RESUMO
Accurately predicting patient outcomes is essential for optimizing treatment and improving outcomes in pediatric acute myeloid leukemia (AML). In recent years, microRNAs have emerged as a promising prognostic marker, with a growing body of evidence supporting their potential predictive value. We systematically reviewed all previous studies that have analyzed the expression of microRNAs as predictors of survival in pediatric AML and found 16 microRNAs and 4 microRNA signatures previously proposed as predictors of survival. We then used a public access cohort of 1414 pediatric AML patients from the TARGET project to develop a new predictive model using penalized lasso Cox regression based on microRNA expression. Here we propose a new score based on a 37-microRNA signature that is associated with AML and is able to predict survival more accurately than previous microRNA-based methods.
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Human embryonic stem cells (hESCs) differentiate into specialized cells, including midbrain dopaminergic neurons (DANs), and Non-human primates (NHPs) injected with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine develop some alterations observed in Parkinson's disease (PD) patients. Here, we obtained well-characterized DANs from hESCs and transplanted them into two parkinsonian monkeys to assess their behavioral and imaging changes. DANs from hESCs expressed dopaminergic markers, generated action potentials, and released dopamine (DA) in vitro. These neurons were transplanted bilaterally into the putamen of parkinsonian NHPs, and using magnetic resonance imaging techniques, we calculated the fractional anisotropy (FA) and mean diffusivity (MD), both employed for the first time for these purposes, to detect in vivo axonal and cellular density changes in the brain. Likewise, positron-emission tomography scans were performed to evaluate grafted DANs. Histological analyses identified grafted DANs, which were quantified stereologically. After grafting, animals showed signs of partially improved motor behavior in some of the HALLWAY motor tasks. Improvement in motor evaluations was inversely correlated with increases in bilateral FA. MD did not correlate with behavior but presented a negative correlation with FA. We also found higher 11C-DTBZ binding in positron-emission tomography scans associated with grafts. Higher DA levels measured by microdialysis after stimulation with a high-potassium solution or amphetamine were present in grafted animals after ten months, which has not been previously reported. Postmortem analysis of NHP brains showed that transplanted DANs survived in the putamen long-term, without developing tumors, in immunosuppressed animals. Although these results need to be confirmed with larger groups of NHPs, our molecular, behavioral, biochemical, and imaging findings support the integration and survival of human DANs in this pre-clinical PD model.
Assuntos
Células-Tronco Embrionárias Humanas , Doença de Parkinson , Animais , Humanos , Neurônios Dopaminérgicos/metabolismo , Células-Tronco Embrionárias Humanas/metabolismo , Haplorrinos/metabolismo , Mesencéfalo/metabolismo , Dopamina/metabolismo , Doença de Parkinson/terapia , Doença de Parkinson/metabolismoRESUMO
Bernard-Soulier syndrome (BSS) is a rare congenital disease characterized by macrothrombocytopenia and frequent bleeding. It is caused by pathogenic variants in three genes (GP1BA, GP1BB, or GP9) that encode for the GPIbα, GPIbß, and GPIX subunits of the GPIb-V-IX complex, the main platelet surface receptor for von Willebrand factor, being essential for platelet adhesion and aggregation. According to the affected gene, we distinguish BSS type A1 (GP1BA), type B (GP1BB), or type C (GP9). Pathogenic variants in these genes cause absent, incomplete, or dysfunctional GPIb-V-IX receptor and, consequently, a hemorrhagic phenotype. Using gene-editing tools, we generated knockout (KO) human cellular models that helped us to better understand GPIb-V-IX complex assembly. Furthermore, we developed novel lentiviral vectors capable of correcting GPIX expression, localization, and functionality in human GP9-KO megakaryoblastic cell lines. Generated GP9-KO induced pluripotent stem cells produced platelets that recapitulated the BSS phenotype: absence of GPIX on the membrane surface and large size. Importantly, gene therapy tools reverted both characteristics. Finally, hematopoietic stem cells from two unrelated BSS type C patients were transduced with the gene therapy vectors and differentiated to produce GPIX-expressing megakaryocytes and platelets with a reduced size. These results demonstrate the potential of lentiviral-based gene therapy to rescue BSS type C.
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Pediatric Acute Myeloid Leukemia (AML) is a rare and heterogeneous disease characterized by a high prevalence of gene fusions as driver mutations. Despite the improvement of survival in the last years, about 50% of patients still experience a relapse. It is not possible to improve prognosis only with further intensification of chemotherapy, as come with a severe cost to the health of patients, often resulting in treatment-related death or long-term sequels. To design more effective and less toxic therapies we need a better understanding of pediatric AML biology. The NUP98-KDM5A chimeric protein is exclusively found in a particular subgroup of young pediatric AML patients with complex karyotypes and poor prognosis. In this study, we investigated the impact of NUP98-KDM5A expression on cellular processes in human Pluripotent Stem Cell models and a patient-derived cell line. We found that NUP98-KDM5A generates genomic instability through two complementary mechanisms that involve accumulation of DNA damage and direct interference of RAE1 activity during mitosis. Overall, our data support that NUP98-KDM5A promotes genomic instability and likely contributes to malignant transformation.
Assuntos
Leucemia Mieloide Aguda , Proteínas de Fusão Oncogênica , Humanos , Criança , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Proteínas Oncogênicas/genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Instabilidade Genômica , Proteína 2 de Ligação ao Retinoblastoma/metabolismoRESUMO
Osteosarcomas are frequently associated to a poor prognosis and a modest response to current treatments. EC-8042 is a well-tolerated mithramycin analog that has demonstrated an efficient ability to eliminate tumor cells, including cancer stem cell subpopulations (CSC), in sarcomas. In transcriptomic and protein expression analyses, we identified NOTCH1 signaling as one of the main pro-stemness pathways repressed by EC-8042 in osteosarcomas. Overexpression of NOTCH-1 resulted in a reduced anti-tumor effect of EC-8042 in CSC-enriched 3D tumorspheres cultures. On the other hand, the depletion of the NOTCH-1 downstream target HES-1 was able to enhance the action of EC-8042 on CSCs. Moreover, HES1 depleted cells failed to recover after treatment withdrawal and showed reduced tumor growth potential in vivo. In contrast, mice xenografted with NOTCH1-overexpressing cells responded worse than parental cells to EC-8042. Finally, we found that active NOTCH1 levels in sarcoma patients was associated to advanced disease and lower survival. Overall, these data highlight the relevant role that NOTCH1 signaling plays in mediating stemness in osteosarcoma. Moreover, we demonstrate that EC-8042 is powerful inhibitor of NOTCH signaling and that the anti-CSC activity of this mithramycin analog highly rely on its ability to repress this pathway.
Assuntos
Neoplasias Ósseas , Osteossarcoma , Animais , Camundongos , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Células-Tronco Neoplásicas/metabolismo , Osteossarcoma/patologia , Plicamicina/farmacologia , Receptor Notch1/metabolismo , Receptores Notch/metabolismoRESUMO
Pediatric acute myeloid leukemia (AML) is a rare and heterogeneous disease that remains the major cause of mortality in children with leukemia. To improve the outcome of pediatric AML we need to gain knowledge on the biological bases of this disease. NUP98-KDM5A (NK5A) fusion protein is present in a particular subgroup of young pediatric patients with poor outcome. We report the generation and characterization of human Embryonic Stem Cell (hESC) clonal lines with inducible expression of NK5A. Temporal control of NK5A expression during hematopoietic differentiation from hESC will be critical for elucidating its participation during the leukemogenic process.
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In 1975 two independent groups noticed the presence of immune cells with a unique ability to recognize and eliminate transformed hematopoietic cells without any prior sensitization or expansion of specific clones. Since then, NK cells have been the axis of thousands of studies that have resulted until June 2021, in more than 70 000 publications indexed in PubMed. As result of this work, which include approaches in vitro, in vivo, and in natura, it has been possible to appreciate the role played by the NK cells, not only as effectors against specific pathogens, but also as regulators of the immune response. Recent advances have revealed previous unidentified attributes of NK cells including the ability to adapt to new conditions under the context of chronic infections, or their ability to develop some memory-like characteristics. In this review, we will discuss significant findings that have rule our understanding of the NK cell biology, the developing of these findings into new concepts in immunology, and how these conceptual platforms are being used in the design of strategies for cancer immunotherapy.
Assuntos
Neoplasias Hematológicas , Neoplasias , Neoplasias Hematológicas/terapia , Humanos , Imunoterapia/métodos , Células Matadoras Naturais , Neoplasias/terapiaRESUMO
Acute myeloid leukemia (AML) is the most common acute leukemia in adults. Disease heterogeneity is well documented, and patient stratification determines treatment decisions. Patient-derived xenografts (PDXs) from risk-stratified AML are crucial for studying AML biology and testing novel therapeutics. Despite recent advances in PDX modeling of AML, reproducible engraftment of human AML is primarily limited to high-risk (HR) cases, with inconsistent or very protracted engraftment observed for favorable-risk (FR) and intermediate-risk (IR) patients. We used NSGS mice to characterize the engraftment robustness/kinetics of 28 AML patient samples grouped according to molecular/cytogenetic classification and assessed whether the orthotopic coadministration of patient-matched bone marrow mesenchymal stromal cells (BM MSCs) improves AML engraftment. PDX event-free survival correlated well with the predictable prognosis of risk-stratified AML patients. The majority (85-94%) of the mice were engrafted in bone marrow (BM) independently of the risk group, although HR AML patients showed engraftment levels that were significantly superior to those of FR or IR AML patients. Importantly, the engraftment levels observed in NSGS mice by week 6 remained stable over time. Serial transplantation and long-term culture-initiating cell (LTC-IC) assays revealed long-term engraftment limited to HR AML patients, fitter leukemia-initiating cells (LICs) in HR AML samples, and the presence of AML LICs in the CD34- leukemic fraction, regardless of the risk group. Finally, orthotopic coadministration of patient-matched BM MSCs and AML cells was dispensable for BM engraftment levels but favored peripheralization of engrafted AML cells. This comprehensive characterization of human AML engraftment in NSGS mice offers a valuable platform for in vivo testing of targeted therapies in risk-stratified AML patient samples.
Assuntos
Leucemia Mieloide Aguda , Animais , Antígenos CD34 , Medula Óssea , Humanos , Leucemia Mieloide Aguda/terapia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCIDRESUMO
Mammals optimize their physiology to the light-dark cycle by synchronization of the master circadian clock in the brain with peripheral clocks in the rest of the tissues of the body. Circadian oscillations rely on a negative feedback loop exerted by the molecular clock that is composed by transcriptional activators Bmal1 and Clock, and their negative regulators Period and Cryptochrome. Components of the molecular clock are expressed during early development, but onset of robust circadian oscillations is only detected later during embryogenesis. Here, we have used naïve pluripotent mouse embryonic stem cells (mESCs) to study the role of Bmal1 during early development. We found that, compared to wild-type cells, Bmal1-/- mESCs express higher levels of Nanog protein and altered expression of pluripotency-associated signalling pathways. Importantly, Bmal1-/- mESCs display deficient multi-lineage cell differentiation capacity during the formation of teratomas and gastrula-like organoids. Overall, we reveal that Bmal1 regulates pluripotent cell differentiation and propose that the molecular clock is an hitherto unrecognized regulator of mammalian development.
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Fatores de Transcrição ARNTL/metabolismo , Diferenciação Celular/fisiologia , Células-Tronco Embrionárias Murinas/metabolismo , Fatores de Transcrição ARNTL/fisiologia , Animais , Proteínas CLOCK/genética , Proteínas CLOCK/metabolismo , Relógios Circadianos/fisiologia , Ritmo Circadiano/genética , Retroalimentação Fisiológica/fisiologia , Expressão Gênica/genética , Células-Tronco Pluripotentes Induzidas/citologia , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Proteínas Circadianas Period/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Transcrição GênicaRESUMO
Multipotent mesenchymal stromal cells (MSCs) have emerged as a promising cell therapy in regenerative medicine and for autoimmune/inflammatory diseases. However, a main hurdle for MSCs-based therapies is the loss of their proliferative potential in vitro. Here we report that glycoprotein A repetitions predominant (GARP) is required for the proliferation and survival of adipose-derived MSCs (ASCs) via its regulation of transforming growth factor-ß (TGF-ß) activation. Silencing of GARP in human ASCs increased their activation of TGF-ß which augmented the levels of mitochondrial reactive oxygen species (mtROS), resulting in DNA damage, a block in proliferation and apoptosis. Inhibition of TGF-ß signaling reduced the levels of mtROS and DNA damage and restored the ability of GARP-/low ASCs to proliferate. In contrast, overexpression of GARP in ASCs increased their proliferative capacity and rendered them more resistant to etoposide-induced DNA damage and apoptosis, in a TGF-ß-dependent manner. In summary, our data show that the presence or absence of GARP on ASCs gives rise to distinct TGF-ß responses with diametrically opposing effects on ASC proliferation and survival.
Assuntos
Células-Tronco Mesenquimais/metabolismo , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , HumanosRESUMO
Familial Platelet Disorder with associated Myeloid Malignancy (FPDMM) is a rare platelet disorder caused by mutations in RUNX1. We generated an iPSC line (GENYOi005-A) from a FPDMM patient with a non-previously reported variant p.Thr196Ala. Non-integrative Sendai viruses expressing the Yamanaka reprogramming factors were used to reprogram peripheral blood mononuclear cells from this FPDMM patient. Characterization of GENYOi005-A included genetic analysis of RUNX1 locus, Short Tandem Repeats profiling, alkaline phosphatase enzymatic activity, expression of pluripotency-associated factors and differentiation studies in vitro and in vivo. This iPSC line will provide a powerful tool to study developmental alterations of FPDMM patients.
Assuntos
Transtornos Herdados da Coagulação Sanguínea/genética , Transtornos Herdados da Coagulação Sanguínea/patologia , Transtornos Plaquetários/genética , Transtornos Plaquetários/patologia , Diferenciação Celular , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Células-Tronco Pluripotentes Induzidas/patologia , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Leucócitos Mononucleares/patologia , Mutação , Células Cultivadas , Reprogramação Celular , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Leucócitos Mononucleares/metabolismo , Pessoa de Meia-IdadeRESUMO
ADNP syndrome is an intellectual disability associated with Autism spectrum disorder caused by mutations in ADNP. We generated an iPSC line from an ADNP syndrome pediatric patient harboring the mutation p.Trp719* (GENYOi004-A). Peripheral blood mononuclear cells were reprogrammed using a non-transmissible form of Sendai viruses expressing the four Yamanaka factors (Oct3/4, SOX2, KLF4 and c-MYC). Characterization of GENYOi004-A included mutation analysis of ADNP by allele-specific PCR, genetic identity by Short Tandem Repeats polymorphism profiling, alkaline phosphatase enzymatic activity, expression of pluripotency-associated factors and pluripotency studies in vivo. GENYOi004-A will be useful to evaluate ADNP syndrome alterations at early developmental stages.
Assuntos
Transtorno do Espectro Autista/genética , Diferenciação Celular , Proteínas de Homeodomínio/genética , Células-Tronco Pluripotentes Induzidas/patologia , Leucócitos Mononucleares/patologia , Mutação , Proteínas do Tecido Nervoso/genética , Teratoma/etiologia , Animais , Transtorno do Espectro Autista/patologia , Células Cultivadas , Reprogramação Celular , Criança , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Fator 4 Semelhante a Kruppel , Leucócitos Mononucleares/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Fenótipo , Teratoma/patologiaRESUMO
The Notch ligand DLL4 has key roles during embryonic development of different tissues, but most of the data comes from animal models. Here we describe the generation and characterization of 2 human Pluripotent Stem Cell (hPSC) lines that overexpress DLL4, as well as the two corresponding control hPSC lines. DLL4 expression can be detected at the mRNA and protein level, and does not affect the pluripotency of the cells. These hPSC lines can be used to study the role of DLL4 during human embryonic development.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Células-Tronco Pluripotentes/citologia , Proteínas Adaptadoras de Transdução de Sinal , Adulto , Proteínas de Ligação ao Cálcio , Diferenciação Celular , Linhagem Celular , Proliferação de Células , Células Cultivadas , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Células-Tronco Pluripotentes/metabolismo , Receptores Notch/genética , Receptores Notch/metabolismoRESUMO
HoxA9 is an evolutionarily conserved homeobox gene implicated in embryo development. To study the roles of Hoxa9 during human development we generated a transgenic H9 (hESC) line that overexpresses HoxA9 and the Enhanced Green Fluorescent Protein (EGFP), and a control H9 with a stable expression of the EGFP. The resulting H9-HoxA9-EGFP and H9-EGFP cell lines allow an efficient visualization of hESCs by fluorescent microscopy, quantification by flow cytometry and cell differentiation tracking. Both transgenic cell lines maintained the pluripotent phenotype, the ability to differentiate into all three germ layers and a normal karyotype.
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Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário , Células-Tronco Embrionárias/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Homeodomínio/metabolismo , Diferenciação Celular , Células Cultivadas , Células-Tronco Embrionárias/citologia , Feminino , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Homeodomínio/genética , Humanos , TransfecçãoRESUMO
Runt-related transcription factor 1 (Runx1) is a master hematopoietic transcription factor essential for hematopoietic stem cell (HSC) emergence. Runx1-deficient mice die during early embryogenesis due to the inability to establish definitive hematopoiesis. Here, we have used human pluripotent stem cells (hPSCs) as model to study the role of RUNX1 in human embryonic hematopoiesis. Although the three RUNX1 isoforms a, b, and c were induced in CD45+ hematopoietic cells, RUNX1c was the only isoform induced in hematoendothelial progenitors (HEPs)/hemogenic endothelium. Constitutive expression of RUNX1c in human embryonic stem cells enhanced the appearance of HEPs, including hemogenic (CD43+) HEPs and promoted subsequent differentiation into blood cells. Conversely, specific deletion of RUNX1c dramatically reduced the generation of hematopoietic cells from HEPs, indicating that RUNX1c is a master regulator of human hematopoietic development. Gene expression profiling of HEPs revealed a RUNX1c-induced proinflammatory molecular signature, supporting previous studies demonstrating proinflammatory signaling as a regulator of HSC emergence. Collectively, RUNX1c orchestrates hematopoietic specification of hPSCs, possibly in cooperation with proinflammatory signaling. Stem Cells 2017;35:2253-2266.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/genética , Perfilação da Expressão Gênica/métodos , Células-Tronco Pluripotentes/metabolismo , Animais , Diferenciação Celular , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Humanos , Camundongos , Transdução de SinaisRESUMO
Bernard Soulier Syndrome (BSS) is a rare autosomal platelet disorder characterized by mutations in the von Willebrand factor platelet receptor complex GPIb-V-IX. In this work we have generated an induced pluripotent stem cell (BSS3-PBMC-iPS4F8) from peripheral blood mononuclear cells of a BSS patient with a p.Phe55Ser mutation in the GPIX gene. Characterization of BSS3-PBMC-iPS4F8 showed that these cells maintained the original mutation present in the BSS patient, expressed pluripotent stem cell markers and were able to differentiate into the three germline layers. This new iPSC line will contribute to better understand the biology of BSS disease.
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Síndrome de Bernard-Soulier/patologia , Células-Tronco Pluripotentes Induzidas/citologia , Complexo Glicoproteico GPIb-IX de Plaquetas/genética , Sequência de Bases , Síndrome de Bernard-Soulier/genética , Síndrome de Bernard-Soulier/metabolismo , Diferenciação Celular , Linhagem Celular , Reprogramação Celular , Análise Mutacional de DNA , Corpos Embrioides/metabolismo , Corpos Embrioides/patologia , Feminino , Homozigoto , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Cariótipo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Polimorfismo de Nucleotídeo Único , Sequências de Repetição em Tandem/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Pediatric Acute Megakaryoblastic Leukemia not associated to Down Syndrome (non-DS AMKL) is a rare disease with a dismal prognosis. Around 15% of patients carry the chromosomal translocation t(1;22) that originates the fusion oncogene RBM15-MKL1, which is linked to an earlier disease onset (median of 6months of age) and arises in utero. Here we report the generation of two hPSC cell lines constitutively expressing the oncogene RBM15-MKL1, resulting in an increased expression of known RBM15-MKL1 gene targets. These cell lines represent new disease models of pediatric AMKL to study the impact of the RBM15-MKL1 oncogene on human embryonic hematopoietic development.
Assuntos
Leucemia Megacarioblástica Aguda/patologia , Proteínas de Fusão Oncogênica/metabolismo , Células-Tronco Pluripotentes/citologia , Biomarcadores/metabolismo , Linhagem Celular , Criança , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Humanos , Leucemia Megacarioblástica Aguda/metabolismo , Modelos Biológicos , Proteínas de Fusão Oncogênica/genética , Células-Tronco Pluripotentes/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Bernard Soulier Syndrome (BSS) is an inherited rare platelet disorder characterized by mutations in the platelet glycoprotein complex GPIb-IX-V. We generated an induced pluripotent stem cell (iPSC) line from a BSS patient with a mutation p.Asn45Ser in the GPIX locus (BSS2-PBMC-iPS4F24). Peripheral blood mononuclear cells were reprogrammed using non-integrative viral transduction. Characterization of BSS2-PBMC-iPS4F24 included mutational analysis of GPIX locus, analysis of conventional pluripotency-associated factors at mRNA and protein level and in vitro and in vivo differentiation studies. This iPSC line will provide a powerful tool to study the biology of BSS disease.
Assuntos
Síndrome de Bernard-Soulier/patologia , Células-Tronco Pluripotentes Induzidas/citologia , Complexo Glicoproteico GPIb-IX de Plaquetas/genética , Animais , Sequência de Bases , Síndrome de Bernard-Soulier/genética , Diferenciação Celular , Linhagem Celular , Reprogramação Celular , Análise Mutacional de DNA , Corpos Embrioides/citologia , Corpos Embrioides/metabolismo , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/transplante , Cariótipo , Leucócitos Mononucleares/citologia , Camundongos , Camundongos SCID , Polimorfismo de Nucleotídeo Único , Teratoma/patologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Induced pluripotent stem cells (iPSCs) are a powerful tool for disease modeling. They are routinely generated from healthy donors and patients from multiple cell types at different developmental stages. However, reprogramming leukemias is an extremely inefficient process. Few studies generated iPSCs from primary chronic myeloid leukemias, but iPSC generation from acute myeloid or lymphoid leukemias (ALL) has not been achieved. We attempted to generate iPSCs from different subtypes of B-ALL to address the developmental impact of leukemic fusion genes. OKSM(L)-expressing mono/polycistronic-, retroviral/lentiviral/episomal-, and Sendai virus vector-based reprogramming strategies failed to render iPSCs in vitro and in vivo. Addition of transcriptomic-epigenetic reprogramming "boosters" also failed to generate iPSCs from B cell blasts and B-ALL lines, and when iPSCs emerged they lacked leukemic fusion genes, demonstrating non-leukemic myeloid origin. Conversely, MLL-AF4-overexpressing hematopoietic stem cells/B progenitors were successfully reprogrammed, indicating that B cell origin and leukemic fusion gene were not reprogramming barriers. Global transcriptome/DNA methylome profiling suggested a developmental/differentiation refractoriness of MLL-rearranged B-ALL to reprogramming into pluripotency.