Histone deacetylation, a common hallmark in malignant tumors, strongly alters the transcription of genes involved in the control of proliferation, cell survival, differentiation and genetic stability. We have previously shown that HDAC1, HDAC2, and HDAC3 (HDAC1-3) genes encoding histone deacetylases 1-3 are upregulated in primary human hepatocellular carcinoma (HCC). The aim of this study was to characterize the functional effects of HDAC1-3 downregulation and to identify functionally important target genes of histone deacetylation in HCC.

Fusion genes are potent driver mutations in cancer. In this study, we delineate the fusion gene landscape in a consecutive series of 195 paediatric B-cell precursor acute lymphoblastic leukaemia (BCP ALL). Using RNA sequencing, we find in-frame fusion genes in 127 (65%) cases, including 27 novel fusions. We describe a subtype characterized by recurrent IGH-DUX4 or ERG-DUX4 fusions, representing 4% of cases, leading to overexpression of DUX4 and frequently co-occurring with intragenic ERG deletions. Furthermore, we identify a subtype characterized by an ETV6-RUNX1-like gene-expression profile and coexisting ETV6 and IKZF1 alterations. Thus, this study provides a detailed overview of fusion genes in paediatric BCP ALL and adds new pathogenetic insights, which may improve risk stratification and provide therapeutic options for this disease.

Unbalanced (major route) additional cytogenetic aberrations (ACA) at diagnosis of chronic myeloid leukemia (CML) indicate an increased risk of progression and shorter survival. Moreover, newly arising ACA under imatinib treatment and clonal evolution are considered features of acceleration and define failure of therapy according to the European LeukemiaNet (ELN) recommendations. On the basis of 1151 Philadelphia chromosome positive chronic phase patients of the randomized CML-study IV, we examined the incidence of newly arising ACA under imatinib treatment with regard to the p210BCR-ABL breakpoint variants b2a2 and b3a2. We found a preferential acquisition of unbalanced ACA in patients with b3a2 vs. b2a2 fusion type (ratio: 6.3 vs. 1.6, p = 0.0246) concurring with a faster progress to blast crisis for b3a2 patients (p = 0.0124). ESPL1/Separase, a cysteine endopeptidase, is a key player in chromosomal segregation during mitosis. Separase overexpression and/or hyperactivity has been reported from a wide range of cancers and cause defective mitotic spindles, chromosome missegregation and aneuploidy. We investigated the influence of p210BCR-ABL breakpoint variants and imatinib treatment on expression and proteolytic activity of Separase as measured with a specific fluorogenic assay on CML cell lines (b2a2: KCL-22, BV-173; b3a2: K562, LAMA-84). Despite a drop in Separase protein levels an up to 5.4-fold increase of Separase activity under imatinib treatment was observed exclusively in b3a2 but not in b2a2 cell lines. Mimicking the influence of imatinib on BV-173 and LAMA-84 cells by ESPL1 silencing stimulated Separase proteolytic activity in both b3a2 and b2a2 cell lines. Our data suggest the existence of a fusion type-related feedback mechanism that posttranslationally stimulates Separase proteolytic activity after therapy-induced decreases in Separase protein levels. This could render b3a2 CML cells more prone to aneuploidy and clonal evolution than b2a2 progenitors and may therefore explain the cytogenetic results of CML patients.

Long non-coding RNAs (lncRNAs) have emerged as a novel class of RNA due to its diverse mechanism in cancer development and progression. However, the role and expression pattern of lncRNAs in molecular subtypes of B cell acute lymphoblastic leukemia (BCP-ALL) have not yet been investigated. Here, we assess to what extent lncRNA expression and DNA methylation is driving the progression of relapsed BCP-ALL subtypes and we determine if the expression and DNA methylation profile of lncRNAs correlates with established BCP-ALL subtypes.

The significance of human biorepositories for modern medical research, particularly for comprehensive population-based genetic analyses, is constantly growing. While large and centralized institutions are usually considered best suited to meet the increasing demand for high-quality "biobanks," most medical research institutions still host rather heterogeneous and fragmented biobanking activities, undertaken by clinical departments with oftentimes rather different scientific scope. Undoubtedly, most clinicians and medical researchers would appreciate infrastructural support in terms of the storage and handling of their biosamples, but they are also likely to expect access to their samples avoiding extensive formal requirements. We report on the establishment of the PopGen 2.0 Network (P2N), an overarching alliance of initially seven biobanks from Northern Germany which adopted a joint but lean governance structure and use-and-access policy for their samples and data. In addition, the members of P2N have pursued an intense collaboration on ethical, legal and social issues and maintain a common IT infrastructure. The implementation of P2N has substantially improved the prospects of biobank-based research at the participating institutions. The network may thus serve as a role model for similar initiatives geared at linking pre-existing biorepositories for the benefit of research quality, efficiency, and transparency.

There is no international consensus up to which age women with a diagnosis of triple-negative breast cancer (TNBC) and no family history of breast or ovarian cancer should be offered genetic testing for germline BRCA1 and BRCA2 (gBRCA) mutations. Here, we explored the association of age at TNBC diagnosis with the prevalence of pathogenic gBRCA mutations in this patient group.

Amyotrophic lateral sclerosis (ALS) is a fatal disorder of the motor neuron system with poor prognosis and marginal therapeutic options. Current clinical diagnostic criteria are based on electrophysiological examination and exclusion of other ALS-mimicking conditions. Neuroprotective treatments are, however, most promising in early disease stages. Identification of disease-specific CSF biomarkers and associated biochemical pathways is therefore most relevant to monitor disease progression, response to neuroprotective agents and to enable early inclusion of patients into clinical trials.

Evidence for distinct human cancer stem cells (CSCs) remains contentious and the degree to which different cancer cells contribute to propagating malignancies in patients remains unexplored. In low- to intermediate-risk myelodysplastic syndromes (MDS), we establish the existence of rare multipotent MDS stem cells (MDS-SCs), and their hierarchical relationship to lineage-restricted MDS progenitors. All identified somatically acquired genetic lesions were backtracked to distinct MDS-SCs, establishing their distinct MDS-propagating function in vivo. In isolated del(5q)-MDS, acquisition of del(5q) preceded diverse recurrent driver mutations. Sequential analysis in del(5q)-MDS revealed genetic evolution in MDS-SCs and MDS-progenitors prior to leukemic transformation. These findings provide definitive evidence for rare human MDS-SCs in vivo, with extensive implications for the targeting of the cells required and sufficient for MDS-propagation.

In the European Intergroup EURO-LB02 trial, children and adolescents with lymphoblastic lymphoma underwent the non-Hodgkin lymphoma Berlin-Frankfurt-Münster protocol without prophylactic cranial radiotherapy. The primary aims of this trial were to test whether replacing prednisone with dexamethasone during induction increases event-free survival in the subgroups with T-cell lymphoblastic lymphoma and whether therapy duration could be reduced from 24 to 18 months (factorial design, randomizations). These questions could not be answered due to premature closure of the trial. Here we report on the secondary aims of the trial: whether the results of the NHL-BFM90 study could be reproduced and evaluation of disease features and prognostic factors. Three hundred and nineteen patients (66 with precursor B-cell lymphoblastic lymphoma, 233 with T-cell lymphoblastic lymphoma, 12 with mixed phenotype, 8 not classifiable) were enrolled. In induction, 215 patients received prednisone and 104 patients received dexamethasone. The median follow-up was 6.8 years (range, 3.0-10.3). The 5-year event-free survival was 82±2% [12 toxic deaths, 5 secondary malignancies, 43 non-response/relapse (central nervous system n=9; all received prednisone during induction)]. The event-free survival rate was 80±5% for patients with precursor B-cell lymphoblastic lymphoma, 82±3% for those with T-cell lymphoblastic lymphoma, and 100% for patients with a mixed phenotype. During induction, significantly more grade III/IV toxicities were observed in patients receiving dexamethasone, resulting in significant treatment delays. The number of toxic deaths did not differ significantly. The only variable associated with outcome was performance status at diagnosis. The 90% event-free survival rate for patients with T-cell lymphoblastic lymphoma shown in study NHL-BFM90 was not replicated, mainly due to more toxic deaths and central nervous system relapses. Dexamethasone in induction may prevent central nervous system relapse more effectively than prednisone but produces a higher burden of toxicity. (#NCT00275106).

Leukaemia-propagating cells are more frequent in high-risk acute B lymphoblastic leukaemia than in many malignancies that follow a hierarchical cancer stem cell model. It is unclear whether this characteristic can be more universally applied to patients from non-'high-risk' sub-groups and across a broad range of cellular immunophenotypes. Here, we demonstrate in a wide range of primary patient samples and patient samples previously passaged through mice that leukaemia-propagating cells are found in all populations defined by high or low expression of the lymphoid differentiation markers CD10, CD20 or CD34. The frequency of leukaemia-propagating cells and their engraftment kinetics do not differ between these populations. Transcriptomic analysis of CD34(high) and CD34(low) blasts establishes their difference and their similarity to comparable normal progenitors at different stages of B-cell development. However, consistent with the functional similarity of these populations, expression signatures characteristic of leukaemia propagating cells in acute myeloid leukaemia fail to distinguish between the different populations. Together, these findings suggest that there is no stem cell hierarchy in acute B lymphoblastic leukaemia.

We investigated whether, in myelodysplastic syndromes (MDS), aberrant expression of miR-150/miR-221/miR-222 and their designated target mRNA molecules MYB, p27 and c-KIT may be involved in insufficient haematopoiesis. In a series of MDS (n=52), an aberrant increase of miR-150 was found only in MDS with associated del(5q) (n=9; p<0.01). The mRNA expression of transcription factor MYB, the designated target of miR-150, was shown to correlate inversely with the miR-150 level. Acute leukaemia evolving from MDS (n=11) showed significantly decreased levels of miR-221 but not miR-222. We conclude that inhibition of proliferation via over-expressed miR-150 might contribute to myelodysplastic haematopoiesis in MDS-del(5q).

HER2/ERBB2 amplification/overexpression determines the eligibility of breast cancer patients to HER2-targeted therapy. This study evaluates the agreement between ERBB2 copy number assessment by fluorescence in situ hybridization, a standard method recommended by the American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP), and newly available DNA extraction-based methods. A series of n=29 formalin-fixed paraffin-embedded breast cancers were subjected to ERBB2 copy number assessment by fluorescence in situ hybridization (FISH, Vysis, Abbott). Following macrodissection of invasive breast cancer tissue and DNA extraction, ERBB2 copy number was also determined by molecular inversion probe array analysis (MIP, OncoScan, Affymetrix) and next generation sequencing combined with normalized amplicon coverage analysis (NGS/NAC, AmpliSeq, Ion Torrent). ERBB2 copy number values obtained by MIP or NGS/NAC were tightly correlated with ERBB2 copy number values obtained by conventional FISH (rs = 0.940 and rs = 0.894, P < 0.001). Using ASCO/CAP guideline-conform thresholds for categorization of breast cancers as HER2-negative, equivocal or positive, nearly perfect concordance was observed for HER2 classification by FISH and MIP (93% concordant classifications, κ = 0.87). Substantial concordance was observed for FISH and NGS/NAC (83% concordant classifications, κ = 0.62). In conclusion, MIP facilitates precise ERBB2 copy number detection and should be considered as an ancillary method for clinical HER2 testing.

Despite rapid progress in genomic profiling in acute lymphoblastic leukemia (ALL), identification of actionable targets and prediction of response to drugs remains challenging. To identify specific vulnerabilities in ALL, we performed a drug screen using primary human ALL samples cultured in a model of the bone marrow microenvironment combined with high content image analysis. Among the 2487 FDA-approved compounds tested, anthelmintic agents of the class of macrocyclic lactones exhibited potent anti-leukemia activity, similar to the already known anti-leukemia agents currently used in induction chemotherapy. Ex vivo validation in 55 primary ALL samples of both precursor B cell and T-ALL including refractory relapse cases confirmed strong anti-leukemia activity with IC50 values in the low micromolar range. Anthelmintic agents increased intracellular chloride levels in primary leukemia cells, inducing mitochondrial outer membrane depolarization and cell death. Supporting the notion that simultaneously targeting cell death machineries at different angles may enhance the cell death response, combination of anthelmintic agents with the BCL-2 antagonist navitoclax or with the chemotherapeutic agent dexamethasone showed synergistic activity in primary ALL. These data reveal anti-leukemia activity of anthelmintic agents and support exploiting drug repurposing strategies to identify so far unrecognized anti-cancer agents with potential to eradicate even refractory leukemia.

Acute pancreatitis (AP) is a serious, mechanistically not entirely resolved side effect of L-asparaginase-containing treatment for acute lymphoblastic leukemia (ALL). To find new candidate variations for AP, we conducted a genome-wide association study (GWAS).

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 DUX4-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., PAX5 and IKZF1. Moreover, we identified novel fusion partners of lymphoid lineage transcriptional factors ETV6, IKZF1 and PAX5. 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.

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.

B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is a genetically heterogenous malignancy with poor prognosis in relapsed adult patients. The genetic basis for relapse in aneuploid subtypes such as near haploid (NH) and high hyperdiploid (HeH) BCP-ALL is only poorly understood. Pathogenic genetic alterations remain to be identified. To this end, we investigated the dynamics of genetic alterations in a matched initial diagnosis-relapse (ID-REL) BCP-ALL cohort. Here, we firstly report the identification of the novel genetic alteration CYB5Aalt, an alternative transcript of CYB5A, in two independent cohorts.

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., IKZF1, PAX5) 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 JAK2 and NPAT 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.

In this multicenter survey (July 07 to August 08, 2020) in pediatric oncology centers (POCs) belonging to the German Society for Pediatric Oncology and Hematology (GPOH), 36 POCs participated (response rate 70.6%). Home schooling practice was judged as satisfying by 79% prior to and by 38% during the pandemic (P=0.0007). The individual risk of a SARS-CoV-2 infection and the risk of transmission to other patients/caregivers were arguments against attendance. Most POCs recommended regular social participation/school attendance after the end of intensive therapy. 81% stated that persisting restrictions result in serious negative psychosocial consequences for the patients and their families. In-hospital school education, home schooling and re-attendance of school and kindergarten among pediatric cancer patients have suffered a severe setback during the SARS-CoV-2 pandemic. Continuous communication and education concerning protective measures as well as an individual risk assessment are required to avoid the detrimental exclusion of pediatric oncology patients from kindergarten and school.

Acute myeloid leukemia with complex karyotype (CK-AML) is a distinct biological entity associated with a very poor outcome. Since complex karyotypes frequently contain deletions of the chromosomal region 12p13 encompassing the tumor suppressor genes ETV6 and CDKN1B, we aimed to unravel their modes of inactivation in CK-AML.