BACKGROUND:Central nervous system (CNS) involvement in childhood acute lymphoblastic leukemia (ALL) is assessed by cell counting and cytomorphology from cerebrospinal fluid (CSF) and is used for treatment stratification worldwide. The ratio of "CNS2" patients in clinical trials ranges from 3% to 40%, with unclear prognostic significance. PROCEDURE:To assess real-world practice, a survey was distributed globally. Questions focused on the systemic treatment allowed before the staging lumbar puncture, sample handling, methodological details of different analytics, and staging definitions. RESULTS:Eighty-two centers from 47 countries from 5 continents responded. Contraindications and timing of sampling and applied CSF test modalities are heterogeneous. CSF volumes used for cytospins and flow cytometry range widely (15-3000 and 200-4000 µL, respectively). Test positivity definitions vary between ≥1 and ≥5 identified blasts for cytospin and ≥1 to ≥50 blasts for flow cytometry. Differences in practice were seen between countries in the same ALL consortium and within individual countries. CONCLUSIONS:The observed heterogeneity impacts CNS staging and treatment decisions worldwide. Our results may explain the conflicting published evidence on the prognostic value of minimal leukemic CNS involvement. A global consensus on CNS diagnostics and a more detailed reporting of CNS staging methods in clinical studies are urgently needed.

Monitoring MYCN-driven high-risk neuroblastoma presents challenges to capture dynamics of all tumor cell clones at their earliest divergence to current clinical course. Not all clones may enter the bone marrow, the most important monitoring site for minimal residual disease (MRD), causing relapse in ∼50% of patients. We developed mediator-probe PCR assays to detect up to four multiplexed patient-individual genetic alterations in 37 longitudinally collected bone marrow aspirates from 8 patients with MYCN-amplified disease. Multiplexed biomarkers, including MYCN amplicon breakpoints, detected diverse neuroblastoma clones, surpassing conventional GD2 immunocytology accuracy. We provide proof-of-principle for clonally heterogeneous MRD biomarker detection (1 tumor: 106 reference cells). In selected patients, multiplexed biomarkers indicated divergent dynamics, suggesting individual tumor clones differ in their ability to disseminate to the bone marrow and escape therapy. Our pilot data support integrating multiplexed MRD detection in co-clinical trials to monitor the molecular remission state during therapy and follow-up.

BackgroundCREB binding protein (CREBBP) is a key epigenetic regulator, altered in a fifth of relapsed cases of acute lymphoblastic leukemia (ALL). Selectively targeting epigenetic signaling may be an effective novel therapeutic approach to overcome drug resistance. Anti-tumor effects have previously been demonstrated for GSK-J4, a selective H3K27 histone demethylase inhibitor, in several animal models of cancers.MethodsTo characterize the effect of GSK-J4, drug response profiling, CRISPR-Dropout Screening, BH3 profiling and immunoblotting were carried out in ALL cell lines or patient derived samples.ResultsHere we provide evidence that GSK-J4 downregulates cyclic AMP-responsive element-binding protein (CREB) and CREBBP in B-cell precursor-ALL cell lines and patient samples. High CREBBP expression in BCP-ALL cell lines correlated with high GSK-J4 sensitivity and low dexamethasone sensitivity. GSK-J4 treatment also induced Bcl-2 and Bcl-XL dependency and apoptosis.ConclusionsThis study proposes H3K27 demethylase inhibition as a potential treatment strategy for patients with treatment-resistant ALL, using CREBBP as a biomarker for drug response and combining GSK-J4 with venetoclax and navitoclax as synergistic partners.

A ddPCR assay detecting unique TERT rearrangement breakpoints for improved in-time molecular disease monitoring. A, Representative original data output from the uniplex ddPCR assay designed to quantify copies of the TERT breakpoint unique to the GI-ME-N cell line (left). The 1D amplitude plot depicts event number (droplets) versus fluorescence amplitude with positive (blue) and negative (gray) droplets separated by the threshold line (pink). Quantification of TERT breakpoint copies detected in GI-ME-N cells grown in vitro, conditioned cell culture medium, and subcutaneous GI-ME-N xenografts grown in mice (mean ± SD, n ≥ 3). B, The TERT rearrangement breakpoint copies detected in genomic DNA from neuroblastoma tissue samples by ddPCR are shown for patient P1. Neg. co., negative control; NTC, nontemplate control; M, metastasis; PT, primary tumor.

Abstract This photographic essay explores solid waste management and recycling practices among residents of Zaandam and other towns in the Netherlands. The study was based on ethnographic research conducted in September 2019 and December 2021 that included street ethnography, landscape observation, and recorded interviews. The photo essay presents a narrative of the researcher’s observations and experiences, highlighting the challenges and successes of waste management and recycling practices in the Netherlands.

T-ALL relapses are characterized by chemotherapy resistance, cellular diversity and dismal outcome. To gain a deeper understanding of the mechanisms underlying relapses, we conduct single-cell RNA sequencing on 13 matched pediatric T-ALL patient-derived samples at diagnosis and relapse, along with samples derived from 5 non-relapsing patients collected at diagnosis. This comprehensive longitudinal single-cell study in T-ALL reveals significant transcriptomic diversity. Notably, 11 out of 18 samples exhibit a subpopulation of T-ALL cells with stem-like features characterized by a common set of active regulons, expression patterns and splice isoforms. This subpopulation, accounting for a small proportion of leukemia cells at diagnosis, expands substantially at relapse, indicating resistance to therapy. Strikingly, increased stemness at diagnosis is associated with higher risk of treatment induction failure. Chemotherapy resistance is validated through in-vitro and in-vivo drug testing. Thus, we report the discovery of treatment-resistant stem-like cells in T-ALL, underscoring the potential for devising future therapeutic strategies targeting stemness-related pathways.

ABSTRACT:Telomerase is reactivated by genomic TERT rearrangements in ∼30% of diagnosed high-risk neuroblastomas. Dismal patient prognosis results if the RAS/MAPK/ALK signaling transduction network also harbors mutations. We present a liquid biopsy–based monitoring strategy for this particularly vulnerable pediatric patient subgroup for which real-time molecular diagnostic tools are limited to date. Droplet digital PCR assays quantifying patient-individualized TERT rearrangement breakpoint copies, ALK copy numbers, and allelic ALK p.R1275Q mutation frequencies were applied to longitudinally collected liquid biopsies (peripheral blood and bone marrow [BM] plasma, n = 44 biosamples), the mononuclear cell fraction from BM and matched tumor samples. Marker detection was compared with current gold-standard diagnostics. Reanalysis of whole-genome and targeted panel sequencing data from 169 patients identified 64 TERT-rearranged neuroblastoma samples collected at initial and/or relapse diagnosis from 55 patients (254 total TERT rearrangement events). Detection and quantification of unique TERT rearrangement breakpoints in as little as 1 ng of total cell-free DNA in peripheral blood plasma improved therapy response assessment and early relapse detection in individual patients. Proof-of-concept is provided for minimal residual disease detection in the BM niche, from which relapses frequently arise, by analyzing unique TERT rearrangement breakpoints in BM plasma–derived cell-free DNA. TERT rearrangement breakpoints, as a single marker or combined with mutations in the RAS/MAPK/ALK signaling transduction network, can serve as robust and highly sensitive biomarkers for disease activity and spatially and temporally resolve disease better than current gold-standard diagnostics in individual patients with TERT-driven neuroblastoma. SIGNIFICANCE:Real-time molecular monitoring of TERT-rearranged high-risk neuroblastoma is an unmet clinical need. We tested liquid biopsy-based assays for patient-individualized TERT breakpoint sequences to monitor disease in pediatric patients. Our digital PCR approach provides high resolution of spatial and temporal disease quantification in individual patients and is applicable for clinical routine.

No ano de 2023 a Associação Brasileira de Antropologia comemorou os 70 anos de Reuniões Brasileiras de Antropologias, as RBAs, e promoveu uma série de eventos em parceria com Programas de Pós-graduação em Antropologia, Ciências Sociais e Sociologia e Antropologia. O livro reúne participações e trabalhos apresentados nestes eventos comemorativos que trazem um testemunho sobre a história da antropologia brasileira em diversos contextos institucionais pelo país.

O capítulo trata das comemorações dos 50 anos de Reuniões Brasileiras de Antropologia e das comemorações dos 50 anos da Associação Brasileira de Antropologia, tendo por fonte o livro publicado e intitulado Homenagens de 2006, com selo ABA.

O artigo contempla os temas da ciência aberta, das políticas de memória e do patrimônio cultural nas cidades brasileiras como parte dos processos de acessibilidade às comunidades urbanas do país através da comunicação pública dos dados das pesquisas sobre seus patrimônios culturais e históricos. Tais estudos vêm sendo desenvolvidos no interior do Banco de Imagens e Efeitos Visuais, projeto de pesquisa do Programa de Pós Graduação em Antropologia Social na Universidade Federal do Rio Grande do Sul. Trata-se de aprofundar os dilemas enfrentados pela comunidade científica tanto no que se refere à construção de repositórios digitais de pesquisa etnográfica na área de Antropologia Urbana quanto a sua divulgação, que deve visar não apenas a redenacional de museus e centros de documentação do país, mas também um público mais amplo, via redes digitais e eletrônicas. Objetiva-se ampliar a pesquisa etnográfica em hipermídia e contribuir para os estudos de patrimonialização da produção antropológica audiovisual, considerando-a como portadora de um valor cultural específico. Esse processo complexo, importante e fundamental já vem sendo realizado no âmbito dos ateliês de produção e criação do BIEV, sob a supervisão das coordenadoras do grupo de pesquisa, com a participação de bolsistas e com o acompanhamento de pesquisadores associados(as) com formação em web design. Os acervos sob a guarda de laboratórios e núcleos de pesquisa em Antropologia Visual, como o caso do BIEV, concentram produções culturais que resultam das relações de colaboração dos pesquisadores com seus parceiros de pesquisa, devendo, portanto, a eles retornar. PALAVRAS-CHAVE: Repositórios digitais. Restituição. Etnografia. Imagem. Cidade

BACKGROUND In childhood acute lymphoblastic leukemia (ALL), central nervous system (CNS) involvement is associated with increased risk of relapse. Traditionally, CNS involvement has been staged using cell counting and microscopic cytomorphological assessment of centrifuged and stained samples (cytospins) of the cerebrospinal fluid (CSF). More recently CSF flow cytometry has been shown to be more sensitive than cytology and to have prognostic significance. CNS staging is a basis for treatment stratification worldwide. CSF diagnostic protocols of study groups differ in small details, however, larger variability in practice was suspected based on informal discussions within the childhood ALL community. AIMS To systematically evaluate real world practice of CSF diagnostics and staging in pediatric ALL patients across hospitals globally. METHODS A survey was developed and distributed in 2021-2022 with target hospital categories and hospital numbers per country. Questions focused on numerous details of routine practice at the given hospitals in year 2021. RESULTS Eighty-two centers from 47 countries across five continents responded with a median 20 new pediatric ALL patients per year per center. Significant heterogeneity was observed in testing methodologies and CNS staging. The staging LP is contraindicated by high peripheral WBC, with threshold varying between 50 and 200 x109/L in 46% of the centers, while 54% of the centers set no threshold (LP is performed even in case of extreme leukocytosis). The timing of staging LP relative to the initiation of systemic antileukemic therapy also varies, with 57% of hospitals performing the LP strictly before therapy, while others routinely allowing some (1 and 7 days depending on hospital) systemic steroid and/or chemotherapy before. There is heterogeneity in the selection of test modalities to analyze the CSF. Manual cell counting is applied in 73%, automated cell counting in 43%, cytospins in 84% and flow cytometry in 33% of the hospitals during routine CNS-staging. Ten percent of the centers use only one modality, 54% of them use 2 modalities. A larger proportion of centers in high income countries analyze WBCs in cytospins routinely as compared to centers in middle income countries (92% versus 65%, p = 0.006). A similar trend was observed regarding CSF flow cytometry (39% vs 17%, p = 0.07). Similarly, a heterogenous set of testing modalities and cut offs/definitions are applied for CSF red cell quantification and for defining traumatic LP (TLP). Among hospitals which assess cytospins, the CSF volume centrifuged to prepare these specimens varies between 15 and 3000 microliters. Typical CSF volumes analyzed by flow cytometry range between 200 and 4000 microliters. There is also wide variation regarding the cut-off blast numbers to define positive findings in case of cytospins (≥1 to ≥5 cells) and flow cytometry (1 to 50 events characterized as test positivity in different centers). Interestingly, there is no correlation between the CSF volume used and the minimum number of blasts to define the tests' positivity. These heterogeneities are seen both within study groups and within individual countries in the same study group. DISCUSSION The above inconsistencies and differences in practice must have obvious clinical implications by significantly impacting CNS staging and thus CNS1/2/3/TLP frequencies and therefore treatment decisions for ALL patients worldwide. Moreover, internationally well-connected, larger pediatric oncology centers involved in research are thought to be overrepresented among survey participants. In view of this bias, global diagnostic heterogeneity can be larger and the quality of diagnostics poorer for a large proportion of patients than what our survey reflects. These results raise concern over published findings on the prognostic value of leukemic CNS involvement, particularly the CNS2 stage, and the comparability of published studies. We propose that details on CNS staging methods and adherence to the diagnostic guidelines should be included in publications of clinical studies. There is an urgent need for standardization, ideally through the development of international consensus guidelines in this field.

Background: T-cell lymphoblastic lymphoma (T-LBL) and acute lymphoblastic leukemia (T-ALL) originate from the malignant transformation of similar subsets of immature T-cell precursors in the thymus. Unique genetic and epigenetic aberrations lead to distinct clinical presentations. Both are distinguished by bone marrow infiltration. Patient's age at diagnosis is associated with the distribution of molecular subgroups in T-ALL (Neumann, et al. Leukemia, 2024). Despite sharing many clinical features and molecular alterations, there is ongoing debate about whether T-LBL and T-ALL represent a spectrum of a disease or are distinct entities. Recent papers identified TRB::NOTCH1 fusions exclusively in T-LBL and associated with high relapse risk (te Vrugt, et al. Blood, 2024). Both diseases are characterized by poor survival in resistant or relapsed cases, highlighting a clear unmet medical need. Understanding varying therapy resistance and molecular pathogenesis across ages is crucial. Clonal evolution analysis can help reconstruct tumor development, explore treatment responses, and identify reasons for therapy failure (Sandmann, et al. Int J Environ Res Public Health, 2023). This study compares DNA samples from pediatric and adult T-LBL and T-ALL patients to uncover genetic differences and tackle therapy resistance. Methods: DNA samples include material from initial diagnosis, corresponding germline, and, if applicable, relapse. Subcohorts include 87 pediatric T-LBL cases (27 relapsed, 60 non-relapsed), 36 pediatric T-ALL (12 relapsed, 24 non-relapsed), 47 adult T-LBL (of which 3 corresponding relapse samples were available), and 41 adult T-ALL, (of which 4 corresponding relapse samples were available). Besides targeted sequencing, CNV analysis is performed by Illumina Infinium Global Screening Array v3.0. Results and Discussion: For T-LBL and T-ALL, there were no significant differences in the average number of single nucleotide variants (SNV) per sample, either within or between the groups. For CNVs, a clear increase with age was observed for both T-LBL and T-ALL, except deletions and LOH affecting chromosome 9, which were more frequent in pediatric cases and decreased with age. Detailed analysis revealed subgroup-specific characteristics in initial samples. In adult T-LBL vs pediatric T-LBL samples, variants in PHF6 (34% vs 13%), JAK1 (9% vs 0%), TP53 (6% vs 0%), NOTCH3 (11% vs 5%), JAK3 (11% vs 5%), STAT5B (11% vs 5%), TET2 (6% vs 1%), and NRAS (9% vs 3%) were more prevalent. NOTCH1 (60% vs 49%), PIK3CA (10% vs 2%), PTEN (15% vs 9%), PIK3R1 (8% vs 3%), CCND3 (6% vs 0%), and KMT2C (6% vs 0%) were more frequent in pediatric T-LBL than adult T-LBL patients. In the T-ALL cohort, some variants, such as DNM2 (17%) and RUNX1 (10%) were almost exclusively found in adult cases. The increased prevalence of TP53 mutations in pediatric relapse samples underscored the critical role of TP53 in treatment resistance and disease progression (T-ALL: 8% primary vs 29% relapse; T-LBL: 0% primary vs 13% relapse). Clonal evolution analysis showed that CNVs appeared early, especially deletions or LOH affecting chromosome 9. Subsequently, additional subclones with point mutations evolved. Pediatric cases revealed differential timing and frequency of mutations in T-LBL vs in T-ALL: MYB (nested level 4.6 vs 1.0), SMARCA4 (4.7 vs 1.5), or TET2 (4.0 vs 1.0). In pediatric T-LBL, deletions and LOH on chromosome 9 occurred equally in primary samples of patients who suffered/did not suffer relapse. In relapse samples, the clonal population with CNV increased, indicating a growth advantage. Frequent CNVs in chromosome 9, encompassing genes like CDKN2A/B and MTAP, underscored the importance of this region in both T-LBL and T-ALL. These CNVs, involving tumor suppressor genes such as p16INK4a and p14ARF, were early events in clonal evolution. Conclusion: Our integrated genomic analysis highlights differences and similarities between pediatric and adult T-LBL and T-ALL cases. The analysis of CNVs revealed their role in clonal evolution, particularly with frequent deletions and LOH affecting chromosome 9 in pediatric cases. This emphasize the importance of genomic profiling for each subgroup. By understanding distinct genetic landscapes and mutation patterns, we aim to discover potential novel targets to improve risk group stratification of patients with T-LBL and T-ALL.

Minimal/measurable residual disease (MRD) diagnostics using real-time quantitative PCR analysis of rearranged immunoglobulin and T-cell receptor gene rearrangements are nowadays implemented in most treatment protocols for patients with acute lymphoblastic leukemia (ALL). Within the EuroMRD Consortium, we aim to provide comparable, high-quality MRD diagnostics, allowing appropriate risk-group classification for patients and inter-protocol comparisons. To this end, we set up a quality assessment scheme, that was gradually optimized and updated over the last 20 years, and that now includes participants from around 70 laboratories worldwide. We here describe the design and analysis of our quality assessment scheme. In addition, we here report revised data interpretation guidelines, based on our newly generated data and extensive discussions between experts. The main novelty is the partial re-definition of the “positive below quantitative range” category by two new categories, “MRD low positive, below quantitative range” and “MRD of uncertain significance”. The quality assessment program and revised guidelines will ensure reproducible and accurate MRD data for ALL patients. Within the Consortium, similar programs and guidelines have been introduced for other lymphoid diseases (e.g., B-cell lymphoma), for new technological platforms (e.g., digital droplet PCR or Next-Generation Sequencing), and for other patient-specific MRD PCR-based targets (e.g., fusion genes).

KMT2A-rearranged acute lymphoblastic infant leukemia (KMT2A-r iALL) is associated with outsize risk of relapse and relapse mortality. We previously reported strong upregulation of the immediate early gene EGR3 in KMT2A::AFF1 iALL at relapse; now we provide analyses of the EGR3 regulome, which we assessed through binding and expression target analysis of an EGR3-overexpressing t(4;11) cell culture model. Our data identify EGR3 as a regulator of early B-lineage commitment. Principal component analysis of 50 KMT2A-r iALL patients at diagnosis and 18 at relapse provided strictly dichotomous separation of patients based on the expression of four B-lineage genes. Absence of B-lineage gene expression translates to more than two-fold poorer long-term event-free survival. In conclusion, our study presents four B-lineage genes with prognostic significance, suitable for gene expression-based risk stratification of KMT2A-r iALL patients.

Infant acute lymphoblastic leukemia is a devastating disease which arises mainly due to the occurrence of KMT2A rearrangements (KMT2A-r). In this context, risk stratification is an indispensable pre-treatment step enabling assignment of patients to appropriate intervention subgroups. KMT2A-r, young age, poor Glucocorticoid response rates, and high blast counts are the primarily applied risk-determining factors. Our group recently described 100-fold increased expression of the immediate early gene EGR3 at time of relapse compared to diagnosis. Here, we present the EGR3 regulome of infant KMT2A-r ALL which was assessed through integration of ChIP-seq and MACE-seq data of an EGR3-overexpressing KMT2A::AFF1 cell culture model. Binding and expression target analysis followed by gene set enrichment analysis, motif scanning, and flow cytometry identified EGR3 as a mediator of B lineage commitment. Moreover, EGR3 was found to collaborate with a set of B lineage transcription factors including PAX5. Gene expression analysis of patient cohorts from diagnosis (n=50) and relapse (n=18) uncovered strict bimodal clustering of patients determined by four EGR3-regulated B lineage marker genes. Although the whole diagnosis cohort was assigned to the high-risk group, event free survival analysis of patient clusters demonstrated absence of B lineage gene expression translating to clearly inferior long-term event-free survival. Possible confounders including age, blast percentage, and HOXA status did not affect dichotomous separation of patients. In conclusion, our study identified EGR3 as a mediator of B lineage specification and commitment in infant KMT2A::AFF1 ALL and characterized the EGR3 regulome in detail. Furthermore, we present four B lineage markers whose gene expression can be applied for substratification of high-risk patients potentially improving clinical decision making. Infant acute lymphoblastic leukemia is a devastating disease which arises mainly due to the occurrence of KMT2A rearrangements (KMT2A-r). In this context, risk stratification is an indispensable pre-treatment step enabling assignment of patients to appropriate intervention subgroups. KMT2A-r, young age, poor Glucocorticoid response rates, and high blast counts are the primarily applied risk-determining factors. Our group recently described 100-fold increased expression of the immediate early gene EGR3 at time of relapse compared to diagnosis. Here, we present the EGR3 regulome of infant KMT2A-r ALL which was assessed through integration of ChIP-seq and MACE-seq data of an EGR3-overexpressing KMT2A::AFF1 cell culture model. Binding and expression target analysis followed by gene set enrichment analysis, motif scanning, and flow cytometry identified EGR3 as a mediator of B lineage commitment. Moreover, EGR3 was found to collaborate with a set of B lineage transcription factors including PAX5. Gene expression analysis of patient cohorts from diagnosis (n=50) and relapse (n=18) uncovered strict bimodal clustering of patients determined by four EGR3-regulated B lineage marker genes. Although the whole diagnosis cohort was assigned to the high-risk group, event free survival analysis of patient clusters demonstrated absence of B lineage gene expression translating to clearly inferior long-term event-free survival. Possible confounders including age, blast percentage, and HOXA status did not affect dichotomous separation of patients. In conclusion, our study identified EGR3 as a mediator of B lineage specification and commitment in infant KMT2A::AFF1 ALL and characterized the EGR3 regulome in detail. Furthermore, we present four B lineage markers whose gene expression can be applied for substratification of high-risk patients potentially improving clinical decision making.