Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 44
Filtrar
1.
Haematologica ; 109(1): 72-83, 2024 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-37470150

RESUMEN

Treatment options for relapsed and refractory acute myeloid leukemia patients (R/R AML) are limited. This retrospective cohort study compares safety and efficacy of fludarabine, cytarabine, and idarubicin (FLA-IDA) without or with venetoclax (FLAVIDA) in patients with R/R AML. Thirty-seven and 81 patients received one course FLA-IDA with or without a 7-day course of venetoclax, respectively. The overall response rate (ORR) was significantly higher in FLAVIDA compared to FLAIDA- treated patients (78% vs. 47%; P=0.001), while measurable residual disease was negative at a similar proportion in responding patients (50% vs. 57%), respectively. Eighty-one percent and 79% of patients proceeded to allogeneic hematopoietic cell transplantation or donor lymphocyte infusion after FLAVIDA and FLA-IDA, respectively. Event-free and overall survival were similar in FLAVIDA- and FLA-IDA-treated patients. Refractory patients could be salvaged more successfully after FLA-IDA compared to FLAVIDA pretreatment. Neutrophil and platelet recovery times were similar in the venetoclax and the control group. In conclusion, short-term venetoclax in combination with FLA-IDA represents an effective treatment regimen in R/R AML identifying chemosensitive patients rapidly and inducing measurable residual disease-negative remission in a high proportion of R/R AML patients.


Asunto(s)
Idarrubicina , Leucemia Mieloide Aguda , Humanos , Idarrubicina/uso terapéutico , Citarabina , Estudios Retrospectivos , Factor Estimulante de Colonias de Granulocitos , Leucemia Mieloide Aguda/tratamiento farmacológico , Vidarabina , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos
2.
Cytometry A ; 105(3): 181-195, 2024 03.
Artículo en Inglés | MEDLINE | ID: mdl-37984809

RESUMEN

Multiparameter flow cytometry (MFC) has emerged as a standard method for quantifying measurable residual disease (MRD) in acute myeloid leukemia. However, the limited number of available channels on conventional flow cytometers requires the division of a diagnostic sample into several tubes, restricting the number of cells and the complexity of immunophenotypes that can be analyzed. Full spectrum flow cytometers overcome this limitation by enabling the simultaneous use of up to 40 fluorescent markers. Here, we used this approach to develop a good laboratory practice-conform single-tube 19-color MRD detection assay that complies with recommendations of the European LeukemiaNet Flow-MRD Working Party. We based our assay on clinically-validated antibody clones and evaluated its performance on an IVD-certified full spectrum flow cytometer. We measured MRD and normal bone marrow samples and compared the MRD data to a widely used reference MRD-MFC panel generating highly concordant results. Using our newly developed single-tube panel, we established reference values in healthy bone marrow for 28 consensus leukemia-associated immunophenotypes and introduced a semi-automated dimensionality-reduction, clustering and cell type identification approach that aids the unbiased detection of aberrant cells. In summary, we provide a comprehensive full spectrum MRD-MFC workflow with the potential for rapid implementation for routine diagnostics due to reduced cell requirements and ease of data analysis with increased reproducibility in comparison to conventional FlowMRD routines.


Asunto(s)
Leucemia Mieloide Aguda , Humanos , Citometría de Flujo/métodos , Reproducibilidad de los Resultados , Leucemia Mieloide Aguda/diagnóstico , Médula Ósea/metabolismo , Neoplasia Residual/diagnóstico
3.
Mol Ther Methods Clin Dev ; 30: 515-533, 2023 Sep 14.
Artículo en Inglés | MEDLINE | ID: mdl-37693949

RESUMEN

Safety assessment in retroviral vector-mediated gene therapy remains challenging. In clinical trials for different blood and immune disorders, insertional mutagenesis led to myeloid and lymphoid leukemia. We previously developed the In Vitro Immortalization Assay (IVIM) and Surrogate Assay for Genotoxicity Assessment (SAGA) for pre-clinical genotoxicity prediction of integrating vectors. Murine hematopoietic stem and progenitor cells (mHSPCs) transduced with mutagenic vectors acquire a proliferation advantage under limiting dilution (IVIM) and activate stem cell- and cancer-related transcriptional programs (SAGA). However, both assays present an intrinsic myeloid bias due to culture conditions. To detect lymphoid mutants, we differentiated mHSPCs to mature T cells and analyzed their phenotype, insertion site pattern, and gene expression changes after transduction with retroviral vectors. Mutagenic vectors induced a block in differentiation at an early progenitor stage (double-negative 2) compared to fully differentiated untransduced mock cultures. Arrested samples harbored high-risk insertions close to Lmo2, frequently observed in clinical trials with severe adverse events. Lymphoid insertional mutants displayed a unique gene expression signature identified by SAGA. The gene expression-based highly sensitive molecular readout will broaden our understanding of vector-induced oncogenicity and help in pre-clinical prediction of retroviral genotoxicity.

4.
Mol Cancer Ther ; 22(11): 1290-1303, 2023 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-37643767

RESUMEN

DLBCL are aggressive, rapidly proliferating tumors that critically depend on the ATF4-mediated integrated stress response (ISR) to adapt to stress caused by uncontrolled growth, such as hypoxia, amino acid deprivation, and accumulation of misfolded proteins. Here, we show that ISR hyperactivation is a targetable liability in DLBCL. We describe a novel class of compounds represented by BTM-3528 and BTM-3566, which activate the ISR through the mitochondrial protease OMA1. Treatment of tumor cells with compound leads to OMA1-dependent cleavage of DELE1 and OPA1, mitochondrial fragmentation, activation of the eIF2α-kinase HRI, cell growth arrest, and apoptosis. Activation of OMA1 by BTM-3528 and BTM-3566 is mechanistically distinct from inhibitors of mitochondrial electron transport, as the compounds induce OMA1 activity in the absence of acute changes in respiration. We further identify the mitochondrial protein FAM210B as a negative regulator of BTM-3528 and BTM-3566 activity. Overexpression of FAM210B prevents both OMA1 activation and apoptosis. Notably, FAM210B expression is nearly absent in healthy germinal center B-lymphocytes and in derived B-cell malignancies, revealing a fundamental molecular vulnerability which is targeted by BTM compounds. Both compounds induce rapid apoptosis across diverse DLBCL lines derived from activated B-cell, germinal center B-cell, and MYC-rearranged lymphomas. Once-daily oral dosing of BTM-3566 resulted in complete regression of xenografted human DLBCL SU-DHL-10 cells and complete regression in 6 of 9 DLBCL patient-derived xenografts. BTM-3566 represents a first-of-its kind approach of selectively hyperactivating the mitochondrial ISR for treating DLBCL.


Asunto(s)
Linfoma de Células B , Péptido Hidrolasas , Humanos , Péptido Hidrolasas/metabolismo , GTP Fosfohidrolasas/metabolismo , Mitocondrias/metabolismo , Proteínas Mitocondriales , Linfoma de Células B/tratamiento farmacológico , Linfoma de Células B/genética , Linfoma de Células B/metabolismo
5.
Front Oncol ; 12: 867356, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36059667

RESUMEN

Preemptive and therapeutic donor lymphocyte infusions (preDLI and tDLI) are widely used in relapsing and relapsed hematopoietic malignancies after allogeneic stem cell transplantation (alloSCT) to enhance the graft-versus-malignancy effect. However, in advanced myeloid malignancies, long-term survival after preDLI and tDLI remains low, reflecting our inability to master the double-edged sword of alloreactivity, balancing anti-neoplastic activity versus graft-versus-host disease (GvHD). We previously evaluated a quantitative PCR-based high-sensitivity chimerism (hs-chimerism) based on insertion/deletion polymorphisms instead of short tandem repeats, where increasing host chimerism in peripheral blood predicts relapse more than a month before clinical diagnosis, and declining host chimerism signals anti-host alloreactivity. Here we report 32 consecutive patients with advanced myeloid malignancies receiving preDLI or tDLI "navigated" by hs-chimerism ("navigated DLI"). We compared them to a historical cohort of 110 consecutive preDLI or tDLI recipients, prior to implementation of hs-chimerism at our institution ("controls"). Both groups were comparable regarding age, gender, conditioning, donor type, and time to DLI. With longer median follow-up of the navigated DLI group (8.5 versus 5 months), their landmark overall (64%) and disease-free survival (62%) at 2 years from first DLI compared favorably with controls (23% and 21%, respectively). Improved survival of navigated DLI was due to both reduced relapse incidence (38% versus 60%) and non-relapse mortality (17% versus 44%) at 2 years. Early relapse prediction by hs-chimerism allowed a preemptive approach in 28% of navigated DLI versus 7% in controls. Our results confirm hs-chimerism as a highly valuable tool for monitoring and steering immune interventions after alloSCT.

6.
Hemasphere ; 6(1): e676, 2022 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-34964040

RESUMEN

Measurable residual disease (MRD) quantified by multiparameter flow cytometry (MFC) is a strong and independent prognostic factor in acute myeloid leukemia (AML). However, several technical factors may affect the final read-out of the assay. Experts from the MRD Working Party of the European LeukemiaNet evaluated which aspects are crucial for accurate MFC-MRD measurement. Here, we report on the agreement, obtained via a combination of a cross-sectional questionnaire, live discussions, and a Delphi poll. The recommendations consist of several key issues from bone marrow sampling to final laboratory reporting to ensure quality and reproducibility of results. Furthermore, the experiences were tested by comparing two 8-color MRD panels in multiple laboratories. The results presented here underscore the feasibility and the utility of a harmonized theoretical and practical MFC-MRD assessment and are a next step toward further harmonization.

7.
Blood ; 139(7): 1080-1097, 2022 02 17.
Artículo en Inglés | MEDLINE | ID: mdl-34695195

RESUMEN

In an effort to identify novel drugs targeting fusion-oncogene-induced acute myeloid leukemia (AML), we performed high-resolution proteomic analysis. In AML1-ETO (AE)-driven AML, we uncovered a deregulation of phospholipase C (PLC) signaling. We identified PLCgamma 1 (PLCG1) as a specific target of the AE fusion protein that is induced after AE binding to intergenic regulatory DNA elements. Genetic inactivation of PLCG1 in murine and human AML inhibited AML1-ETO dependent self-renewal programs, leukemic proliferation, and leukemia maintenance in vivo. In contrast, PLCG1 was dispensable for normal hematopoietic stem and progenitor cell function. These findings are extended to and confirmed by pharmacologic perturbation of Ca++-signaling in AML1-ETO AML cells, indicating that the PLCG1 pathway poses an important therapeutic target for AML1-ETO+ leukemic stem cells.


Asunto(s)
Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Regulación Leucémica de la Expresión Génica , Células Madre Hematopoyéticas/patología , Leucemia Mieloide Aguda/patología , Células Madre Neoplásicas/patología , Proteínas de Fusión Oncogénica/metabolismo , Fosfolipasa C gamma/metabolismo , Proteína 1 Compañera de Translocación de RUNX1/metabolismo , Animales , Autorrenovación de las Células , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Células Madre Hematopoyéticas/metabolismo , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Ratones , Células Madre Neoplásicas/metabolismo , Proteínas de Fusión Oncogénica/genética , Fosfolipasa C gamma/genética , Proteoma , Proteína 1 Compañera de Translocación de RUNX1/genética , Transcriptoma , Translocación Genética
9.
Int J Mol Sci ; 22(17)2021 Aug 30.
Artículo en Inglés | MEDLINE | ID: mdl-34502319

RESUMEN

HOXA9 and MEIS1 are frequently upregulated in acute myeloid leukemia (AML), including those with MLL-rearrangement. Because of their pivotal role in hemostasis, HOXA9 and MEIS1 appear non-druggable. We, thus, interrogated gene expression data of pre-leukemic (overexpressing Hoxa9) and leukemogenic (overexpressing Hoxa9 and Meis1; H9M) murine cell lines to identify cancer vulnerabilities. Through gene expression analysis and gene set enrichment analyses, we compiled a list of 15 candidates for functional validation. Using a novel lentiviral multiplexing approach, we selected and tested highly active sgRNAs to knockout candidate genes by CRISPR/Cas9, and subsequently identified a H9M cell growth dependency on the cytosolic phospholipase A2 (PLA2G4A). Similar results were obtained by shRNA-mediated suppression of Pla2g4a. Remarkably, pharmacologic inhibition of PLA2G4A with arachidonyl trifluoromethyl ketone (AACOCF3) accelerated the loss of H9M cells in bulk cultures. Additionally, AACOCF3 treatment of H9M cells reduced colony numbers and colony sizes in methylcellulose. Moreover, AACOCF3 was highly active in human AML with MLL rearrangement, in which PLA2G4A was significantly higher expressed than in AML patients without MLL rearrangement, and is sufficient as an independent prognostic marker. Our work, thus, identifies PLA2G4A as a prognostic marker and potential therapeutic target for H9M-dependent AML with MLL-rearrangement.


Asunto(s)
Biomarcadores de Tumor/metabolismo , Sistemas CRISPR-Cas , Regulación Neoplásica de la Expresión Génica , Fosfolipasas A2 Grupo IV/antagonistas & inhibidores , Proteínas de Homeodominio/metabolismo , Leucemia Mieloide Aguda/patología , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide/metabolismo , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Fosfolipasas A2 Grupo IV/genética , Ensayos Analíticos de Alto Rendimiento , Proteínas de Homeodominio/genética , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide/genética , Células Tumorales Cultivadas
10.
Mol Ther ; 29(12): 3383-3397, 2021 12 01.
Artículo en Inglés | MEDLINE | ID: mdl-34174440

RESUMEN

Hematopoietic stem cell gene therapy is emerging as a promising therapeutic strategy for many diseases of the blood and immune system. However, several individuals who underwent gene therapy in different trials developed hematological malignancies caused by insertional mutagenesis. Preclinical assessment of vector safety remains challenging because there are few reliable assays to screen for potential insertional mutagenesis effects in vitro. Here we demonstrate that genotoxic vectors induce a unique gene expression signature linked to stemness and oncogenesis in transduced murine hematopoietic stem and progenitor cells. Based on this finding, we developed the surrogate assay for genotoxicity assessment (SAGA). SAGA classifies integrating retroviral vectors using machine learning to detect this gene expression signature during the course of in vitro immortalization. On a set of benchmark vectors with known genotoxic potential, SAGA achieved an accuracy of 90.9%. SAGA is more robust and sensitive and faster than previous assays and reliably predicts a mutagenic risk for vectors that led to leukemic severe adverse events in clinical trials. Our work provides a fast and robust tool for preclinical risk assessment of gene therapy vectors, potentially paving the way for safer gene therapy trials.


Asunto(s)
Terapia Genética , Vectores Genéticos , Animales , Daño del ADN , Expresión Génica , Terapia Genética/efectos adversos , Vectores Genéticos/efectos adversos , Vectores Genéticos/genética , Células Madre Hematopoyéticas , Humanos , Aprendizaje Automático , Ratones , Mutagénesis Insercional
11.
Mol Ther ; 29(8): 2535-2553, 2021 08 04.
Artículo en Inglés | MEDLINE | ID: mdl-33831558

RESUMEN

Cellular therapies based on induced pluripotent stem cells (iPSCs) come out of age and an increasing number of clinical trials applying iPSC-based transplants are ongoing or in preparation. Recent studies, however, demonstrated a high number of small-scale mutations in iPSCs. Although the mutational load in iPSCs seems to be largely derived from their parental cells, it is still unknown whether reprogramming may enrich for individual mutations that could lead to loss of functionality and tumor formation from iPSC derivatives. 30 hiPSC lines were analyzed by whole exome sequencing. High accuracy amplicon sequencing showed that all analyzed small-scale variants pre-existed in their parental cells and that individual mutations present in small subpopulations of parental cells become enriched among hiPSC clones during reprogramming. Among those, putatively actionable driver mutations affect genes related to cell-cycle control, cell death, and pluripotency and may confer a selective advantage during reprogramming. Finally, a short hairpin RNA (shRNA)-based experimental approach was applied to provide additional evidence for the individual impact of such genes on the reprogramming efficiency. In conclusion, we show that enriched mutations in curated onco- and tumor suppressor genes may account for an increased tumor risk and impact the clinical value of patient-derived hiPSCs.


Asunto(s)
Células Clonales/citología , Secuenciación del Exoma/métodos , Células Madre Pluripotentes Inducidas/citología , Mutación , Neoplasias/genética , Anciano , Ciclo Celular , Muerte Celular , Diferenciación Celular , Línea Celular , Células Cultivadas , Reprogramación Celular , Células Clonales/química , Células Endoteliales de la Vena Umbilical Humana , Humanos , Células Madre Pluripotentes Inducidas/química , Neoplasias/patología
12.
Sci Rep ; 11(1): 5294, 2021 03 05.
Artículo en Inglés | MEDLINE | ID: mdl-33674652

RESUMEN

Regulation of haematopoietic stem cell fate through conditional gene expression could improve understanding of healthy haematopoietic and leukaemia initiating cell (LIC) biology. We established conditionally immortalised myeloid progenitor cell lines co-expressing constitutive Hoxa9.EGFP and inducible Meis1.dTomato (H9M-ciMP) to study growth behaviour, immunophenotype and morphology under different cytokine/microenvironmental conditions ex vivo upon doxycycline (DOX) induction or removal. The vector design and drug-dependent selection approach identified new retroviral insertion (RVI) sites that potentially collaborate with Meis1/Hoxa9 and define H9M-ciMP fate. For most cell lines, myelomonocytic conditions supported reversible H9M-ciMP differentiation into neutrophils and macrophages with DOX-dependent modulation of Hoxa9/Meis1 and CD11b/Gr-1 expression. Here, up-regulation of Meis1/Hoxa9 promoted reconstitution of exponential expansion of immature H9M-ciMPs after DOX reapplication. Stem cell maintaining conditions supported selective H9M-ciMP exponential growth. H9M-ciMPs that had Ninj2 RVI and were cultured under myelomonocytic or stem cell maintaining conditions revealed the development of DOX-dependent acute myeloid leukaemia in a murine transplantation model. Transcriptional dysregulation of Ninj2 and distal genes surrounding RVI (Rad52, Kdm5a) was detected. All studied H9M-ciMPs demonstrated adaptation to T-lymphoid microenvironmental conditions while maintaining immature myelomonocytic features. Thus, the established system is relevant to leukaemia and stem cell biology.


Asunto(s)
Transformación Celular Neoplásica/efectos de los fármacos , Transformación Celular Neoplásica/genética , Doxiciclina/farmacología , Proteínas de Homeodominio/metabolismo , Leucemia Mieloide Aguda/metabolismo , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide/metabolismo , Células Progenitoras Mieloides/metabolismo , Animales , Proliferación Celular/genética , Trasplante de Células/métodos , Modelos Animales de Enfermedad , Femenino , Vectores Genéticos , Células HEK293 , Proteínas de Homeodominio/genética , Humanos , Leucemia Mieloide Aguda/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide/genética , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Transfección
13.
Internist (Berl) ; 62(7): 768-771, 2021 Jul.
Artículo en Alemán | MEDLINE | ID: mdl-33580307

RESUMEN

This article presents a case of pure red cell aplasia in a 35-year-old heart transplant recipient on chronic hemodialysis. Elevated parvovirus B19 immunoglobulin M blood levels were detected along with a high viral load of 80 billion IU/ml quantified by polymerase chain reaction. Bone marrow examination revealed giant proerythroblasts confirming parvovirus B19 infection. High-dose intravenous immunoglobulin was used for treatment. Anaemia had significantly improved 4 weeks later. Parvovirus B19 infection should be excluded in organ transplant recipients with anaemia due to ineffective erythropoiesis.


Asunto(s)
Anemia Refractaria , Trasplante de Corazón , Infecciones por Parvoviridae , Parvovirus B19 Humano , Adulto , Trasplante de Corazón/efectos adversos , Humanos , Inmunoglobulinas Intravenosas , Infecciones por Parvoviridae/diagnóstico , Infecciones por Parvoviridae/terapia , Diálisis Renal/efectos adversos
16.
Leukemia ; 34(11): 2951-2963, 2020 11.
Artículo en Inglés | MEDLINE | ID: mdl-32576961

RESUMEN

To establish novel and effective treatment combinations for chronic myelomonocytic leukemia (CMML) preclinically, we hypothesized that supplementation of CMML cells with the human oncogene Meningioma 1 (MN1) promotes expansion and serial transplantability in mice, while maintaining the functional dependencies of these cells on their original genetic profile. Using lentiviral expression of MN1 for oncogenic supplementation and transplanting transduced primary mononuclear CMML cells into immunocompromised mice, we established three serially transplantable CMML-PDX models with disease-related gene mutations that recapitulate the disease in vivo. Ectopic MN1 expression was confirmed to enhance the proliferation of CMML cells, which otherwise did not engraft upon secondary transplantation. Furthermore, MN1-supplemented CMML cells were serially transplantable into recipient mice up to 5 generations. This robust engraftment enabled an in vivo RNA interference screening targeting CMML-related mutated genes including NRAS, confirming that their functional relevance is preserved in the presence of MN1. The novel combination treatment with azacitidine and the MEK-inhibitor trametinib additively inhibited ERK-phosphorylation and thus depleted the signal from mutated NRAS. The combination treatment significantly prolonged survival of CMML mice compared to single-agent treatment. Thus, we identified the combination of azacitidine and trametinib as an effective treatment in NRAS-mutated CMML and propose its clinical development.


Asunto(s)
Antineoplásicos/farmacología , Evaluación Preclínica de Medicamentos , Leucemia Mielomonocítica Crónica/tratamiento farmacológico , Ensayos Antitumor por Modelo de Xenoinjerto , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Azacitidina/farmacología , Evolución Clonal , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos/métodos , Evaluación Preclínica de Medicamentos/normas , Sinergismo Farmacológico , Femenino , GTP Fosfohidrolasas/genética , Humanos , Leucemia Mielomonocítica Crónica/genética , Leucemia Mielomonocítica Crónica/mortalidad , Leucemia Mielomonocítica Crónica/patología , Proteínas de la Membrana/genética , Ratones , Mutación , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Piridonas/farmacología , Piridonas/uso terapéutico , Pirimidinonas/farmacología , Pirimidinonas/uso terapéutico , ARN Interferente Pequeño/genética , Receptor Notch1/genética , Ensayos Antitumor por Modelo de Xenoinjerto/métodos
17.
Stem Cell Reports ; 14(1): 122-137, 2020 01 14.
Artículo en Inglés | MEDLINE | ID: mdl-31839543

RESUMEN

Induced pluripotent stem cells (iPSCs) offer a promising platform to model early embryonic developmental processes, to create disease models that can be evaluated by drug screens as well as proof-of-concept experiments for regenerative medicine. However, generation of iPSC-derived hemato-endothelial and hematopoietic progenitor cells for these applications is challenging due to variable and limited cell numbers, which necessitates enormous up-scaling or development of demanding protocols. Here, we unravel the function of key transcriptional regulators SCL, LMO2, GATA2, and ETV2 (SLGE) on early hemato-endothelial specification and establish a fully inducible and stepwise hemato-endothelial forward programming system based on SLGE-regulated overexpression. Regulated induction of SLGE in stable SLGE-iPSC lines drives very efficient generation of large numbers of hemato-endothelial progenitor cells (CD144+/CD73-), which produce hematopoietic progenitor cells (CD45+/CD34+/CD38-/CD45RA-/CD90+/CD49f+) through a gradual process of endothelial-to-hematopoietic transition (EHT).

18.
Blood Adv ; 3(24): 4252-4263, 2019 12 23.
Artículo en Inglés | MEDLINE | ID: mdl-31867596

RESUMEN

HOX genes are highly conserved, and their precisely controlled expression is crucial for normal hematopoiesis. Accordingly, deregulation of HOX genes can cause leukemia. However, despite of intensive research on the coding HOX genes, the role of the numerous long noncoding RNAs (lncRNAs) within the HOX clusters during hematopoiesis and their contribution to leukemogenesis are incompletely understood. Here, we show that the lncRNA HOXA10-AS, located antisense to HOXA10 and mir-196b in the HOXA cluster, is highly expressed in hematopoietic stem cells (HSCs) as well as in KMT2A-rearranged and NPM1 mutated acute myeloid leukemias (AMLs). Using short hairpin RNA- and locked nucleic acid-conjugated chimeric antisense oligonucleotide (LNA-GapmeR)-mediated HOXA10-AS-knockdown and CRISPR/Cas9-mediated excision in vitro, we demonstrate that HOXA10-AS acts as an oncogene in KMT2A-rearranged AML. Moreover, HOXA10-AS knockdown severely impairs the leukemic growth of KMT2A-rearranged patient-derived xenografts in vivo, while high HOXA10-AS expression can serve as a marker of poor prognosis in AML patients. Lentiviral expression of HOXA10-AS blocks normal monocytic differentiation of human CD34+ hematopoietic stem and progenitor cells. Mechanistically, we show that HOXA10-AS localizes in the cytoplasm and acts in trans to induce NF-κB target genes. In total, our data imply that the normally HSC-specific HOXA10-AS is an oncogenic lncRNA in KMT2A-r AML. Thus, it may also represent a potential therapeutic target in KMT2A-rearranged AML.


Asunto(s)
Reordenamiento Génico , N-Metiltransferasa de Histona-Lisina/genética , Proteínas Homeobox A10/genética , Leucemia/genética , Proteína de la Leucemia Mieloide-Linfoide/genética , Células Madre Neoplásicas/metabolismo , ARN sin Sentido , ARN Largo no Codificante , Animales , Modelos Animales de Enfermedad , Citometría de Flujo , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Leucemia/diagnóstico , Leucemia/mortalidad , Leucemia/terapia , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Ratones , FN-kappa B/metabolismo , Células Madre Neoplásicas/patología , Nucleofosmina , Pronóstico , Transcriptoma
20.
Cancer Cell ; 36(2): 123-138.e10, 2019 08 12.
Artículo en Inglés | MEDLINE | ID: mdl-31303423

RESUMEN

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


Asunto(s)
Biomarcadores de Tumor/genética , Transformación Celular Neoplásica/genética , Cromosomas Humanos Par 21 , Subunidad beta Común de los Receptores de Citocinas/genética , Síndrome de Down/genética , Factor de Transcripción GATA1/genética , Leucemia Mieloide/genética , Reacción Leucemoide/genética , Mutación , Animales , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Síndrome de Down/diagnóstico , Factor de Transcripción GATA1/metabolismo , Regulación Leucémica de la Expresión Génica , Predisposición Genética a la Enfermedad , Células HEK293 , Humanos , Leucemia Mieloide/diagnóstico , Leucemia Mieloide/patología , Reacción Leucemoide/diagnóstico , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones Transgénicos , Fenotipo , Transcripción Genética
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA