RESUMEN
Diamond Blackfan Anemia (DBA) is a rare macrocytic red blood cell aplasia that usually presents within the first year of life. The vast majority of patients carry a mutation in one of approximately 20 genes that results in ribosomal insufficiency with the most significant clinical manifestations being anemia and a predisposition to cancers. Nemo-like Kinase (NLK) is hyperactivated in the erythroid progenitors of DBA patients and inhibition of this kinase improves erythropoiesis, but how NLK contributes to the pathogenesis of the disease is unknown. Here we report that activated NLK suppresses the critical upregulation of mitochondrial biogenesis required in early erythropoiesis. During normal erythropoiesis, mTORC1 facilitates the translational upregulation of Transcription factor A, mitochondrial (TFAM), and Prohibin 2 (PHB2) to increase mitochondrial biogenesis. In our models of DBA, active NLK phosphorylates the regulatory component of mTORC1, thereby suppressing mTORC1 activity and preventing mTORC1-mediated TFAM and PHB2 upregulation and subsequent mitochondrial biogenesis. Improvement of erythropoiesis that accompanies NLK inhibition is negated when TFAM and PHB2 upregulation is prevented. These data demonstrate that a significant contribution of NLK on the pathogenesis of DBA is through loss of mitochondrial biogenesis.
RESUMEN
Pyruvate kinase (PK) deficiency, a rare, congenital haemolytic anaemia caused by mutations in the PKLR gene, is associated with many clinical manifestations, but the full disease burden has yet to be characterised. The Peak Registry (NCT03481738) is an observational, longitudinal registry of adult and paediatric patients with PK deficiency. Here, we described comorbidities and complications in these patients by age at most recent visit and PKLR genotype. As of 13 May 2022, 241 patients were included in the analysis. In total, 48.3% had undergone splenectomy and 50.5% had received chelation therapy. History of iron overload (before enrolment/during follow-up) was common (52.5%), even in never-transfused patients (20.7%). Neonatal complications and symptoms included jaundice, splenomegaly and hepatomegaly, with treatment interventions required in 41.5%. Among adults, osteopenia/osteoporosis occurred in 19.0% and pulmonary hypertension in 6.7%, with median onset ages of 37, 33 and 22 years, respectively. Biliary events and bone health problems were common across PKLR genotypes. Among 11 patients who had thromboembolic events, eight had undergone prior splenectomy. Patients with PK deficiency may have many complications, which can occur early in and throughout life. Awareness of their high disease burden may help clinicians better provide appropriate monitoring and management of these patients.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica , Piruvato Quinasa , Errores Innatos del Metabolismo del Piruvato , Sistema de Registros , Humanos , Piruvato Quinasa/deficiencia , Piruvato Quinasa/genética , Masculino , Femenino , Adulto , Niño , Anemia Hemolítica Congénita no Esferocítica/genética , Anemia Hemolítica Congénita no Esferocítica/epidemiología , Errores Innatos del Metabolismo del Piruvato/genética , Errores Innatos del Metabolismo del Piruvato/epidemiología , Adolescente , Preescolar , Lactante , Comorbilidad , Persona de Mediana Edad , Esplenectomía , Adulto Joven , Hipertensión Pulmonar/etiología , Hipertensión Pulmonar/genética , Hipertensión Pulmonar/epidemiología , Sobrecarga de Hierro/etiología , Sobrecarga de Hierro/epidemiología , Enfermedades Óseas Metabólicas/etiología , Enfermedades Óseas Metabólicas/epidemiología , Recién NacidoRESUMEN
Master regulators, such as the hematopoietic transcription factor (TF) GATA1, play an essential role in orchestrating lineage commitment and differentiation. However, the precise mechanisms by which such TFs regulate transcription through interactions with specific cis-regulatory elements remain incompletely understood. Here, we describe a form of congenital hemolytic anemia caused by missense mutations in an intrinsically disordered region of GATA1, with a poorly understood role in transcriptional regulation. Through integrative functional approaches, we demonstrate that these mutations perturb GATA1 transcriptional activity by partially impairing nuclear localization and selectively altering precise chromatin occupancy by GATA1. These alterations in chromatin occupancy and concordant chromatin accessibility changes alter faithful gene expression, with failure to both effectively silence and activate select genes necessary for effective terminal red cell production. We demonstrate how disease-causing mutations can reveal regulatory mechanisms that enable the faithful genomic targeting of master TFs during cellular differentiation.
Asunto(s)
Anemia , Factor de Transcripción GATA1 , Diferenciación Celular/genética , Cromatina/genética , Inmunoprecipitación de Cromatina , Eritropoyesis/genética , Factor de Transcripción GATA1/genética , Factor de Transcripción GATA1/metabolismo , HumanosRESUMEN
BACKGROUND: Identification of hemoglobin (Hb) variants is of significant value in the clinical diagnosis of hemoglobinopathy. However, conventional methods for identification of Hb variants in clinical laboratories can be inadequate due to the lack of structural characterization. We describe the use of neutral-coating capillary electrophoresis coupled with high-resolution mass spectrometry (CE-HR-MS) to achieve high-performance top-down identification of Hb variants. METHODS: An Orbitrap Q-Exactive Plus mass spectrometer was coupled with an ECE-001 capillary electrophoresis (CE) unit through an EMASS-II ion source. A PS1 neutral-coating capillary was used for CE. Samples of red blood cells were lysed in water and diluted in 10 mM ammonium formate buffer for analysis. Deconvolution of raw mass spectrometry data was carried out to merge multiple charge states and isotopic peaks of an analyte to obtain its monoisotopic mass. RESULTS: The neutral-coating CE could baseline separate individual Hb subunits dissociated from intact Hb forms, and the HR-MS could achieve both intact-protein analysis and top-down analysis of analytes. A number of patient samples that contain Hb subunit variants were analyzed, and the variants were successfully identified using the CE-HR-MS method. CONCLUSIONS: The CE-HR-MS method has been demonstrated as a useful tool for top-down identification of Hb variants. With the ability to characterize the primary structures of Hb subunits, the CE-HR-MS method has significant advantages to complement or partially replace the conventional methods for the identification of Hb variants.
Asunto(s)
Electroforesis Capilar , Hemoglobinopatías , Humanos , Espectrometría de Masas/métodos , Electroforesis Capilar/métodos , Eritrocitos , Hemoglobinas/genéticaRESUMEN
Nemo-like kinase (NLK) is a member of the mitogen-activated protein kinase family of kinases and shares a highly conserved kinase domain with other mitogen-activated protein kinase family members. The activation of NLK contributes to the pathogenesis of Diamond-Blackfan anemia (DBA), reducing c-myb expression and mechanistic target of rapamycin activity, and is therefore a potential therapeutic target. Unlike other anemias, the hematopoietic effects of DBA are largely restricted to the erythroid lineage. Mutations in ribosomal genes induce ribosomal insufficiency and reduced protein translation, dramatically impacting early erythropoiesis in the bone marrow of patients with DBA. We sought to identify compounds that suppress NLK and increases erythropoiesis in ribosomal insufficiency. We report that the active component of ginseng, ginsenoside Rb1, suppresses NLK expression and improves erythropoiesis in in vitro models of DBA. Ginsenoside Rb1-mediated suppression of NLK occurs through the upregulation of miR-208, which binds to the 3'-UTR of NLK mRNA and targets it for degradation. We also compare ginsenoside Rb1-mediated upregulation of miR-208 with metformin-mediated upregulation of miR-26. We conclude that targeting NLK expression through miRNA binding of the unique 3'-UTR is a viable alternative to the challenges of developing small-molecule inhibitors to target the highly conserved kinase domain of this specific kinase.
Asunto(s)
Anemia de Diamond-Blackfan/patología , Eritropoyesis/efectos de los fármacos , Ginsenósidos/farmacología , Panax/química , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Regiones no Traducidas 3' , Animales , HumanosRESUMEN
BACKGROUND: Pyruvate kinase deficiency is caused by mutations in PKLR and leads to congenital hemolytic anemia. Mitapivat is an oral, small-molecule allosteric activator of pyruvate kinase in red cells. METHODS: In this uncontrolled, phase 2 study, we evaluated the safety and efficacy of mitapivat in 52 adults with pyruvate kinase deficiency who were not receiving red-cell transfusions. The patients were randomly assigned to receive either 50 mg or 300 mg of mitapivat twice daily for a 24-week core period; eligible patients could continue treatment in an ongoing extension phase. RESULTS: Common adverse events, including headache and insomnia, occurred at the time of drug initiation and were transient; 92% of the episodes of headache and 47% of the episodes of insomnia resolved within 7 days. The most common serious adverse events, hemolytic anemia and pharyngitis, each occurred in 2 patients (4%). A total of 26 patients (50%) had an increase of more than 1.0 g per deciliter in the hemoglobin level. Among these patients, the mean maximum increase was 3.4 g per deciliter (range, 1.1 to 5.8), and the median time until the first increase of more than 1.0 g per deciliter was 10 days (range, 7 to 187); 20 patients (77%) had an increase of more than 1.0 g per deciliter in the hemoglobin level at more than 50% of visits during the core study period, with improvement in markers of hemolysis. The response was sustained in all 19 patients remaining in the extension phase, with a median follow-up of 29 months (range, 22 to 35). Hemoglobin responses were observed only in patients who had at least one missense PKLR mutation and were associated with the red-cell pyruvate kinase protein level at baseline. CONCLUSIONS: The administration of mitapivat was associated with a rapid increase in the hemoglobin level in 50% of adults with pyruvate kinase deficiency, with a sustained response during a median follow-up of 29 months during the extension phase. Adverse effects were mainly low-grade and transient. (Funded by Agios Pharmaceuticals; ClinicalTrials.gov number, NCT02476916.).
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica/tratamiento farmacológico , Hemoglobinas/metabolismo , Piperazinas/administración & dosificación , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato/tratamiento farmacológico , Quinolinas/administración & dosificación , Administración Oral , Adolescente , Adulto , Anemia Hemolítica Congénita no Esferocítica/sangre , Anemia Hemolítica Congénita no Esferocítica/genética , Catecoles , Esquema de Medicación , Femenino , Estudios de Seguimiento , Cefalea/inducido químicamente , Humanos , Masculino , Mutación , Piperazinas/efectos adversos , Piruvato Quinasa/sangre , Piruvato Quinasa/genética , Errores Innatos del Metabolismo del Piruvato/sangre , Errores Innatos del Metabolismo del Piruvato/genética , Quinolinas/efectos adversos , Trastornos del Inicio y del Mantenimiento del Sueño/inducido químicamente , Tirfostinos , Adulto JovenRESUMEN
The congenital sideroblastic anemias (CSAs) are a heterogeneous group of inherited disorders of erythropoiesis characterized by pathologic deposits of iron in the mitochondria of developing erythroblasts. Mutations in the mitochondrial glycine carrier SLC25A38 cause the most common recessive form of CSA. Nonetheless, the disease is still rare, there being fewer than 70 reported families. Here we describe the clinical phenotype and genotypes of 31 individuals from 24 families, including 11 novel mutations. We also review the spectrum of reported mutations and genotypes associated with the disease, describe the unique localization of missense mutations in transmembrane domains and account for the presence of several alleles in different populations.
Asunto(s)
Anemia Sideroblástica/congénito , Genotipo , Proteínas de Transporte de Membrana Mitocondrial/genética , Mutación , Fenotipo , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , MasculinoRESUMEN
Diamond-Blackfan anemia (DBA) is a rare bone marrow failure disorder that affects 7 out of 1,000,000 live births and has been associated with mutations in components of the ribosome. In order to characterize the genetic landscape of this heterogeneous disorder, we recruited a cohort of 472 individuals with a clinical diagnosis of DBA and performed whole-exome sequencing (WES). We identified relevant rare and predicted damaging mutations for 78% of individuals. The majority of mutations were singletons, absent from population databases, predicted to cause loss of function, and located in 1 of 19 previously reported ribosomal protein (RP)-encoding genes. Using exon coverage estimates, we identified and validated 31 deletions in RP genes. We also observed an enrichment for extended splice site mutations and validated their diverse effects using RNA sequencing in cell lines obtained from individuals with DBA. Leveraging the size of our cohort, we observed robust genotype-phenotype associations with congenital abnormalities and treatment outcomes. We further identified rare mutations in seven previously unreported RP genes that may cause DBA, as well as several distinct disorders that appear to phenocopy DBA, including nine individuals with biallelic CECR1 mutations that result in deficiency of ADA2. However, no new genes were identified at exome-wide significance, suggesting that there are no unidentified genes containing mutations readily identified by WES that explain >5% of DBA-affected case subjects. Overall, this report should inform not only clinical practice for DBA-affected individuals, but also the design and analysis of rare variant studies for heterogeneous Mendelian disorders.
Asunto(s)
Anemia de Diamond-Blackfan/genética , Adolescente , Niño , Preescolar , Estudios de Cohortes , Exoma/genética , Exones/genética , Femenino , Eliminación de Gen , Estudios de Asociación Genética/métodos , Humanos , Péptidos y Proteínas de Señalización Intercelular/genética , Masculino , Mutación/genética , Fenotipo , Proteínas Ribosómicas/genética , Ribosomas/genética , Análisis de Secuencia de ARN/métodos , Secuenciación del Exoma/métodosRESUMEN
Diagnosis of pyruvate kinase deficiency (PKD), the most common cause of hereditary non-spherocytic haemolytic anaemia, remains challenging in routine practice and no biomarkers for clinical severity have been characterised. This prospective study enrolled 41 patients with molecularly confirmed PKD from nine North American centres to evaluate the diagnostic sensitivity of pyruvate kinase (PK) enzyme activity and PK:hexokinase (HK) enzyme activity ratio, and evaluate the erythrocyte PK (PK-R) protein level and erythrocyte metabolites as biomarkers for clinical severity. In this population not transfused for ≥90 days before sampling, the diagnostic sensitivity of the PK enzyme assay was 90% [95% confidence interval (CI) 77-97%], whereas the PK:HK ratio sensitivity was 98% (95% CI 87-100%). There was no correlation between PK enzyme activity and clinical severity. Transfusion requirements correlated with normalised erythrocyte ATP levels (r = 0·527, P = 0·0016) and PK-R protein levels (r = -0·527, P = 0·0028). PK-R protein levels were significantly higher in the never transfused [median (range) 40·1 (9·8-73·9)%] versus ever transfused [median (range) 7·7 (0·4-15·1)%] patients (P = 0·0014). The PK:HK ratio had excellent sensitivity for PK diagnosis, superior to PKLR exon sequencing. Given that the number of PKLR variants and genotype combinations limits prognostication based on molecular findings, PK-R protein level may be a useful prognostic biomarker of disease severity and merits further study.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica/sangre , Eritrocitos/enzimología , Hexoquinasa/sangre , Piruvato Quinasa/sangre , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato/sangre , Adolescente , Adulto , Anemia Hemolítica Congénita no Esferocítica/genética , Biomarcadores/sangre , Niño , Preescolar , Femenino , Hexoquinasa/genética , Humanos , Lactante , Masculino , Persona de Mediana Edad , Piruvato Quinasa/genética , Errores Innatos del Metabolismo del Piruvato/genética , Índice de Severidad de la EnfermedadRESUMEN
OBJECTIVES: Pyruvate kinase (PK) deficiency is caused by PKLR gene mutations, leading to defective red blood cell glycolysis and hemolytic anemia. Rates of comorbidities and complications by transfusion history and relative to the general population remain poorly quantified. METHODS: Data for patients aged ≥ 18 years with two confirmed PKLR mutations were obtained from the PK deficiency Natural History Study (NCT02053480). Frequencies of select conditions were compared with an age- and sex-matched cohort from a general insured US population without PK deficiency. RESULTS: Compared with the matched population (n = 1220), patients with PK deficiency (n = 122) had significantly higher lifetime rates of osteoporosis, liver cirrhosis, and pulmonary hypertension; splenectomy and cholecystectomy rates were also significantly higher in the 8 years before the index date. Sixty-five (53.3%) patients with PK deficiency were classified as regularly transfused, 30 (24.6%) as occasionally transfused, and 27 (22.1%) as never transfused. Regularly transfused patients were significantly more likely than never transfused patients to have had splenectomy, cholecystectomy, and/or thrombosis. Liver iron overload was reported in 62% of patients and occurred regardless of transfusion cohort. CONCLUSIONS: Even never transfused patients with PK deficiency had higher rates of select comorbidities and complications than individuals without PK deficiency.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica/epidemiología , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato/epidemiología , Adulto , Alelos , Anemia Hemolítica Congénita no Esferocítica/etiología , Comorbilidad , Femenino , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Mutación , Prevalencia , Piruvato Quinasa/genética , Errores Innatos del Metabolismo del Piruvato/etiología , Adulto JovenRESUMEN
BACKGROUND: Pyruvate kinase deficiency (PKD) is a rare, autosomal recessive red blood cell enzyme disorder, which leads to lifelong hemolytic anemia and associated complications from the disease and its management. METHODS: An international, multicenter registry enrolled 124 individuals younger than 18 years old with molecularly confirmed PKD from 29 centers. Retrospective and prospective clinical data were collected. RESULTS: There was a wide range in the age at diagnosis from 0 to 16 years. Presentation in the newborn period ranged from asymptomatic to neonatal jaundice to fulminant presentations of fetal distress, myocardial depression, and/or liver failure. Children <5 years old were significantly more likely to be transfused than children >12 to <18 years (53% vs. 14%, p = .0006), which correlated with the timing of splenectomy. Regular transfusions were most common in children with two severe PKLR variants. In regularly transfused children, the nadir hemoglobin goal varied considerably. Impact on quality of life was a common reason for treatment with regular blood transfusions and splenectomy. Splenectomy increased the hemoglobin and decreased transfusion burden in most children but was associated with infection or sepsis (12%) and thrombosis (1.3%) even during childhood. Complication rates were high, including iron overload (48%), perinatal complications (31%), and gallstones (20%). CONCLUSIONS: There is a high burden of disease in children with PKD, with wide practice variation in monitoring and treatment. Clinicians must recognize the spectrum of the manifestations of PKD for early diagnostic testing, close monitoring, and management to avoid serious complications in childhood.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato , Adolescente , Anemia Hemolítica Congénita no Esferocítica/diagnóstico , Anemia Hemolítica Congénita no Esferocítica/genética , Anemia Hemolítica Congénita no Esferocítica/terapia , Niño , Preescolar , Humanos , Estudios Prospectivos , Errores Innatos del Metabolismo del Piruvato/diagnóstico , Errores Innatos del Metabolismo del Piruvato/genética , Errores Innatos del Metabolismo del Piruvato/terapia , Calidad de Vida , Estudios RetrospectivosRESUMEN
An international, multicenter registry was established to collect retrospective and prospective clinical data on patients with pyruvate kinase (PK) deficiency, the most common glycolytic defect causing congenital nonspherocytic hemolytic anemia. Medical history and laboratory and radiologic data were retrospectively collected at enrollment for 254 patients with molecularly confirmed PK deficiency. Perinatal complications were common, including anemia that required transfusions, hyperbilirubinemia, hydrops, and prematurity. Nearly all newborns were treated with phototherapy (93%), and many were treated with exchange transfusions (46%). Children age 5 years and younger were often transfused until splenectomy. Splenectomy (150 [59%] of 254 patients) was associated with a median increase in hemoglobin of 1.6 g/dL and a decreased transfusion burden in 90% of patients. Predictors of a response to splenectomy included higher presplenectomy hemoglobin (P = .007), lower indirect bilirubin (P = .005), and missense PKLR mutations (P = .0017). Postsplenectomy thrombosis was reported in 11% of patients. The most frequent complications included iron overload (48%) and gallstones (45%), but other complications such as aplastic crises, osteopenia/bone fragility, extramedullary hematopoiesis, postsplenectomy sepsis, pulmonary hypertension, and leg ulcers were not uncommon. Overall, 87 (34%) of 254 patients had both a splenectomy and cholecystectomy. In those who had a splenectomy without simultaneous cholecystectomy, 48% later required a cholecystectomy. Although the risk of complications increases with severity of anemia and a genotype-phenotype relationship was observed, complications were common in all patients with PK deficiency. Diagnostic testing for PK deficiency should be considered in patients with apparent congenital hemolytic anemia and close monitoring for iron overload, gallstones, and other complications is needed regardless of baseline hemoglobin. This trial was registered at www.clinicaltrials.gov as #NCT02053480.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica/diagnóstico , Estudios de Asociación Genética , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato/diagnóstico , Adolescente , Adulto , Anemia Hemolítica Congénita no Esferocítica/etiología , Anemia Hemolítica Congénita no Esferocítica/metabolismo , Anemia Hemolítica Congénita no Esferocítica/terapia , Transfusión Sanguínea , Niño , Preescolar , Colecistectomía/efectos adversos , Colecistectomía/métodos , Terapia Combinada , Activación Enzimática , Femenino , Genotipo , Humanos , Lactante , Recién Nacido , Masculino , Persona de Mediana Edad , Mutación , Fenotipo , Piruvato Quinasa/metabolismo , Errores Innatos del Metabolismo del Piruvato/etiología , Errores Innatos del Metabolismo del Piruvato/metabolismo , Errores Innatos del Metabolismo del Piruvato/terapia , Esplenectomía/efectos adversos , Esplenectomía/métodos , Evaluación de Síntomas , Resultado del Tratamiento , Adulto JovenRESUMEN
Pyruvate kinase deficiency (PKD) is the most common cause of chronic hereditary non-spherocytic hemolytic anemia and results in a broad spectrum of disease. The diagnosis of PKD requires a high index of suspicion and judicious use of laboratory tests that may not always be informative, including pyruvate kinase enzyme assay and genetic analysis of the PKLR gene. A significant minority of patients with PKD have occult mutations in non-coding regions of PKLR which are missed on standard genetic tests. The biochemical consequences of PKD result in hemolytic anemia due to red cell pyruvate and ATP deficiency while simultaneously causing increased red cell 2,3-diphosphoglycerate, which facilitates oxygen unloading. This phenomenon, in addition to numerous other factors such as genetic background and differences in splenic function result in a poor correlation between symptoms and degree of anemia from patient to patient. Red cell transfusions should, therefore, be symptom-directed and not based on a hemoglobin threshold. Patients may experience specific complications, such as paravertebral extramedullary hematopoiesis and chronic debilitating icterus, which require personalized treatment. The decision to perform splenectomy or hematopoietic stem cell transplantation is nuanced and depends on disease burden and long-term outlook given that targeted therapeutics are in development. In recognition of the complicated nature of the disease and its management and the limitations of the PKD literature, an international working group of ten PKD experts convened to better define the disease burden and manifestations. This article summarizes the conclusions of this working group and is a guide for clinicians and investigators caring for patients with PKD.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica , Errores Innatos del Metabolismo del Piruvato , Anemia Hemolítica Congénita no Esferocítica/diagnóstico , Anemia Hemolítica Congénita no Esferocítica/genética , Anemia Hemolítica Congénita no Esferocítica/terapia , Eritrocitos , Humanos , Piruvato Quinasa/deficiencia , Piruvato Quinasa/genética , Errores Innatos del Metabolismo del Piruvato/etiología , Errores Innatos del Metabolismo del Piruvato/genéticaRESUMEN
Pyruvate kinase (PK) deficiency is a rare recessive congenital hemolytic anemia caused by mutations in the PKLR gene. This study reports the molecular features of 257 patients enrolled in the PKD Natural History Study. Of the 127 different pathogenic variants detected, 84 were missense and 43 non-missense, including 20 stop-gain, 11 affecting splicing, five large deletions, four in-frame indels, and three promoter variants. Within the 177 unrelated patients, 35 were homozygous and 142 compound heterozygous (77 for two missense, 48 for one missense and one non-missense, and 17 for two non-missense variants); the two most frequent mutations were p.R510Q in 23% and p.R486W in 9% of mutated alleles. Fifty-five (21%) patients were found to have at least one previously unreported variant with 45 newly described mutations. Patients with two non-missense mutations had lower hemoglobin levels, higher numbers of lifetime transfusions, and higher rates of complications including iron overload, extramedullary hematopoiesis, and pulmonary hypertension. Rare severe complications, including lower extremity ulcerations and hepatic failure, were seen more frequently in patients with non-missense mutations or with missense mutations characterized by severe protein instability. The PKLR genotype did not correlate with the frequency of complications in utero or in the newborn period. With ICCs ranging from 0.4 to 0.61, about the same degree of clinical similarity exists within siblings as it does between siblings, in terms of hemoglobin, total bilirubin, splenectomy status, and cholecystectomy status. Pregnancy outcomes were similar across genotypes in PK deficient women. This report confirms the wide genetic heterogeneity of PK deficiency.
Asunto(s)
Anemia Hemolítica Congénita no Esferocítica/genética , Estudios de Asociación Genética/métodos , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato/genética , Adolescente , Adulto , Niño , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Persona de Mediana Edad , Piruvato Quinasa/genética , Adulto JovenRESUMEN
BACKGROUND: Diamond-Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by anemia, short stature, congenital anomalies, and cancer predisposition. Most cases are due to mutations in genes encoding ribosomal proteins (RP) leading to RP haploinsufficiency. Effective treatments for the anemia of DBA include chronic red cell transfusions, long-term corticosteroid therapy, or hematopoietic stem cell transplantation. In a small patient series and in animal models, there have been hematologic responses to L-leucine with amelioration of anemia. The study objectives of this clinical trial were to determine feasibility, safety, and efficacy of L-leucine in transfusion-dependent patients with DBA. PROCEDURE: Patients ≥2 years of age received L-leucine 700 mg/m2 orally three times daily for nine months to determine a hematologic response and any improvement in growth (NCT01362595). RESULTS: This multicenter, phase I/II study enrolled 55 subjects; 43 were evaluable. There were 21 males; the median age at enrollment was 10.4 years (range, 2.5-46.1 years). No significant adverse events were attributable to L-leucine. Two subjects had a complete erythroid response and five had a partial response. Nine of 25, and 11 of 25, subjects experienced a positive weight and height percentile change, respectively, at the end of therapy. CONCLUSIONS: L-leucine is safe, resulted in an erythroid response in 16% of subjects with DBA, and led to an increase in weight and linear growth velocity in 36% and 44% of evaluable subjects, respectively. Further studies will be critical to understand the role of L-leucine in the management of patients with DBA.
Asunto(s)
Anemia de Diamond-Blackfan/terapia , Transfusión Sanguínea/métodos , Leucina/uso terapéutico , Adolescente , Adulto , Anemia de Diamond-Blackfan/patología , Niño , Preescolar , Terapia Combinada , Estudios de Factibilidad , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Proyectos Piloto , Pronóstico , Adulto JovenRESUMEN
Quality of response to immunosuppressive therapy and long-term outcomes for pediatric severe aplastic anemia remain incompletely characterized. Contemporary evidence to inform treatment of relapsed or refractory severe aplastic anemia for pediatric patients is also limited. The clinical features and outcomes for 314 children treated from 2002 to 2014 with immunosuppressive therapy for acquired severe aplastic anemia were analyzed retrospectively from 25 institutions in the North American Pediatric Aplastic Anemia Consortium. The majority of subjects (n=264) received horse anti-thymocyte globulin (hATG) plus cyclosporine (CyA) with a median 61 months follow up. Following hATG/CyA, 71.2% (95%CI: 65.3,76.6) achieved an objective response. In contrast to adult studies, the quality of response achieved in pediatric patients was high, with 59.8% (95%CI: 53.7,65.8) complete response and 68.2% (95%CI: 62.2,73.8) achieving at least a very good partial response with a platelet count ≥50×109L. At five years post-hATG/CyA, overall survival was 93% (95%CI: 89,96), but event-free survival without subsequent treatment was only 64% (95%CI: 57,69) without a plateau. Twelve of 171 evaluable patients (7%) acquired clonal abnormalities after diagnosis after a median 25.2 months (range: 4.3-71 months) post treatment. Myelodysplastic syndrome or leukemia developed in 6 of 314 (1.9%). For relapsed/refractory disease, treatment with a hematopoietic stem cell transplant had a superior event-free survival compared to second immunosuppressive therapy treatment in a multivariate analysis (HR=0.19, 95%CI: 0.08,0.47; P=0.0003). This study highlights the need for improved therapies to achieve sustained high-quality remission for children with severe aplastic anemia.
Asunto(s)
Anemia Aplásica/tratamiento farmacológico , Suero Antilinfocítico/administración & dosificación , Ciclosporina/administración & dosificación , Terapia de Inmunosupresión , Anemia Aplásica/epidemiología , Anemia Aplásica/patología , Suero Antilinfocítico/efectos adversos , Preescolar , Ciclosporina/efectos adversos , Femenino , Estudios de Seguimiento , Humanos , Lactante , Masculino , Estudios Retrospectivos , Estados Unidos/epidemiologíaRESUMEN
Pyruvate kinase deficiency (PKD) is the most common enzyme defect of glycolysis and an important cause of hereditary, nonspherocytic hemolytic anemia. The disease has a worldwide geographical distribution but there are no verified data regarding its frequency. Difficulties in the diagnostic workflow and interpretation of PK enzyme assay likely play a role. By the creation of a global PKD International Working Group in 2016, involving 24 experts from 20 Centers of Expertise we studied the current gaps in the diagnosis of PKD in order to establish diagnostic guidelines. By means of a detailed survey and subsequent discussions, multiple aspects of the diagnosis of PKD were evaluated and discussed by members of Expert Centers from Europe, USA, and Asia directly involved in diagnosis. Broad consensus was reached among the Centers on many clinical and technical aspects of the diagnosis of PKD. The results of this study are here presented as recommendations for the diagnosis of PKD and used to prepare a diagnostic algorithm. This information might be helpful for other Centers to deliver timely and appropriate diagnosis and to increase awareness in PKD.
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Anemia Hemolítica Congénita no Esferocítica/diagnóstico , Piruvato Quinasa/deficiencia , Errores Innatos del Metabolismo del Piruvato/diagnóstico , Anemia Hemolítica Congénita no Esferocítica/sangre , Anemia Hemolítica Congénita no Esferocítica/genética , Anemia Hemolítica Congénita no Esferocítica/fisiopatología , Artefactos , Recuento de Células Sanguíneas , Conservación de la Sangre , Análisis Mutacional de ADN , Eritrocitos/enzimología , Reacciones Falso Negativas , Reacciones Falso Positivas , Humanos , Piruvato Quinasa/sangre , Piruvato Quinasa/genética , Errores Innatos del Metabolismo del Piruvato/sangre , Errores Innatos del Metabolismo del Piruvato/genética , Errores Innatos del Metabolismo del Piruvato/fisiopatología , Valores de Referencia , Reticulocitos , Sensibilidad y Especificidad , Análisis de Secuencia de ADN , Espectrofotometría , Factores de TiempoRESUMEN
Alpha thalassemia due to nondeletional mutations usually leads to more severe disease than that caused by deletional mutations. Devastating outcomes such as hydrops fetalis can occur with two nondeletional mutations, therefore warranting DNA-based workup for suspected carriers with subtle hematological abnormalities for family counseling purposes. We describe three cases with hemoglobin (Hb) Adana, a nondeletional alpha chain mutation, compounded with an alpha globin gene deletion resulting in thalassemia intermedia. We review the literature, draw genotype-phenotype correlations from published cases of Hb Adana, and propose that this correlation can be used by clinicians to help direct diagnostic studies and urge hematologists to thoroughly workup high-risk patients.
Asunto(s)
Hemoglobinas Anormales/genética , Globinas alfa/genética , Talasemia alfa/genética , Preescolar , Codón/genética , Femenino , Eliminación de Gen , Estudios de Asociación Genética , Asesoramiento Genético , Humanos , Hidropesía Fetal/etiología , Hidropesía Fetal/genética , Masculino , Mutación Puntual , Eliminación de SecuenciaRESUMEN
Forkhead box M1 (FOXM1) belongs to the forkhead/winged-helix family of transcription factors and regulates a network of proliferation-associated genes. Its abnormal upregulation has been shown to be a key driver of cancer progression and an initiating factor in oncogenesis. FOXM1 is also highly expressed in stem/progenitor cells and inhibits their differentiation, suggesting that FOXM1 plays a role in the maintenance of multipotency. However, the exact molecular mechanisms by which FOXM1 regulates human stem/progenitor cells are still uncharacterized. To understand the role of FOXM1 in normal hematopoiesis, human cord blood CD34+ cells were transduced with FOXM1 short hairpin ribonucleic acid (shRNA) lentivirus. Knockdown of FOXM1 resulted in a 2-fold increase in erythroid cells compared to myeloid cells. Additionally, knockdown of FOXM1 increased bromodeoxyuridine (BrdU) incorporation in erythroid cells, suggesting greater proliferation of erythroid progenitors. We also observed that the defective phosphorylation of FOXM1 by checkpoint kinase 2 (CHK2) or cyclin-dependent kinases 1/2 (CDK1/2) increased the erythroid population in a manner similar to knockdown of FOXM1. Finally, we found that an inhibitor of FOXM1, forkhead domain inhibitor-6 (FDI-6), increased red blood cell numbers through increased proliferation of erythroid precursors. Overall, our data suggest a novel function of FOXM1 in normal human hematopoiesis.