of these two activating hot spot positions. We hypothesize that pathogenic missense variants in the RAP1B gene cause congenital syndromic thrombocytopenia with a spectrum of associated malformations and dysmorphism, possibly through a gain of function mechanism.
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http://dx.doi.org/10.1111/cge.13807DOI Listing
October 2020

Frequency and prognostic impact of PAX5 p.P80R in pediatric acute lymphoblastic leukemia patients treated on an AIEOP-BFM acute lymphoblastic leukemia protocol.

Genes Chromosomes Cancer 2020 11 7;59(11):667-671. Epub 2020 Jul 7.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

PAX5 is a member of the paired box (PAX) family of transcription factors involved in B-cell development. PAX5 has recently been described as a distinct genetic B-cell precursor (BCP) acute lymphoblastic leukemia (ALL) subtype with a favorable prognosis in adults. In contrast, an unfavorable outcome has been observed in children. Our aim was to determine the frequency of PAX5 in childhood BCP-ALL treated according to the Associazione Italiana Ematologia ed Oncologia Pediatrica-Berlin-Frankfurt-Muenster (AIEOP-BFM) ALL 2000 protocol and to evaluate its clinical significance within this study cohort. The analyses included 1237 patients with ALL treated in the AIEOP-BFM ALL 2000 trial with complete information for copy number variations (CNVs) of IKZF1, PAX5, ETV6, RB1, BTG1, EBF1, CDKN2A, CDKN2B, and ERG. A customized TaqMan genotyping assay was used to screen for PAX5 . Sanger sequencing was used to confirm PAX5 -positive results as well as to screen for second variants in PAX5. Agilent CGH + SNP arrays (e-Array design 85 320; Agilent Technologies) were performed in PAX5 -positive patients to verify additional CNVs. Almost 2% (20/1028) of our BCP-ALL cohort were PAX5 -positive. White blood cell counts higher than 50 000/μl as well as male sex were significantly (P < .05) associated with PAX5 . Most of the PAX5 -positive cases were 10 years of age or older. PAX5 -positive samples were enriched for deletions affecting PAX5, IKZF1, CDKN2A, and CDKN2B. Compared to PAX5 -wildtype BCP-ALL, PAX5 -positive patients showed a significantly reduced 5-year overall survival (P = .042). Further studies should evaluate the interaction of PAX5 with other genetic aberrations to further stratify intermediate risk pediatric BCP-ALL.
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http://dx.doi.org/10.1002/gcc.22882DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7540392PMC
November 2020

High-risk additional chromosomal abnormalities at low blast counts herald death by CML.

Leukemia 2020 08 7;34(8):2074-2086. Epub 2020 May 7.

Department of Hematology-Oncology, Policlinico S.Orsola-Malpighi, University of Bologna, Bologna, Italy.

Blast crisis is one of the remaining challenges in chronic myeloid leukemia (CML). Whether additional chromosomal abnormalities (ACAs) enable an earlier recognition of imminent blastic proliferation and a timelier change of treatment is unknown. One thousand five hundred and ten imatinib-treated patients with Philadelphia-chromosome-positive (Ph+) CML randomized in CML-study IV were analyzed for ACA/Ph+ and blast increase. By impact on survival, ACAs were grouped into high risk (+8, +Ph, i(17q), +17, +19, +21, 3q26.2, 11q23, -7/7q abnormalities; complex) and low risk (all other). The presence of high- and low-risk ACAs was linked to six cohorts with different blast levels (1%, 5%, 10%, 15%, 20%, and 30%) in a Cox model. One hundred and twenty-three patients displayed ACA/Ph+ (8.1%), 91 were high risk. At low blast levels (1-15%), high-risk ACA showed an increased hazard to die compared to no ACA (ratios: 3.65 in blood; 6.12 in marrow) in contrast to low-risk ACA. No effect was observed at blast levels of 20-30%. Sixty-three patients with high-risk ACA (69%) died (n = 37) or were alive after progression or progression-related transplantation (n = 26). High-risk ACA at low blast counts identify end-phase CML earlier than current diagnostic systems. Mortality was lower with earlier treatment. Cytogenetic monitoring is indicated when signs of progression surface or response to therapy is unsatisfactory.
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http://dx.doi.org/10.1038/s41375-020-0826-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7387244PMC
August 2020

The complex genetic landscape of familial MDS and AML reveals pathogenic germline variants.

Nat Commun 2020 02 25;11(1):1044. Epub 2020 Feb 25.

UMC Utrecht Cancer Center, Universitair Medisch Centrum Utrecht, Huispostnummer, Utrecht, Netherlands.

The inclusion of familial myeloid malignancies as a separate disease entity in the revised WHO classification has renewed efforts to improve the recognition and management of this group of at risk individuals. Here we report a cohort of 86 acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) families with 49 harboring germline variants in 16 previously defined loci (57%). Whole exome sequencing in a further 37 uncharacterized families (43%) allowed us to rationalize 65 new candidate loci, including genes mutated in rare hematological syndromes (ADA, GP6, IL17RA, PRF1 and SEC23B), reported in prior MDS/AML or inherited bone marrow failure series (DNAH9, NAPRT1 and SH2B3) or variants at novel loci (DHX34) that appear specific to inherited forms of myeloid malignancies. Altogether, our series of MDS/AML families offer novel insights into the etiology of myeloid malignancies and provide a framework to prioritize variants for inclusion into routine diagnostics and patient management.
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http://dx.doi.org/10.1038/s41467-020-14829-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7042299PMC
February 2020

Implementation of RNA sequencing and array CGH in the diagnostic workflow of the AIEOP-BFM ALL 2017 trial on acute lymphoblastic leukemia.

Ann Hematol 2020 Apr 20;99(4):809-818. Epub 2020 Feb 20.

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

Risk-adapted therapy has significantly contributed to improved survival rates in pediatric acute lymphoblastic leukemia (ALL) and reliable detection of chromosomal aberrations is mandatory for risk group stratification. This study evaluated the applicability of panel-based RNA sequencing and array CGH within the diagnostic workflow of the German study group of the international AIEOP-BFM ALL 2017 trial. In a consecutive cohort of 117 children with B cell precursor (BCP) ALL, array analysis identified twelve cases with an IKZF1 profile of gene deletions and one case of masked hypodiploidy. Genetic markers BCR-ABL1 (n = 1), ETV6-RUNX1 (n = 25), and rearrangements involving KMT2A (n = 3) or TCF3 (n = 3) were assessed by established conventional techniques such as karyotyping, FISH, and RT-PCR. Comparison of these results with RNA sequencing analysis revealed overall consistency in n=115/117 cases, albeit with one undetected AFF1-KMT2A fusion in RNA sequencing and one undetected ETV6-RUNX1 fusion in conventional analyses. The combined application of RNA sequencing, FISH, and CGH+SNP array reliably detected all genetic markers necessary for risk stratification and will be used as the diagnostic standard workflow for BCP-ALL patients enrolled in the AIEOP-BFM ALL 2017 study. Prospectively, consistent collection of genome-wide CGH+SNP array as well as RNA sequencing data will be a valuable source to elucidate new prognostic lesions beyond established markers of pediatric ALL. In this respect, RNA sequencing identified various gene fusions in up to half of the IKZF1 (n = 6/12) and B-other (n = 19/36) cases but not in cases with hyperdiploid karyotypes (n = 35). Among these fusions, this study reports several previously undescribed in frame PAX5 fusions, including PAX5-MYO1G and PAX5-NCOA6.
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http://dx.doi.org/10.1007/s00277-020-03953-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7069912PMC
April 2020

From a variant of unknown significance to pathogenic: Reclassification of a large novel duplication in BRCA2 by high-throughput sequencing.

Mol Genet Genomic Med 2020 09 13;8(9):e1045. Epub 2019 Nov 13.

Department of Human Genetics, Hannover Medical School, Hannover, Germany.

Background: Germline mutations in BRCA1/2 significantly contribute to hereditary breast and/or ovarian cancer. Here, we report a novel BRCA2 duplication of exons 22-24 in a female patient with bilateral breast cancer at age 35 and 44. The duplicated region was initially detected by gene panel sequencing and multiplex ligation-dependent probe amplification. However, the location and orientation of the duplicated region was unknown. Therefore, it was initially classified as a variant of unknown significance.

Methods: The spatial directional characterization of the BRCA2 duplication was achieved by targeted enrichment of the whole-genomic BRCA2 locus including exons and introns, and subsequent high-throughput sequencing. Subsequently, bioinformatics tools and a breakpoint-spanning PCR were used for identification of location and orientation of the duplication.

Results: The duplicated region was arranged in tandem and direct orientation (Chr13(GRCh37):g.32951579_32960394dup; NM_000059.3 c.8754 + 651_9256+6112dup p.(Ala3088Phefs*3)). It is predicted to result in a frameshift and a premature stop codon likely triggering nonsense-mediated mRNA decay. Consequently, it is regarded as pathogenic.

Conclusion: This case study demonstrates that a comprehensive characterization of a structural variant by breakpoint assessment is crucial for its correct classification. Therefore, sequencing strategies including non-coding regions might be necessary to identify cancer predispositions in affected families.
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http://dx.doi.org/10.1002/mgg3.1045DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7506983PMC
September 2020
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