ABSTRACT
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.
Subject(s)
Leukemia, Myeloid, Acute , Oncogene Proteins, Fusion , Humans , Child , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Nuclear Pore Complex Proteins/genetics , Nuclear Pore Complex Proteins/metabolism , Oncogene Proteins/genetics , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/pathology , Genomic Instability , Retinoblastoma-Binding Protein 2/metabolismABSTRACT
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.
ABSTRACT
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.
Subject(s)
Intercellular Signaling Peptides and Proteins/metabolism , Pluripotent Stem Cells/cytology , Adaptor Proteins, Signal Transducing , Adult , Calcium-Binding Proteins , Cell Differentiation , Cell Line , Cell Proliferation , Cells, Cultured , Female , Humans , Intercellular Signaling Peptides and Proteins/genetics , Pluripotent Stem Cells/metabolism , Receptors, Notch/genetics , Receptors, Notch/metabolismABSTRACT
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.
Subject(s)
Embryo, Mammalian/metabolism , Embryonic Development , Embryonic Stem Cells/metabolism , Gene Expression Regulation, Developmental , Green Fluorescent Proteins/metabolism , Homeodomain Proteins/metabolism , Cell Differentiation , Cells, Cultured , Embryonic Stem Cells/cytology , Female , Genes, Reporter , Green Fluorescent Proteins/genetics , Homeodomain Proteins/genetics , Humans , TransfectionABSTRACT
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.
