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1.
Blood Adv ; 3(18): 2751-2763, 2019 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-31540902

RESUMO

The del(5q) myelodysplastic syndrome (MDS) is a distinct subtype of MDS, associated with deletion of the ribosomal protein S14 (RPS14) gene that results in macrocytic anemia. This study sought to identify novel targets for the treatment of patients with del(5q) MDS by performing an in vivo drug screen using an rps14-deficient zebrafish model. From this, we identified the secreted gelatinase matrix metalloproteinase 9 (MMP9). MMP9 inhibitors significantly improved the erythroid defect in rps14-deficient zebrafish. Similarly, treatment with MMP9 inhibitors increased the number of colony forming unit-erythroid colonies and the CD71+ erythroid population from RPS14 knockdown human BMCD34+ cells. Importantly, we found that MMP9 expression is upregulated in RPS14-deficient cells by monocyte chemoattractant protein 1. Double knockdown of MMP9 and RPS14 increased the CD71+ population compared with RPS14 single knockdown, suggesting that increased expression of MMP9 contributes to the erythroid defect observed in RPS14-deficient cells. In addition, transforming growth factor ß (TGF-ß) signaling is activated in RPS14 knockdown cells, and treatment with SB431542, a TGF-ß inhibitor, improved the defective erythroid development of RPS14-deficient models. We found that recombinant MMP9 treatment decreases the CD71+ population through increased SMAD2/3 phosphorylation, suggesting that MMP9 directly activates TGF-ß signaling in RPS14-deficient cells. Finally, we confirmed that MMP9 inhibitors reduce SMAD2/3 phosphorylation in RPS14-deficient cells to rescue the erythroid defect. In summary, these study results support a novel role for MMP9 in the pathogenesis of del(5q) MDS and the potential for the clinical use of MMP9 inhibitors in the treatment of patients with del(5q) MDS.

2.
N Engl J Med ; 381(10): 933-944, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31483964

RESUMO

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.).


Assuntos
Anemia Hemolítica Congênita não Esferocítica/tratamento farmacológico , Hemoglobinas/metabolismo , Piruvato Quinase/deficiência , Erros Inatos do Metabolismo dos Piruvatos/tratamento farmacológico , Administração Oral , Adolescente , Adulto , Anemia Hemolítica Congênita não Esferocítica/sangue , Anemia Hemolítica Congênita não Esferocítica/genética , Catecóis , Esquema de Medicação , Feminino , Seguimentos , Cefaleia/induzido quimicamente , Humanos , Masculino , Mutação , Piruvato Quinase/sangue , Piruvato Quinase/genética , Erros Inatos do Metabolismo dos Piruvatos/sangue , Erros Inatos do Metabolismo dos Piruvatos/genética , Distúrbios do Início e da Manutenção do Sono/induzido quimicamente , Tirfostinas , Adulto Jovem
3.
Haematologica ; 104(10): 1974-1983, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30948484

RESUMO

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.

5.
Am J Hum Genet ; 2018 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-30503522

RESUMO

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.

6.
Am J Hematol ; 2018 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-30358897

RESUMO

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.

9.
Pediatr Blood Cancer ; 65(9): e27220, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29749692

RESUMO

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.

10.
Pediatr Clin North Am ; 65(3): 579-595, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29803284

RESUMO

Mature red blood cells are reliant on the glycolytic pathway for energy production and the hexose monophosphate shunt for cell protection from oxidative insults. The most common red blood cell enzyme disorders are characterized by hemolysis but with wide clinical variability. Glucose-6-phosphate dehydrogenase deficiency is the most common red cell enzyme disorder worldwide. Frequent clinical presentations include neonatal jaundice and episodic hemolysis after exposure to oxidative stress. Symptoms of pyruvate kinase deficiency and other glycolytic enzyme disorders include neonatal jaundice, chronic hemolytic anemia, gallstones, and transfusion-related and transfusion-independent iron overload. Diagnosis is critical for appropriate supportive care, monitoring, and treatment.


Assuntos
Anemia Hemolítica Congênita não Esferocítica/diagnóstico , Anemia Hemolítica Congênita não Esferocítica/etiologia , Deficiência de Glucosefosfato Desidrogenase/diagnóstico , Deficiência de Glucosefosfato Desidrogenase/etiologia , Piruvato Quinase/deficiência , Erros Inatos do Metabolismo dos Piruvatos/diagnóstico , Erros Inatos do Metabolismo dos Piruvatos/etiologia , Deficiência de Glucosefosfato Desidrogenase/terapia , Humanos , Piruvato Quinase/uso terapêutico
11.
Blood ; 131(20): 2183-2192, 2018 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-29549173

RESUMO

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.

12.
Haematologica ; 102(5): 826-834, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28154085

RESUMO

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.


Assuntos
Proliferação de Células/genética , Células Precursoras Eritroides/metabolismo , Eritropoese/genética , Proteína Forkhead Box M1/genética , Antígenos CD34/sangue , Diferenciação Celular/genética , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Quinase do Ponto de Checagem 2/metabolismo , Células Eritroides/metabolismo , Sangue Fetal/citologia , Sangue Fetal/metabolismo , Proteína Forkhead Box M1/metabolismo , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Células K562 , Fosforilação , Piridinas/farmacologia , Interferência de RNA , Tiofenos/farmacologia
16.
Transfusion ; 56(5): 1182-1184, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26996653

RESUMO

BACKGROUND: Although rare in the United States, anti-Mur is relatively common in Southeast Asia and has been reported to have clinical significance in Chinese and Taiwanese populations. STUDY DESIGN AND METHODS: The infant was full term and the second child of a Chinese mother and Vietnamese father, presenting with jaundice. He was clinically diagnosed with immune-mediated hemolytic anemia. RESULTS: The direct antiglobulin test indicated that the infant's red blood cells were coated only with anti-IgG. Anti-Mur was identified in the maternal serum and the neonate's plasma. The father was found to be positive for the Mur antigen. The cause of the infant's hemolytic anemia was determined to be most likely anti-Mur. CONCLUSION: Since anti-Mur is implicated in causing hemolytic disease of the newborn, it is important to recognize this antibody more commonly found in Asian patients in the United States as the Mur+ phenotype has a higher prevalence in this population.

18.
J Pediatr Surg ; 51(1): 122-7, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26613837

RESUMO

PURPOSE: The purpose of this study was to define the hematologic response to total splenectomy (TS) or partial splenectomy (PS) in children with hereditary spherocytosis (HS) or sickle cell disease (SCD). METHODS: The Splenectomy in Congenital Hemolytic Anemia (SICHA) consortium registry collected hematologic outcomes of children with CHA undergoing TS or PS to 1 year after surgery. Using random effects mixed modeling, we evaluated the association of operative type with change in hemoglobin, reticulocyte counts, and bilirubin. We also compared laparoscopic to open splenectomy. RESULTS: The analysis included 130 children, with 62.3% (n=81) undergoing TS. For children with HS, all hematologic measures improved after TS, including a 4.1g/dl increase in hemoglobin. Hematologic parameters also improved after PS, although the response was less robust (hemoglobin increase 2.4 g/dl, p<0.001). For children with SCD, there was no change in hemoglobin. Laparoscopy was not associated with differences in hematologic outcomes compared to open. TS and laparoscopy were associated with shorter length of stay. CONCLUSION: Children with HS have an excellent hematologic response after TS or PS, although the hematologic response is more robust following TS. Children with SCD have smaller changes in their hematologic parameters. These data offer guidance to families and clinicians considering TS or PS.


Assuntos
Anemia Falciforme/sangue , Anemia Falciforme/cirurgia , Esferocitose Hereditária/sangue , Esferocitose Hereditária/cirurgia , Esplenectomia/métodos , Adolescente , Bilirrubina/sangue , Criança , Feminino , Hemoglobinas/metabolismo , Humanos , Laparoscopia , Masculino , Sistema de Registros , Contagem de Reticulócitos
19.
Pediatrics ; 136(5): e1268-75, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26459646

RESUMO

BACKGROUND AND OBJECTIVES: Widespread newborn screening on a point-of-care basis could prevent bilirubin neurotoxicity in newborns with glucose-6-phosphate dehydrogenase (G6PD) deficiency. We evaluated a quantitative G6PD assay on a digital microfluidic platform by comparing its performance with standard clinical methods. METHODS: G6PD activity was measured quantitatively by using digital microfluidic fluorescence and the gold standard fluorescence biochemical test on a convenience sample of 98 discarded blood samples. Twenty-four samples were designated as G6PD deficient. RESULTS: Mean ± SD G6PD activity for normal samples using the digital microfluidic method and the standard method, respectively, was 9.7 ± 2.8 and 11.1 ± 3.0 U/g hemoglobin (Hb), respectively; for G6PD-deficient samples, it was 0.8 ± 0.7 and 1.4 ± 0.9 U/g Hb. Bland-Altman analysis determined a mean difference of -0.96 ± 1.8 U/g Hb between the digital microfluidic fluorescence results and the standard biochemical test results. The lower and upper limits for the digital microfluidic platform were 4.5 to 19.5 U/g Hb for normal samples and 0.2 to 3.7 U/g Hb for G6PD-deficient samples. The lower and upper limits for the Stanford method were 5.5 to 20.7 U/g Hb for normal samples and 0.1 to 2.8 U/g Hb for G6PD-deficient samples. The measured activity discriminated between G6PD-deficient samples and normal samples with no overlap. CONCLUSIONS: Pending further validation, a digital microfluidics platform could be an accurate point-of-care screening tool for rapid newborn G6PD screening.


Assuntos
Deficiência de Glucosefosfato Desidrogenase/diagnóstico , Testes Imediatos , Adulto , Criança , Feminino , Humanos , Lactente , Recém-Nascido , Masculino
20.
Am J Hematol ; 90(9): 825-30, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26087744

RESUMO

Over the last several decades, our understanding of the genetic variation, pathophysiology, and complications of the hemolytic anemia associated with red cell pyruvate kinase deficiency (PKD) has expanded. Nonetheless, there remain significant gaps in our knowledge with regard to clinical care and monitoring. Treatment remains supportive with phototherapy and/or exchange transfusion in the newborn period, regular or intermittent red cell transfusions in children and adults, and splenectomy to decrease transfusion requirements and/or anemia related symptoms. In this article, we review the clinical diversity of PKD, the current standard of treatment and for supportive care, the complications observed, and future treatment directions.


Assuntos
Anemia Hemolítica Congênita não Esferocítica/terapia , Transfusão de Eritrócitos , Transfusão Total , Piruvato Quinase/deficiência , Erros Inatos do Metabolismo dos Piruvatos/terapia , Adulto , Anemia Hemolítica Congênita não Esferocítica/enzimologia , Anemia Hemolítica Congênita não Esferocítica/patologia , Anemia Hemolítica Congênita não Esferocítica/cirurgia , Criança , Gerenciamento Clínico , Humanos , Recém-Nascido , Fototerapia , Erros Inatos do Metabolismo dos Piruvatos/enzimologia , Erros Inatos do Metabolismo dos Piruvatos/patologia , Erros Inatos do Metabolismo dos Piruvatos/cirurgia , Esplenectomia
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