Late effects after hematopoietic stem cell transplantation for ß-thalassemia major: the French national experience.

In this retrospective study, we evaluate long-term complications in nearly all ß-thalassemia-major patients who successfully received allogeneic hematopoietic stem cell transplantation in France. Ninety-nine patients were analyzed with a median age of 5.9 years at transplantation. The median duration of clinical follow up was 12 years. All conditioning regimens were myeloablative, most were based on busulfan combined with cyclophosphamide, and more than 90% of patients underwent a transplant from a matched sibling donor. After transplantation, 11% of patients developed thyroid dysfunction, 5% diabetes, and 2% heart failure. Hypogonadism was present in 56% of females and 14% of males. Female patients who went on to normal puberty after transplant were significantly younger at transplantation than those who experienced delayed puberty (median age 2.5 vs 8.7 years). Fertility was preserved in 9 of 27 females aged 20 years or older and 2 other patients became pregnant following oocyte donation. In addition to patient's age and higher serum ferritin levels at transplantation, time elapsed since transplant was significantly associated with decreased height growth in multivariate analysis. Weight growth increased after transplantation particularly in females, 36% of adults being overweight at last evaluation. A comprehensive long-term monitoring, especially of endocrine late effects, is required after hematopoietic stem cell transplantation for thalassemia.

Platelet and not erythrocyte microparticles are procoagulant in transfused thalassaemia major patients.

The level of circulating platelet-, erythrocyte-, leucocyte- and endothelial-derived microparticles detected by high-sensitivity flow cytometry was investigated in 37 ß-thalassaemia major patients receiving a regular transfusion regimen. The phospholipid procoagulant potential of the circulating microparticles and the microparticle-dependent tissue factor activity were evaluated. A high level of circulating erythrocyte- and platelet-microparticles was found. In contrast, the number of endothelial microparticles was within the normal range. Platelet microparticles were significantly higher in splenectomized than in non-splenectomized patients, independent of platelet count (P < 0·001). Multivariate analysis indicated that phospholipid-dependent procoagulant activity was influenced by both splenectomy (P = 0·001) and platelet microparticle level (P < 0·001). Erythrocyte microparticles were not related to splenectomy, appear to be devoid of proper procoagulant activity and no relationship between their production and haemolysis, dyserythropoiesis or oxidative stress markers could be established. Intra-microparticle labelling with anti-HbF antibodies showed that they originate only partially (median of 28%) from thalassaemic erythropoiesis. In conclusion, when ß-thalassaemia major patients are intensively transfused, the procoagulant activity associated with thalassaemic erythrocyte microparticles is probably diluted by transfusions. In contrast, platelet microparticles, being both more elevated and more procoagulant, especially after splenectomy, may contribute to the residual thrombotic risk reported in splenectomized multi-transfused ß-thalassaemia major patients.

PRRT2 mutations: a major cause of paroxysmal kinesigenic dyskinesia in the European population.

OBJECTIVE: Paroxysmal kinesigenic dyskinesia (PKD) is a rare disorder characterized by recurrent attacks of hyperkinetic movements. PKD can be isolated or associated with benign infantile seizures as part of the infantile convulsions with choreoathetosis (ICCA) syndrome. Mutations in the PRRT2 gene were recently identified in patients with PKD and ICCA. We studied the prevalence of PRRT2 mutations and characteristics of the patients in a European population of patients with PKD and ICCA. METHODS: Patients were recruited through the 1996-2011 database of our DNA bank, to which physicians refer DNA with a putative diagnosis and clinical information. Two movement disorders experts reviewed the information on patients with a putative diagnosis of PKD. Patients who fulfilled the criteria for PKD and ICCA were included. The PRRT2 coding sequence was analyzed by direct sequencing. RESULTS: Among 42 index cases of unrelated families referred with a putative diagnosis of PKD, a total of 34 patients, including 32 with isolated PKD and 2 with ICCA, were selected for genetic analysis. Mutations introducing premature termination codons were identified in 22 of 34 patients including 13 of 14 families and 9 of 20 patients with sporadic cases. The previously described c.649dupC/pArg217ProfsX8 and c.629dupC/pAla211SerfsX14 were present, respectively, in 17 patients and 1 patient; we also report 3 novel mutations: c.649delC/pArg217GlufsX12 in 2 patients, and c.562C>T/pGln188X and c.649C>T/pArg217X, each in 1 patient. The group with mutations was characterized by a younger age at onset (9 years) compared with the patients without mutations (15 years; p < 0.01). CONCLUSION: Mutations in PRRT2 are a major cause of PKD in familial and sporadic cases in the European population.

Lafora progressive myoclonus epilepsy: NHLRC1 mutations affect glycogen metabolism.

Lafora disease is a fatal autosomal recessive form of progressive myoclonus epilepsy. Patients manifest myoclonus and tonic-clonic seizures, visual hallucinations, intellectual, and progressive neurologic deterioration beginning in adolescence. The two genes known to be involved in Lafora disease are EPM2A and NHLRC1 (EPM2B). The EPM2A gene encodes laforin, a dual-specificity protein phosphatase, and the NHLRC1 gene encodes malin, an E3-ubiquitin ligase. The two proteins interact with each other and, as a complex, are thought to regulate glycogen synthesis. Here, we report three Lafora families with two novel pathogenic mutations (C46Y and L261P) and two recurrent mutations (P69A and D146N) in NHLRC1. Investigation of their functional consequences in cultured mammalian cells revealed that malin(C46Y), malin(P69A), malin(D146N), and malin(L261P) mutants failed to downregulate the level of R5/PTG, a regulatory subunit of protein phosphatase 1 involved in glycogen synthesis. Abnormal accumulation of intracellular glycogen was observed with all malin mutants, reminiscent of the polyglucosan inclusions (Lafora bodies) present in patients with Lafora disease.

Mutations and deletions in PCDH19 account for various familial or isolated epilepsies in females.

Mutations in PCDH19, encoding protocadherin 19 on chromosome X, cause familial epilepsy and mental retardation limited to females or Dravet-like syndrome. Heterozygous females are affected while hemizygous males are spared, this unusual mode of inheritance being probably due to a mechanism called cellular interference. To extend the mutational and clinical spectra associated with PCDH19, we screened 150 unrelated patients (113 females) with febrile and afebrile seizures for mutations or rearrangements in the gene. Fifteen novel point mutations were identified in 15 female patients (6 sporadic and 9 familial cases). In addition, qPCR revealed two whole gene deletions and one partial deletion in 3 sporadic female patients. Clinical features were highly variable but included almost constantly a high sensitivity to fever and clusters of brief seizures. Interestingly, cognitive functions were normal in several family members of 2 families: the familial condition in family 1 was suggestive of Generalized Epilepsy with Febrile Seizures Plus (GEFS+) whereas all three affected females had partial cryptogenic epilepsy. These results show that mutations in PCDH19 are a relatively frequent cause of epilepsy in females and should be considered even in absence of family history and/or mental retardation.

A dominant mutation in the gene encoding the erythroid transcription factor KLF1 causes a congenital dyserythropoietic anemia.

The congenital dyserythropoietic anemias (CDAs) are inherited red blood cell disorders whose hallmarks are ineffective erythropoiesis, hemolysis, and morphological abnormalities of erythroblasts in bone marrow. We have identified a missense mutation in KLF1 of patients with a hitherto unclassified CDA. KLF1 is an erythroid transcription factor, and extensive studies in mouse models have shown that it plays a critical role in the expression of globin genes, but also in the expression of a wide spectrum of genes potentially essential for erythropoiesis. The unique features of this CDA confirm the key role of KLF1 during human erythroid differentiation. Furthermore, we show that the mutation has a dominant-negative effect on KLF1 transcriptional activity and unexpectedly abolishes the expression of the water channel AQP1 and the adhesion molecule CD44. Thus, the study of this disease-causing mutation in KLF1 provides further insights into the roles of this transcription factor during erythropoiesis in humans.

Novel mutations in EPM2A and NHLRC1 widen the spectrum of Lafora disease.

PURPOSE: Lafora disease (LD) is an autosomal recessive form of progressive myoclonus epilepsy with onset in childhood or adolescence and with fatal outcome caused by mutations in two genes: EPM2A and NHLRC1. The aim of this study was to characterize the mutation spectrum in a cohort of unrelated patients with presumed LD. METHODS: Sequencing of the two genes and search for large rearrangements was performed in 46 unrelated patients with suspected LD, 33 originating from France and the others from different countries. Patients were classified into two groups according to the clinical presentation. RESULTS: Mutations of various types were found in EPM2A in 10 patients and in NHLRC1 in 4 patients. Mutations were found in 14 (93%) of 15 patients with classical clinical and electroencephalography (EEG) presentation of LD and in no patients with an atypical presentation. Ten mutations were novel, including the first substitution reported in a donor splice site of EPM2A, leading to the deletion of exon 2 at the RNA level. Four large deletions, including two deletions of exon 2 with different sizes and breakpoints, were found in EPM2A, corresponding to 20% of the alleles of this gene. DISCUSSION: We described several novel mutations of EPM2A and NHLRC1 and brought additional data to the genetic epidemiology of LD. This study emphasized the high mutation rate in patients with classical LD as well as the high negativity rate of skin biopsy.

Complications and treatment of patients with ß-thalassemia in France: results of the National Registry.

BACKGROUND: beta-thalassemia is a rare disease in France, encountered mainly in patients originating from Italy and North Africa. In the setting of the recent French plan for rare diseases, a National Registry for thalassemia has been developed since 2005. Epidemiological and clinical data have been collected on living patients with beta-thalassemia major or intermedia, including those who underwent hematopoietic stem cell transplantation. DESIGN AND METHODS: A standardized questionnaire was sent to clinicians throughout the national professional networks involved in the management of thalassemic patients and data were updated every 18 months. A cross-sectional study was performed in February 2009. RESULTS: Data on 378 patients (267 with thalassemia major) with a median age of 20 were recorded. Hematopoietic stem cell transplantation was performed in 52 patients. Stature, rates of parenthood, splenectomy, and cholecystectomy were no different between non-transplanted thalassemia major and thalassemia intermedia patients, after adjustment for age. Among the 215 non-transplanted thalassemia major patients, the median serum ferritin level was 1240 ng/mL and the rates of iron-related complications were 10%, 6%, 10% and 48% for cardiac failure, diabetes, hypothyroidism, and hypogonadism, respectively. From 2005 to 2008, a dramatic switch in chelation treatment, from deferoxamine to deferasirox, was observed. CONCLUSIONS: The rates of complications of iron overload in French thalassemia major patients appeared similar to those reported in other developed countries in which this condition is not endemic. There were no significant differences in height and parenthood rates between patients with the major and the intermedia forms of the disease, underlining the progress in clinical care. Future developments will focus on mortality and morbidity under oral chelation treatment.