Christine Barnerias

Assessment of interferon-related biomarkers in Aicardi-Goutières syndrome associated with mutations in TREX1, RNASEH2A, RNASEH2B, RNASEH2C, SAMHD1, and ADAR: A case-control study

BACKGROUND Aicardi-Goutières syndrome (AGS) is an inflammatory disorder caused by mutations in any of six genes (TREX1, RNASEH2A, RNASEH2B, RNASEH2C, SAMHD1, and ADAR). The disease is severe and effective treatments are urgently needed. We investigated the status of interferon-related biomarkers in patients with AGS with a view to future use in diagnosis and clinical trials. METHODS In this case-control study, samples were collected prospectively from patients with mutation-proven AGS. The expression of six interferon-stimulated genes (ISGs) was measured by quantitative PCR, and the median fold change, when compared with the median of healthy controls, was used to create an interferon score for each patient. Scores higher than the mean of controls plus two SD (>2·466) were designated as positive. Additionally, we collated historical data for interferon activity, measured with a viral cytopathic assay, in CSF and serum from mutation-positive patients with AGS. We also undertook neutralisation assays of interferon activity in serum, and looked for the presence of autoantibodies against a panel of interferon proteins. FINDINGS 74 (90%) of 82 patients had a positive interferon score (median 12·90, IQR 6·14-20·41) compared with two (7%) of 29 controls (median 0·93, IQR 0·57-1·30). Of the eight patients with a negative interferon score, seven had mutations in RNASEH2B (seven [27%] of all 26 patients with mutations in this gene). Repeat sampling in 16 patients was consistent for the presence or absence of an interferon signature on 39 of 41 occasions. Interferon activity (tested in 147 patients) was negatively correlated with age (CSF, r=-0·604; serum, r=-0·289), and was higher in CSF than in serum in 104 of 136 paired samples. Neutralisation assays suggested that measurable antiviral activity was related to interferon α production. We did not record significantly increased concentrations of autoantibodies to interferon subtypes in patients with AGS, or an association between the presence of autoantibodies and interferon score or serum interferon activity. INTERPRETATION AGS is consistently associated with an interferon signature, which is apparently sustained over time and can thus be used to differentiate patients with AGS from controls. If future studies show that interferon status is a reactive biomarker, the measurement of an interferon score might prove useful in the assessment of treatment efficacy in clinical trials. FUNDING European Unions Seventh Framework Programme; European Research Council.

Pyruvate dehydrogenase complex deficiency: four neurological phenotypes with differing pathogenesis.

Aim  To describe the phenotype and genotype of pyruvate dehydrogenase complex (PDHc) deficiency.

Epilepsy in Menkes Disease: Analysis of Clinical Stages

Summary:  Purpose: Epilepsy is one of the main features of Menkes disease (MD), although it is not described in depth. To determine the spectrum of epilepsy, we studied its main characteristics.

Safety and Efficacy of Rituximab in Severe Juvenile Dermatomyositis: Results from 9 Patients from the French Autoimmunity and Rituximab Registry

Objective. To evaluate the safety and efficacy of rituximab (RTX) in juvenile dermatomyositis (JDM) in off-trial patients. Methods. We conducted a multicenter prospective study of patients with JDM included in the French Autoimmunity and Rituximab (AIR) registry. Results. Nine patients with severe JDM were studied. The main indication for RTX treatment was severe and/or refractory muscle involvement (7 patients), severe calcinosis (1 patient), or severe chronic abdominal pain associated with abdominal lipomatosis (1 patient). RTX was associated with corticosteroids, immunosuppressive drugs, and plasma exchange therapy in 9/9, 5/9, and 2/9 patients, respectively. Mild infections of the calcinosis sites occurred in 2 patients and an infusion-related event in 1. Complete clinical response was achieved in 3/6 patients treated with RTX for muscle involvement. In these responders steroid therapy was stopped or tapered to < 15% of the baseline dosage, with no relapse, with a followup ranging from 1.3 to 3 years. Calcinosis did not improve in the 6 affected patients. Conclusion. This small series suggests that rituximab may be effective for treating muscle and skin involvement in a small subset of children with severe JDM, and that its safety profile was satisfactory. Further studies are needed to identify predictive factors of response to RTX in patients with severe JDM.

Mutation update and genotype-phenotype correlations of novel and previously described mutations in TPM2 and TPM3 causing congenital myopathies.

Mutations affecting skeletal muscle isoforms of the tropomyosin genes may cause nemaline myopathy, cap myopathy, core‐rod myopathy, congenital fiber‐type disproportion, distal arthrogryposes, and Escobar syndrome. We correlate the clinical picture of these diseases with novel (19) and previously reported (31) mutations of the TPM2 and TPM3 genes. Included are altogether 93 families: 53 with TPM2 mutations and 40 with TPM3 mutations. Thirty distinct pathogenic variants of TPM2 and 20 of TPM3 have been published or listed in the Leiden Open Variant Database (http://www.dmd.nl/). Most are heterozygous changes associated with autosomal‐dominant disease. Patients with TPM2 mutations tended to present with milder symptoms than those with TPM3 mutations, DA being present only in the TPM2 group. Previous studies have shown that five of the mutations in TPM2 and one in TPM3 cause increased Ca2+ sensitivity resulting in a hypercontractile molecular phenotype. Patients with hypercontractile phenotype more often had contractures of the limb joints (18/19) and jaw (6/19) than those with nonhypercontractile ones (2/22 and 1/22), whereas patients with the non‐hypercontractile molecular phenotype more often (19/22) had axial contractures than the hypercontractile group (7/19). Our in silico predictions show that most mutations affect tropomyosin–actin association or tropomyosin head‐to‐tail binding.

Molecular characterization of 82 patients with pyruvate dehydrogenase complex deficiency. Structural implications of novel amino acid substitutions in E1 protein

BACKGROUND Pyruvate dehydrogenase complex (PDHc) deficiencies are an important cause of primary lactic acidosis. Most cases result from mutations in the X-linked gene for the pyruvate dehydrogenase E1α subunit (PDHA1) while a few cases result from mutations in genes for E1β (PDHB), E2 (DLAT), E3 (DLD) and E3BP (PDHX) subunits or PDH-phosphatase (PDP1). AIM To report molecular characterization of 82 PDHc-deficient patients and analyze structural effects of novel missense mutations in PDHA1. METHODS PDHA1 variations were investigated first, by exon sequencing using a long range PCR product, gene dosage assay and cDNA analysis. Mutation scanning in PDHX, PDHB, DLAT and DLD cDNAs was further performed in unsolved cases. Novel missense mutations in PDHA1 were located on the tridimensional model of human E1 protein to predict their possible functional consequences. RESULTS PDHA1 mutations were found in 30 girls and 35 boys. Three large rearrangements, including two contiguous gene deletion syndrome were identified. Novel missense, frameshift and splicing mutations were also delineated and a nonsense mutation in a mosaic male. Mutations p.Glu75Ala, p.Arg88Ser, p.Arg119Trp, p.Gly144Asp, p.Pro217Arg, p.Arg235Gly, p.Tyr243Cys, p.Tyr243Ser, p.Arg245Gly, p.Pro250Leu, p.Gly278Arg, p.Met282Val, p.Gly298Glu in PDHA1 were predicted to impair active site channel conformation or subunit interactions. Six out of the seven patients with PDHB mutations displayed the recurrent p.Met101Val mutation; 9 patients harbored PDHX mutations and one patient DLD mutations. CONCLUSION We provide an efficient stepwise strategy for mutation screening in PDHc genes and expand the growing list of PDHA1 mutations analyzed at the structural level.

Detection of interferon alpha protein reveals differential levels and cellular sources in disease

Type I interferons (IFNs) are essential mediators of antiviral responses. These cytokines have been implicated in the pathogenesis of autoimmunity, most notably systemic lupus erythematosus (SLE), diabetes mellitus, and dermatomyositis, as well as monogenic type I interferonopathies. Despite a fundamental role in health and disease, the direct quantification of type I IFNs has been challenging. Using single-molecule array (Simoa) digital ELISA technology, we recorded attomolar concentrations of IFN&agr; in healthy donors, viral infection, and complex and monogenic interferonopathies. IFN&agr; protein correlated well with functional activity and IFN-stimulated gene expression. High circulating IFN&agr; levels were associated with increased clinical severity in SLE patients, and a study of the cellular source of IFN&agr; protein indicated disease-specific mechanisms. Measurement of IFN&agr; attomolar concentrations by digital ELISA will enhance our understanding of IFN biology and potentially improve the diagnosis and stratification of pathologies associated with IFN dysregulation.

Peripheral neuropathy and inborn errors of metabolism in adults

SummaryAlthough they are classically viewed as paediatric diseases, it is now recognized that inborn errors of metabolism (IEMs) can present at any age from childhood to adulthood. IEMs can involve the peripheral nervous system, mostly as part of a more diffuse neurological or systemic clinical picture. However, in some cases, the neuropathy can be the unique initial sign. Here, based on our personal experience and on a comprehensive literature analysis, we review IEMs causing neuropathies in adults. Diseases were classified according to the predominant type of neuropathies into (1) acute neuropathies, (2) mononeuropathy multiplex, (3) chronic axonal polyneuropathies, (4) chronic demyelinating polyneuropathies, (5) small-fibre neuropathies, and (6) lower motor neuron disease.

Mutations in human lipoyltransferase gene LIPT1 cause a Leigh disease with secondary deficiency for pyruvate and alpha-ketoglutarate dehydrogenase

BackgroundSynthesis and apoenzyme attachment of lipoic acid have emerged as a new complex metabolic pathway. Mutations in several genes involved in the lipoic acid de novo pathway have recently been described (i.e., LIAS, NFU1, BOLA3, IBA57), but no mutation was found so far in genes involved in the specific process of attachment of lipoic acid to apoenzymes pyruvate dehydrogenase (PDHc), α-ketoglutarate dehydrogenase (α-KGDHc) and branched chain α-keto acid dehydrogenase (BCKDHc) complexes.MethodsExome capture was performed in a boy who developed Leigh disease following a gastroenteritis and had combined PDH and α-KGDH deficiency with a unique amino acid profile that partly ressembled E3 subunit (dihydrolipoamide dehydrogenase / DLD) deficiency. Functional studies on patient fibroblasts were performed. Lipoic acid administration was tested on the LIPT1 ortholog lip3 deletion strain yeast and on patient fibroblasts.ResultsExome sequencing identified two heterozygous mutations (c.875C > G and c.535A > G) in the LIPT1 gene that encodes a mitochondrial lipoyltransferase which is thought to catalyze the attachment of lipoic acid on PDHc, α-KGDHc, and BCKDHc. Anti-lipoic acid antibodies revealed absent expression of PDH E2, BCKDH E2 and α-KGDH E2 subunits. Accordingly, the production of 14CO2 by patient fibroblasts after incubation with 14Cglucose, 14Cbutyrate or 14C3OHbutyrate was very low compared to controls. cDNA transfection experiments on patient fibroblasts rescued PDH and α-KGDH activities and normalized the levels of pyruvate and 3OHbutyrate in cell supernatants. The yeast lip3 deletion strain showed improved growth on ethanol medium after lipoic acid supplementation and incubation of the patient fibroblasts with lipoic acid decreased lactate level in cell supernatants.ConclusionWe report here a putative case of impaired free or H protein-derived lipoic acid attachment due to LIPT1 mutations as a cause of PDH and α-KGDH deficiencies. Our study calls for renewed efforts to understand the mechanisms of pathology of lipoic acid-related defects and their heterogeneous biochemical expression, in order to devise efficient diagnostic procedures and possible therapies.

MFN2, a new gene responsible for mitochondrial DNA depletion

Sir, we read with great interest the paper by Rouzier et al. (2012) reporting a novel dominant MFN2 missense mutation (c.629A>T, p.D210V) in a large family displaying an optic atrophy ‘plus’ phenotype. Importantly, the authors identified MFN2 as a new gene implicated in mitochondrial DNA instability as they found multiple mitochondrial DNA deletions in skeletal muscle of two adult patients. Moreover, they have shown that mitochondrial fusion is necessary to repair stress-induced damage to mitochondrial DNA. Here, we report a child with an early-onset progressive multi-systemic disorder and carrying a novel MFN2 missense mutation responsible for mitochondrial DNA depletion in skeletal muscle. This girl was the second child of healthy unrelated parents, born after an uneventful pregnancy. Her birth weight was 2700 g [−2 standard deviations (SD)], her length 45.5 cm (−2 SD) and her head circumference 35 cm (mean). At the age of 6 months she was examined because of developmental delay with hypotonia, decreased rate of growth in head circumference (−2 SD), failure to thrive and severe gastro-oesophageal reflux. Brain MRI, EEG, brainstem auditory evoked potentials, metabolic and genetic investigations were all normal. At 3 years of age, the patient could only stand up when aided and had no language acquisition although her level of understanding seemed good as did her social interaction and behaviour. Drooling, abnormal ocular pursuit, ataxia, dysmetria, areflexia, weakness of limbs and near permanent abnormal movements appeared progressively thereafter. EMG disclosed an axonal sensorimotor neuropathy. At 5 years of age, brainstem auditory evoked potentials revealed hearing loss on the right side and the amplitude of visual evoked responses was decreased while the electroretinogram was normal. Complementary aetiological explorations were negative. Orthopaedic …