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The ongoing development of immunotherapies, including chimeric antigen receptor (CAR) T cells, has revolutionized cancer treatment. In paediatric relapsed/refractory B-lineage acute leukaemia antiCD19-CARs induced impressive initial response rates, with event-free survival plateauing at 30-50% in long-term follow-up data. During the interval between diagnosis of relapse or refractoriness and CAR T cell infusion, patients require a bridging therapy. To date, this therapy has consisted of highly variable approaches based on local experience. Here, in an European collaborative effort of paediatric and adult haematologists, we summarise current knowledge with the aim of establishing a guidance for bridging therapy. This includes treatment strategies for different patient subgroups, the advantages and disadvantages of low- and highintensity regimens, and the potential impact of bridging therapy on outcome after CAR T cell infusion. This guidance is a step towards a cross-institutional harmonization of bridging therapy, including personalized approaches. This will allow better comparability of clinical data and increase the level of evidence for the treatment of children and young adults with relapsed/refractory B-lineage ALL until CAR T cell infusion.
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This position paper is based on the authors' many years of clinical experience and basic science research on the diagnosis and treatment of children and adolescents with a presymptomatic early stage of type 1 diabetes. The benefits as well as potential disadvantages of early detection of type 1 diabetes by islet autoantibody screening are critically discussed. In addition, the perspectives of delaying the onset of the clinical metabolic disease through treatment with teplizumab are addressed. Today, we see the chance for a relevant improvement in therapeutic options and life perspectives of affected children and adolescents. Important next steps for the implementation of islet autoantibody screening in Germany are the training of pediatricians who should inform families about the screening, establishment of a few transregional laboratories that carry out the test, and expansion of regional capacities for the training and care of children with an early stage of type 1 diabetes.
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Predisposing CHEK2 germline variants are associated with various adult-type malignancies, whereas their impact on cancer susceptibility in childhood cancer is unclear. To understand the frequency of germline variants in the CHEK2 gene and their impact on pediatric malignancies, we used whole-exome sequencing to search for CHEK2 variants in the germlines of 418 children diagnosed with cancer in our clinics. Moreover, we performed functional analysis of the pathogenic CHEK2 variants to analyze the effect of the alterations on CHK2 protein function. We detected a CHEK2 germline variant in 32/418 (7.7%) pediatric cancer patients and 46.8% of them had leukemia. Functional analysis of the pathogenic variants revealed that 5 pathogenic variants impaired CHK2 protein function. 6/32 patients carried one of these clearly damaging CHEK2 variants and two of them harbored a matching family history of cancer. In conclusion, we detected germline CHEK2 variants in 7.7% of all pediatric cancer patients, of which a minority but still relevant fraction of approximately 20% had a profound impact on protein expression or its phosphorylation after irradiation-induced DNA damage. Accordingly, we conclude that CHEK2 variants increase the risk for not only adult-onset but also pediatric cancer.
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Neoplasias de la Mama , Neoplasias , Adulto , Niño , Femenino , Humanos , Quinasa de Punto de Control 2/genética , Daño del ADN/genética , Predisposición Genética a la Enfermedad , Células Germinativas , Mutación de Línea Germinal , Neoplasias/genéticaRESUMEN
PURPOSE: Clinical checklists are the standard of care to determine whether a child with cancer shows indications for genetic testing. Nevertheless, the efficacy of these tests to reliably detect genetic cancer predisposition in children with cancer is still insufficiently investigated. METHODS: We assessed the validity of clinically recognizable signs to identify cancer predisposition by correlating a state-of-the-art clinical checklist to the corresponding exome sequencing analysis in an unselected single-center cohort of 139 child-parent data sets. RESULTS: In total, one-third of patients had a clinical indication for genetic testing according to current recommendations, and 10.1% (14 of 139) of children harbored a cancer predisposition. Of these, 71.4% (10 of 14) were identified through the clinical checklist. In addition, >2 clinical findings in the checklist increased the likelihood to identifying genetic predisposition from 12.5% to 50%. Furthermore, our data revealed a high rate of genetic predisposition (40%, 4 of 10) in myelodysplastic syndrome cases, while no (likely) pathogenic variants were identified in the sarcoma and lymphoma group. CONCLUSION: In summary, our data show high checklist sensitivity, particularly in identifying childhood cancer predisposition syndromes. Nevertheless, the checklist used here also missed 29% of children with a cancer predisposition, highlighting the drawbacks of sole clinical evaluation and underlining the need for routine germline sequencing in pediatric oncology.
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Neoplasias , Síndromes Neoplásicos Hereditarios , Humanos , Niño , Predisposición Genética a la Enfermedad , Detección Precoz del Cáncer , Neoplasias/diagnóstico , Neoplasias/genética , Neoplasias/patología , Pruebas Genéticas , Genotipo , Síndromes Neoplásicos Hereditarios/diagnóstico , Síndromes Neoplásicos Hereditarios/genética , Mutación de Línea Germinal/genéticaRESUMEN
B-cell precursor acute lymphoblastic leukemia (BCP-ALL) is the most common form of childhood cancer. Chemotherapy is associated with life-long health sequelae and fails in â¼20% of cases. Thus, prevention of leukemia would be preferable to treatment. Childhood leukemia frequently starts before birth, during fetal hematopoiesis. A first genetic hit (eg, the ETV6-RUNX1 gene fusion) leads to the expansion of preleukemic B-cell clones, which are detectable in healthy newborn cord blood (up to 5%). These preleukemic clones give rise to clinically overt leukemia in only â¼0.2% of carriers. Experimental evidence suggests that a major driver of conversion from the preleukemic to the leukemic state is exposure to immune challenges. Novel insights have shed light on immune host responses and how they shape the complex interplay between (1) inherited or acquired genetic predispositions, (2) exposure to infection, and (3) abnormal cytokine release from immunologically untrained cells. Here, we integrate the recently emerging concept of "trained immunity" into existing models of childhood BCP-ALL and suggest future avenues toward leukemia prevention.
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Leucemia-Linfoma Linfoblástico de Células Precursoras B/etiología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/prevención & control , Animales , Niño , Preescolar , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Modelos Animales de Enfermedad , Predisposición Genética a la Enfermedad , Humanos , Inmunidad Innata , Lactante , Infecciones/complicaciones , Infecciones/genética , Infecciones/inmunología , Ratones , Proteínas de Fusión Oncogénica/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/inmunologíaRESUMEN
The impact of LMO2 expression on cell lineage decisions during T-cell leukemogenesis remains largely elusive. Using genetic lineage tracing, we have explored the potential of LMO2 in dictating a T-cell malignant phenotype. We first initiated LMO2 expression in hematopoietic stem/progenitor cells and maintained its expression in all hematopoietic cells. These mice develop exclusively aggressive human-like T-ALL In order to uncover a potential exclusive reprogramming effect of LMO2 in murine hematopoietic stem/progenitor cells, we next showed that transient LMO2 expression is sufficient for oncogenic function and induction of T-ALL The resulting T-ALLs lacked LMO2 and its target-gene expression, and histologically, transcriptionally, and genetically similar to human LMO2-driven T-ALL We next found that during T-ALL development, secondary genomic alterations take place within the thymus. However, the permissiveness for development of T-ALL seems to be associated with wider windows of differentiation than previously appreciated. Restricted Cre-mediated activation of Lmo2 at different stages of B-cell development induces systematically and unexpectedly T-ALL that closely resembled those of their natural counterparts. Together, these results provide a novel paradigm for the generation of tumor T cells through reprogramming in vivo and could be relevant to improve the response of T-ALL to current therapies.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Carcinogénesis , Transformación Celular Neoplásica , Proteínas con Dominio LIM/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patología , Animales , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Células Madre Hematopoyéticas/fisiología , Histocitoquímica , Ratones , Timo/patologíaRESUMEN
The majority of childhood leukemias are precursor B-cell acute lymphoblastic leukemias (pB-ALLs) caused by a combination of prenatal genetic predispositions and oncogenic events occurring after birth. Although genetic predispositions are frequent in children (>1% to 5%), fewer than 1% of genetically predisposed carriers will develop pB-ALL. Although infectious stimuli are believed to play a major role in leukemogenesis, the critical determinants are not well defined. Here, by using murine models of pB-ALL, we show that microbiome disturbances incurred by antibiotic treatment early in life were sufficient to induce leukemia in genetically predisposed mice, even in the absence of infectious stimuli and independent of T cells. By using V4 and full-length 16S ribosomal RNA sequencing of a series of fecal samples, we found that genetic predisposition to pB-ALL (Pax5 heterozygosity or ETV6-RUNX1 fusion) shaped a distinct gut microbiome. Machine learning accurately (96.8%) predicted genetic predisposition using 40 of 3983 amplicon sequence variants as proxies for bacterial species. Transplantation of either wild-type (WT) or Pax5+/- hematopoietic bone marrow cells into WT recipient mice revealed that the microbiome is shaped and determined in a donor genotype-specific manner. Gas chromatography-mass spectrometry (GC-MS) analyses of sera from WT and Pax5+/- mice demonstrated the presence of a genotype-specific distinct metabolomic profile. Taken together, our data indicate that it is a lack of commensal microbiota rather than the presence of specific bacteria that promotes leukemia in genetically predisposed mice. Future large-scale longitudinal studies are required to determine whether targeted microbiome modification in children predisposed to pB-ALL could become a successful prevention strategy.
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Susceptibilidad a Enfermedades , Disbiosis/complicaciones , Heces/microbiología , Microbioma Gastrointestinal , Leucemia Experimental/prevención & control , Factor de Transcripción PAX5/fisiología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/prevención & control , Animales , Femenino , Leucemia Experimental/genética , Leucemia Experimental/microbiología , Leucemia Experimental/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/microbiología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologíaRESUMEN
OBJECTIVE: Recent evidence suggests that illness perceptions in paediatric patients and their parents may differ, with parents holding more negative views compared to their children. Little is known about illness perceptions of very young patients and their parents. This study investigates illness perceptions in paediatric cancer patients aged 4-18 years and their parents in acute treatment or follow-up care, distinguishing patients by age (4-11, 12-18) and stage of medical treatment. METHODS: N = 45 patient-parent dyads in acute treatment and n = 95 dyads in follow-up care were examined. Parents and older children aged 12-18 years completed the Illness Perception Questionnaire-Revised (IPQ-R) and younger children aged 4-11 years were examined using an age-adapted hand puppet interview containing the IPQ-R questions. Difference scores of illness perceptions (symptoms, timeline-acute/chronic, timeline-cyclical, personal control, illness coherence, consequences, emotional representations) between children and parents were tested for significance using Wilcoxon signed-rank tests. RESULTS: Overall, parents perceived more symptoms associated with their child's illness/treatment than the children themselves. In acute treatment, younger children indicated more negative and older children more positive views regarding chronicity than parents. Younger children held less negative views on consequences, and all children reported less negative emotional representations than parents. In follow-up care, all children held less negative views on consequences and emotional representations. Older children reported less negative views on chronicity, cyclicity and illness coherence. CONCLUSION: Differences in illness perceptions of paediatric patients and their parents should be considered during and after treatment/medication and psychosocial care to support illness coping in person- and family-centred interventions.
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Cuidados Posteriores , Neoplasias , Adaptación Psicológica , Adolescente , Niño , Humanos , Neoplasias/terapia , Padres/psicología , Encuestas y CuestionariosRESUMEN
Somatic loss of function mutations in cohesin genes are frequently associated with various cancer types, while cohesin disruption in the germline causes cohesinopathies such as Cornelia-de-Lange syndrome (CdLS). Here, we present the discovery of a recurrent heterozygous RAD21 germline aberration at amino acid position 298 (p.P298S/A) identified in three children with lymphoblastic leukemia or lymphoma in a total dataset of 482 pediatric cancer patients. While RAD21 p.P298S/A did not disrupt the formation of the cohesin complex, it altered RAD21 gene expression, DNA damage response and primary patient fibroblasts showed increased G2/M arrest after irradiation and Mitomycin-C treatment. Subsequent single-cell RNA-sequencing analysis of healthy human bone marrow confirmed the upregulation of distinct cohesin gene patterns during hematopoiesis, highlighting the importance of RAD21 expression within proliferating B- and T-cells. Our clinical and functional data therefore suggest that RAD21 germline variants can predispose to childhood lymphoblastic leukemia or lymphoma without displaying a CdLS phenotype.
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Proteínas de Ciclo Celular , Proteínas de Unión al ADN , Linfoma , Leucemia-Linfoma Linfoblástico de Células Precursoras , Apoptosis , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Niño , Proteínas de Unión al ADN/genética , Síndrome de Cornelia de Lange/genética , Puntos de Control de la Fase G2 del Ciclo Celular , Células Germinativas/metabolismo , Humanos , Linfoma/genética , Mutación , Fenotipo , Leucemia-Linfoma Linfoblástico de Células Precursoras/genéticaRESUMEN
While the shelterin complex guards and coordinates the mechanism of telomere regulation, deregulation of this process is tightly linked to malignant transformation and cancer. Here, we present the novel finding of a germline stop-gain variant (p.Q199*) in the shelterin complex gene POT1, which was identified in a child with acute myeloid leukemia. We show that the cells overexpressing the mutated POT1 display increased DNA damage and chromosomal instabilities compared to the wildtype counterpart. Protein and mRNA expression analyses in the primary patient cells further confirm that, physiologically, the variant leads to a nonfunctional POT1 allele in the patient. Subsequent telomere length measurements in the primary cells carrying heterozygous POT1 p.Q199* as well as POT1 knockdown AML cells revealed telomeric elongation as the main functional effect. These results show a connection between POT1 p.Q199* and telomeric dysregulation and highlight POT1 germline deficiency as a predisposition to myeloid malignancies in childhood.
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Predisposición Genética a la Enfermedad/genética , Leucemia Mieloide Aguda/genética , Trastornos Mieloproliferativos/genética , Complejo Shelterina/genética , Proteínas de Unión a Telómeros/genética , Adulto , Daño del ADN/genética , Células Germinativas , Mutación de Línea Germinal/genética , Células HEK293 , Humanos , Células Mieloides , ARN Mensajero/genética , Telómero/genética , Adulto JovenRESUMEN
Heat shock protein 90 (HSP90) stabilizes many client proteins, including the BCR-ABL1 oncoprotein. BCR-ABL1 is the hallmark of chronic myeloid leukemia (CML) in which treatment-free remission (TFR) is limited, with clinical and economic consequences. Thus, there is an urgent need for novel therapeutics that synergize with current treatment approaches. Several inhibitors targeting the N-terminal domain of HSP90 are under investigation, but side effects such as induction of the heat shock response (HSR) and toxicity have so far precluded their US Food and Drug Administration approval. We have developed a novel inhibitor (aminoxyrone [AX]) of HSP90 function by targeting HSP90 dimerization via the C-terminal domain. This was achieved by structure-based molecular design, chemical synthesis, and functional preclinical in vitro and in vivo validation using CML cell lines and patient-derived CML cells. AX is a promising potential candidate that induces apoptosis in the leukemic stem cell fraction (CD34+CD38-) as well as the leukemic bulk (CD34+CD38+) of primary CML and in tyrosine kinase inhibitor (TKI)-resistant cells. Furthermore, BCR-ABL1 oncoprotein and related pro-oncogenic cellular responses are downregulated, and targeting the HSP90 C terminus by AX does not induce the HSR in vitro and in vivo. We also probed the potential of AX in other therapy-refractory leukemias. Therefore, AX is the first peptidomimetic C-terminal HSP90 inhibitor with the potential to increase TFR in TKI-sensitive and refractory CML patients and also offers a novel therapeutic option for patients with other types of therapy-refractory leukemia because of its low toxicity profile and lack of HSR.
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Antineoplásicos/farmacología , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Proteínas HSP90 de Choque Térmico/química , Respuesta al Choque Térmico/efectos de los fármacos , Mesilato de Imatinib/farmacología , Dominios y Motivos de Interacción de Proteínas , Inhibidores de Proteínas Quinasas/farmacología , Multimerización de Proteína , Animales , Antineoplásicos/química , Sitios de Unión , Biomarcadores de Tumor , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Resistencia a Antineoplásicos/efectos de los fármacos , Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Proteínas de Fusión bcr-abl/química , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Mesilato de Imatinib/química , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Ratones , Modelos Moleculares , Conformación Molecular , Estructura Molecular , Unión Proteica , Inhibidores de Proteínas Quinasas/química , Multimerización de Proteína/efectos de los fármacos , Análisis Espectral , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The homeobox gene HLXB9 encodes for the transcription factor HB9, which is essential for pancreatic as well as motor neuronal development. Beside its physiological expression pattern, aberrant HB9 expression has been observed in several neoplasias. Especially in infant translocation t(7;12) acute myeloid leukemia, aberrant HB9 expression is the only known molecular hallmark and is assumed to be a key factor in leukemic transformation. However, so far, only poor functional data exist addressing the oncogenic potential of HB9 or its influence on hematopoiesis. We investigated the influence of HB9 on cell proliferation and cell cycle in vitro, as well as on hematopoietic stem cell differentiation in vivo using murine and human model systems. In vitro, HB9 expression led to premature senescence in human HT1080 and murine NIH3T3 cells, providing for the first time evidence for an oncogenic potential of HB9. Onset of senescence was characterized by induction of the p53-p21 tumor suppressor network, resulting in growth arrest, accompanied by morphological transformation and expression of senescence-associated ß-galactosidase. In vivo, HB9-transduced primary murine hematopoietic stem and progenitor cells underwent a profound differentiation arrest and accumulated at the megakaryocyte/erythrocyte progenitor stage. In line, gene expression analyses revealed de novo expression of erythropoiesis-related genes in human CD34+hematopoietic stem and progenitor cells upon HB9 expression. In summary, the novel findings of HB9-dependent premature senescence and myeloid-biased perturbed hematopoietic differentiation, for the first time shed light on the oncogenic properties of HB9 in translocation t(7;12) acute myeloid leukemia.
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Ciclo Celular , Diferenciación Celular , Senescencia Celular , Regulación Leucémica de la Expresión Génica , Células Madre Hematopoyéticas/metabolismo , Proteínas de Homeodominio/biosíntesis , Leucemia Mieloide Aguda/metabolismo , Proteínas de Neoplasias/biosíntesis , Factores de Transcripción/biosíntesis , Animales , Eritropoyesis/genética , Células Madre Hematopoyéticas/patología , Proteínas de Homeodominio/genética , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Masculino , Ratones , Células 3T3 NIH , Proteínas de Neoplasias/genética , Factores de Transcripción/genética , Translocación GenéticaRESUMEN
Heritable predisposition is an important cause of cancer in children and adolescents. Although a large number of cancer predisposition genes and their associated syndromes and malignancies have already been described, it appears likely that there are more pediatric cancer patients in whom heritable cancer predisposition syndromes have yet to be recognized. In a consensus meeting in the beginning of 2016, we convened experts in Human Genetics and Pediatric Hematology/Oncology to review the available data, to categorize the large amount of information, and to develop recommendations regarding when a cancer predisposition syndrome should be suspected in a young oncology patient. This review summarizes the current knowledge of cancer predisposition syndromes in pediatric oncology and provides essential information on clinical situations in which a childhood cancer predisposition syndrome should be suspected.
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Predisposición Genética a la Enfermedad , Neoplasias Hematológicas/diagnóstico , Mutación , Proteínas de Neoplasias/genética , Neoplasias/diagnóstico , Adolescente , Niño , Grupos Focales/métodos , Expresión Génica , Asesoramiento Genético/ética , Pruebas Genéticas/métodos , Genética Médica/historia , Genética Médica/instrumentación , Genética Médica/métodos , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/patología , Historia del Siglo XXI , Humanos , Neoplasias/genética , Neoplasias/patología , Sociedades Médicas/historia , SíndromeRESUMEN
Cancer is a clonal malignant disease originated in a single cell and characterized by the accumulation of partially differentiated cells that are phenotypically reminiscent of normal stages of differentiation. According to current models, therapeutic strategies that block oncogene activity are likely to selectively target tumor cells. However, recent evidences have revealed that cancer stem cells could arise through a tumor stem cell reprogramming mechanism, suggesting that genetic lesions that initiate the cancer process might be dispensable for tumor progression and maintenance. This review addresses the impact of these results toward a better understanding of cancer development and proposes new approaches to treat cancer in the future.
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Transformación Celular Neoplásica/genética , Reprogramación Celular/genética , Neoplasias/genética , Células Madre Neoplásicas , Oncogenes/genética , Animales , Diferenciación Celular/genética , Modelos Animales de Enfermedad , Humanos , Ratones , Modelos Biológicos , Neoplasias/terapiaRESUMEN
Acute lymphoblastic leukemia (ALL) accounts for â¼25% of pediatric malignancies. Of interest, the incidence of ALL is observed â¼20% higher in males relative to females. The mechanism behind the phenomenon of sex-specific differences is presently not understood. Employing genome-wide genetic aberration screening in 19 ALL samples, one of the most recurrent lesions identified was monoallelic deletion of the 5' region of SLX4IP. We characterized this deletion by conventional molecular genetic techniques and analyzed its interrelationships with biological and clinical characteristics using specimens and data from 993 pediatric patients enrolled into trial AIEOP-BFM ALL 2000. Deletion of SLX4IP was detected in â¼30% of patients. Breakpoints within SLX4IP were defined to recurrent positions and revealed junctions with typical characteristics of illegitimate V(D)J-mediated recombination. In initial and validation analyses, SLX4IP deletions were significantly associated with male gender and ETV6/RUNX1-rearranged ALL (both overall P < 0.0001). For mechanistic validation, a second recurrent deletion affecting TAL1 and caused by the same molecular mechanism was analyzed in 1149 T-cell ALL patients. Validating a differential role by sex of illegitimate V(D)J-mediated recombination at the TAL1 locus, 128 out of 1149 T-cell ALL samples bore a deletion and males were significantly more often affected (P = 0.002). The repeatedly detected association of SLX4IP deletion with male sex and the extension of the sex bias to deletion of the TAL1 locus suggest that differential illegitimate V(D)J-mediated recombination events at specific loci may contribute to the consistent observation of higher incidence rates of childhood ALL in boys compared with girls.
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Proteínas Portadoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Recombinasas/genética , Recombinación V(D)J , Adolescente , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Niño , Preescolar , Estudios de Cohortes , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Femenino , Eliminación de Gen , Humanos , Lactante , Masculino , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-ets/genética , Proteínas Represoras/genética , Proteína 1 de la Leucemia Linfocítica T Aguda , Proteína ETS de Variante de Translocación 6RESUMEN
BACKGROUND: Recombination-activating gene 1 (RAG1) deficiency results in severe combined immunodeficiency (SCID) caused by a complete lack of T and B lymphocytes. If untreated, patients succumb to recurrent infections. OBJECTIVES: We sought to develop lentiviral gene therapy for RAG1-induced SCID and to test its safety. METHODS: Constructs containing the viral spleen-focus-forming virus (SF), ubiquitous promoters, or cell type-restricted promoters driving sequence-optimized RAG1 were compared for efficacy and safety in sublethally preconditioned Rag1(-/-) mice undergoing transplantation with transduced bone marrow progenitors. RESULTS: Peripheral blood CD3(+) T-cell reconstitution was achieved with SF, ubiquitous promoters, and cell type-restricted promoters but 3- to 18-fold lower than that seen in wild-type mice, and with a compromised CD4(+)/CD8(+) ratio. Mitogen-mediated T-cell responses and T cell-dependent and T cell-independent B-cell responses were not restored, and T-cell receptor patterns were skewed. Reconstitution of mature peripheral blood B cells was approximately 20-fold less for the SF vector than in wild-type mice and often not detectable with the other promoters, and plasma immunoglobulin levels were abnormal. Two months after transplantation, gene therapy-treated mice had rashes with cellular tissue infiltrates, activated peripheral blood CD44(+)CD69(+) T cells, high plasma IgE levels, antibodies against double-stranded DNA, and increased B cell-activating factor levels. Only rather high SF vector copy numbers could boost T- and B-cell reconstitution, but mRNA expression levels during T- and B-cell progenitor stages consistently remained less than wild-type levels. CONCLUSIONS: These results underline that further development is required for improved expression to successfully treat patients with RAG1-induced SCID while maintaining low vector copy numbers and minimizing potential risks, including autoimmune reactions resembling Omenn syndrome.
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Terapia Genética , Vectores Genéticos/genética , Proteínas de Homeodominio/genética , Lentivirus/genética , Inmunodeficiencia Combinada Grave/genética , Inmunodeficiencia Combinada Grave/terapia , Animales , Autoinmunidad/genética , Células de la Médula Ósea/metabolismo , Modelos Animales de Enfermedad , Femenino , Dosificación de Gen , Expresión Génica , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/metabolismo , Humanos , Inmunofenotipificación , Masculino , Ratones , Ratones Noqueados , Fenotipo , Inmunodeficiencia Combinada Grave/inmunología , Bazo/inmunología , Linfocitos T/metabolismo , Timo/inmunología , Transducción Genética , Quimera por TrasplanteRESUMEN
OBJECTIVE: Evaluation of neonatal morbidity after maternal central neurotropic drug exposure. METHODS: Retrospective single-center level-III neonatology cohort analysis of neonates after CND from 2018 to 2021. Control group of neonates born to mothers without CND cared for at the maternity ward. RESULTS: Significantly more frequent therapy need of neonates with CND [OR 23 (95% CI: 7.8-62); RR 14 (95% CI: 5.4-37); p < 0.01]. Neonates after CND had lower Apgar-scores LM 1 [CND 8.1; CG 8.6; p < 0.05]; LM 5 [CND 9; CG 9.7; p < 0.01]; LM 10 [CND 9.6; CG 9.9; p < 0.05]. The first symptom occurred in 95.35% within 24 h (mean: 3.3 h). CND group showed significantly more often preterm delivery [OR 3.5; RR 3.2; p < 0.05], and especially cumulative multiple symptoms [OR 9.4; RR 6.6; p < 0.01] but no correlation to multiple maternal medication use (p = 0.3). CONCLUSIONS: Neonates exposed to CND are at increased risk for postnatal therapy, often due to multiple symptoms. Neonates should be continuously monitored for at least 24 h.
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Enfermedades del Recién Nacido , Humanos , Femenino , Recién Nacido , Estudios Retrospectivos , Embarazo , Adulto , Masculino , Enfermedades del Recién Nacido/epidemiología , Enfermedades del Recién Nacido/inducido químicamente , Fármacos del Sistema Nervioso Central/efectos adversos , Fármacos del Sistema Nervioso Central/uso terapéutico , Efectos Tardíos de la Exposición Prenatal/epidemiología , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Estudios de Casos y Controles , Exposición Materna/efectos adversos , Complicaciones del Embarazo/tratamiento farmacológico , Complicaciones del Embarazo/epidemiologíaRESUMEN
Oncolytic viruses (OVs) selectively replicate in tumor cells resulting in lysis, spreading of new infectious units and induction of antitumor immune responses through abrogating an immunosuppressive tumor microenvironment (TME). Due to their mode of action, OVs are ideal combination partners with targeted immunotherapies. One highly attractive combination is the inhibition of the 'don't-eat-me'-signal CD47, which is known to increase the phagocytic potential of tumor-associated macrophages. In this work, we analyzed the combination approach consisting of the YB-1-based oncolytic adenovirus XVir-N-31 (XVir) and the CD47 inhibitor (CD47i) B6.H12.2 concerning its phagocytic potential. We investigate phagocytosis of XVir-, adenovirus wildtype (AdWT)-, and non-infected established pediatric sarcoma cell lines by different monocytic cells. Phagocytes (immature dendritic cells and macrophages) were derived from THP-1 cells and healthy human donors. Phagocytosis of tumor cells was assessed via FACS analysis in the presence and absence of CD47i. Additional characterization of T cell-stimulatory surface receptors as well as chemo-/cytokine analyses were performed. Furthermore, tumor cells were infected and studied for the surface expression of the 'eat-me'-signal calreticulin (CALR) and the 'don't-eat-me'-signal CD47. We herein demonstrate that (1) XVir-infected tumor cells upregulate both CALR and CD47. XVir induces higher upregulation of CD47 than AdWT. (2) XVir-infection enhances phagocytosis in general and (3) the combination of XVir and CD47i compared to controls showed by far superior enhancement of phagocytosis, tumor cell killing and innate immune activation. In conclusion, the combination of CD47i and XVir causes a significant increase in phagocytosis exceeding the monotherapies considerably accompanied by upregulation of T cell-stimulatory receptor expression and inflammatory chemo/-cytokine secretion.
RESUMEN
Background: Ewing sarcoma (EwS) is an aggressive and highly metastatic bone and soft tissue tumor in pediatric patients and young adults. Cure rates are low when patients present with metastatic or relapsed disease. Therefore, innovative therapy approaches are urgently needed. Cellular- and oncolytic virus-based immunotherapies are on the rise for solid cancers. Methods: Here, we assess the combination of EwS tumor-associated antigen CHM1319-specific TCR-transgenic CD8+ T cells and the YB-1-driven (i.e. E1A13S-deleted) oncolytic adenovirus XVir-N-31 in vitro and in a xenograft mouse model for antitumor activity and immunostimulatory properties. Results: In vitro both approaches specifically kill EwS cell lines in a synergistic manner over controls. This effect was confirmed in vivo, with increased survival using the combination therapy. Further in vitro analyses of immunogenic cell death and antigen presentation confirmed immunostimulatory properties of virus-infected EwS tumor cells. As dendritic cell maturation was also increased by XVir-N-31, we observed superior proliferation of CHM1319-specific TCR-transgenic CD8+ T cells only in virus-tested conditions, emphasizing the superior immune-activating potential of XVir-N-31. Conclusion: Our data prove synergistic antitumor effects in vitro and superior tumor control in a preclinical xenograft setting. Combination strategies of EwS-redirected T cells and YB-1-driven virotherapy are a highly promising immunotherapeutic approach for EwS and warrant further evaluation in a clinical setting.
Asunto(s)
Viroterapia Oncolítica , Sarcoma de Ewing , Humanos , Ratones , Animales , Niño , Linfocitos T CD8-positivos/patología , Xenoinjertos , Modelos Animales de Enfermedad , Animales Modificados Genéticamente , Receptores de Antígenos de Linfocitos T/genética , Factores de TranscripciónRESUMEN
The presence of supernumerary chromosomes is the only abnormality shared by all patients diagnosed with high-hyperdiploid B cell acute lymphoblastic leukemia (HD-ALL). Despite being the most frequently diagnosed pediatric leukemia, the lack of clonal molecular lesions and complete absence of appropriate experimental models have impeded the elucidation of HD-ALL leukemogenesis. Here, we report that for 23 leukemia samples isolated from moribund Eµ-Ret mice, all were characterized by non-random chromosomal gains, involving combinations of trisomy 9, 12, 14, 15, and 17. With a median gain of three chromosomes, leukemia emerged after a prolonged latency from a preleukemic B cell precursor cell population displaying more diverse aneuploidy. Transition from preleukemia to overt disease in Eµ-Ret mice is associated with acquisition of heterogeneous genomic abnormalities affecting the expression of genes implicated in pediatric B-ALL. The development of abnormal centrosomes in parallel with aneuploidy renders both preleukemic and leukemic cells sensitive to inhibitors of centrosome clustering, enabling targeted in vivo depletion of leukemia-propagating cells. This study reveals the Eµ-Ret mouse to be a novel tool for investigating HD-ALL leukemogenesis, including supervision and selection of preleukemic aneuploid clones by the immune system and identification of vulnerabilities that could be targeted to prevent relapse.