RESUMEN
Severe congenital neutropenia (CN) is a preleukemic bone marrow failure syndrome with a 20% risk of evolving into leukemia or myelodysplastic syndrome (MDS). Patterns of acquisition of leukemia-associated mutations were investigated using next-generation deep-sequencing in 31 CN patients who developed leukemia or MDS. Twenty (64.5%) of the 31 patients had mutations in RUNX1. A majority of patients with RUNX1 mutations (80.5%) also had acquired CSF3R mutations. In contrast to their high frequency in CN patients who developed leukemia or MDS, RUNX1 mutations were found in only 9 of 307 (2.9%) patients with de novo pediatric acute myeloid leukemia. A sequential analysis at stages prior to overt leukemia revealed RUNX1 mutations to be late events in leukemic transformation. Single-cell analyses in 2 patients showed that RUNX1 and CSF3R mutations were present in the same malignant clone. Functional studies demonstrated elevated granulocyte colony-stimulating factor (G-CSF)-induced proliferation with diminished myeloid differentiation of hematopoietic CD34(+) cells coexpressing mutated forms of RUNX1 and CSF3R. The high frequency of cooperating RUNX1 and CSF3R mutations in CN patients suggests a novel molecular pathway of leukemogenesis: mutations in the hematopoietic cytokine receptor (G-CSFR) in combination with the second mutations in the downstream hematopoietic transcription fator (RUNX1). The detection of both RUNX1 and CSF3R mutations could be used as a marker for identifying CN patients with a high risk of progressing to leukemia or MDS.
Asunto(s)
Transformación Celular Neoplásica/genética , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Leucemia Mieloide/genética , Mutación , Neutropenia/congénito , Receptores del Factor Estimulante de Colonias/genética , Adolescente , Adulto , Niño , Preescolar , Síndromes Congénitos de Insuficiencia de la Médula Ósea , Análisis Citogenético , Femenino , Humanos , Masculino , Neutropenia/genética , Neutropenia/patología , Transducción de Señal/genética , Adulto JovenRESUMEN
Shwachman-Diamond syndrome is a congenital bone marrow failure disorder characterized by debilitating neutropenia. The disease is associated with loss-of-function mutations in the SBDS gene, implicated in ribosome biogenesis, but the cellular and molecular events driving cell specific phenotypes in ribosomopathies remain poorly defined. Here, we established what is to our knowledge the first mammalian model of neutropenia in Shwachman-Diamond syndrome through targeted downregulation of Sbds in hematopoietic stem and progenitor cells expressing the myeloid transcription factor CCAAT/enhancer binding protein α (Cebpa). Sbds deficiency in the myeloid lineage specifically affected myelocytes and their downstream progeny while, unexpectedly, it was well tolerated by rapidly cycling hematopoietic progenitor cells. Molecular insights provided by massive parallel sequencing supported cellular observations of impaired cell cycle exit and formation of secondary granules associated with the defect of myeloid lineage progression in myelocytes. Mechanistically, Sbds deficiency activated the p53 tumor suppressor pathway and induced apoptosis in these cells. Collectively, the data reveal a previously unanticipated, selective dependency of myelocytes and downstream progeny, but not rapidly cycling progenitors, on this ubiquitous ribosome biogenesis protein, thus providing a cellular basis for the understanding of myeloid lineage biased defects in Shwachman-Diamond syndrome.
Asunto(s)
Diferenciación Celular/genética , Linaje de la Célula/genética , Células Madre Hematopoyéticas/metabolismo , Células Mieloides/citología , Células Mieloides/metabolismo , Neutropenia/genética , Proteínas/genética , Animales , Apoptosis/genética , Enfermedades de la Médula Ósea/genética , Proteínas Potenciadoras de Unión a CCAAT/genética , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Ciclo Celular/genética , Modelos Animales de Enfermedad , Insuficiencia Pancreática Exocrina/genética , Eliminación de Gen , Hematopoyesis/genética , Trasplante de Células Madre Hematopoyéticas , Células Madre Hematopoyéticas/citología , Lipomatosis/genética , Ratones , Ratones Noqueados , Síndrome de Shwachman-Diamond , Transducción de Señal , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Severe congenital neutropenia (SCN) is a BM failure syndrome with a high risk of progression to acute myeloid leukemia (AML). The underlying genetic changes involved in SCN evolution to AML are largely unknown. We obtained serial hematopoietic samples from an SCN patient who developed AML 17 years after the initiation of G-CSF treatment. Next- generation sequencing was performed to identify mutations during disease progression. In the AML phase, we found 12 acquired nonsynonymous mutations. Three of these, in CSF3R, LLGL2, and ZC3H18, co-occurred in a subpopulation of progenitor cells already in the early SCN phase. This population expanded over time, whereas clones harboring only CSF3R mutations disappeared from the BM. The other 9 mutations were only apparent in the AML cells and affected known AML-associated genes (RUNX1 and ASXL1) and chromatin remodelers (SUZ12 and EP300). In addition, a novel CSF3R mutation that conferred autonomous proliferation to myeloid progenitors was found. We conclude that progression from SCN to AML is a multistep process, with distinct mutations arising early during the SCN phase and others later in AML development. The sequential gain of 2 CSF3R mutations implicates abnormal G-CSF signaling as a driver of leukemic transformation in this case of SCN.