Publications by authors named "Caroline Sevin"

16 Publications

  • Page 1 of 1

Adenosine kinase deficiency: Three new cases and diagnostic value of hypermethioninemia.

Mol Genet Metab 2021 Jan 28;132(1):38-43. Epub 2020 Nov 28.

Department of Biochemistry, Reference Center for Mitochondrial Disease, FILNEMUS, Bicêtre University Hospital, University of Paris-Saclay, Assistance Publique-Hôpitaux de Paris, Le Kremlin-Bicêtre, France; Lip(Sys)2, University of Paris-Saclay, Chatenay-Malabry, France. Electronic address:

Adenosine kinase (ADK) deficiency is characterized by liver disease, dysmorphic features, epilepsy and developmental delay. This defect disrupts the adenosine/AMP futile cycle and interferes with the upstream methionine cycle. We report the clinical, histological and biochemical courses of three ADK children carrying two new mutations and presenting with neonatal cholestasis and neurological disorders. One of them died of liver failure whereas the other two recovered from their liver damage. As the phenotype was consistent with a mitochondrial disorder, we studied liver mitochondrial respiratory chain activities in two patients and revealed a combined defect of several complexes. In addition, we retrospectively analyzed methionine plasma concentration, a hallmark of ADK deficiency, in a cohort of children and showed that methionine level in patients with ADK deficiency was strongly increased compared with patients with other liver diseases. ADK deficiency is a cause of neonatal or early infantile liver disease that may mimic primary mitochondrial disorders. In this context, an elevation of methionine plasma levels over twice the upper limit should not be considered as a nonspecific finding. ADK deficiency induced-liver dysfunction is most often transient, but could be life-threatening.
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http://dx.doi.org/10.1016/j.ymgme.2020.11.007DOI Listing
January 2021

The Challenge of Gene Therapy for Neurological Diseases: Strategies and Tools to Achieve Efficient Delivery to the Central Nervous System.

Hum Gene Ther 2021 Jan 26. Epub 2021 Jan 26.

NeuroGenCell, INSERM U1127, Paris Brain Institute (ICM), Sorbonne University, CNRS, AP-HP, University Hospital Pitié-Salpêtrière, Paris, France.

For more than 10 years, gene therapy for neurological diseases has experienced intensive research growth and more recently therapeutic interventions for multiple indications. Beneficial results in several phase 1/2 clinical studies, together with improved vector technology have advanced gene therapy for the central nervous system (CNS) in a new era of development. Although most initial strategies have focused on orphan genetic diseases, such as lysosomal storage diseases, more complex and widespread conditions like Alzheimer's disease, Parkinson's disease, epilepsy, or chronic pain are increasingly targeted for gene therapy. Increasing numbers of applications and patients to be treated will require improvement and simplification of gene therapy protocols to make them accessible to the largest number of affected people. Although vectors and manufacturing are a major field of academic research and industrial development, there is a growing need to improve, standardize, and simplify delivery methods. Delivery is the major issue for CNS therapies in general, and particularly for gene therapy. The blood-brain barrier restricts the passage of vectors; strategies to bypass this obstacle are a central focus of research. In this study, we present the different ways that can be used to deliver gene therapy products to the CNS. We focus on results obtained in large animals that have allowed the transfer of protocols to human patients and have resulted in the generation of clinical data. We discuss the different routes of administration, their advantages, and their limitations. We describe techniques, equipment, and protocols and how they should be selected for safe delivery and improved efficiency for the next generation of gene therapy trials for CNS diseases.
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http://dx.doi.org/10.1089/hum.2020.105DOI Listing
January 2021

Safety of intrathecal delivery of recombinant human arylsulfatase A in children with metachromatic leukodystrophy: Results from a phase 1/2 clinical trial.

Mol Genet Metab 2020 Sep - Oct;131(1-2):235-244. Epub 2020 Jul 16.

Shire, a member of the Takeda group of companies, Lexington, MA, USA. Electronic address:

Background: Metachromatic leukodystrophy (MLD) is an autosomal recessive disorder caused by deficient arylsulfatase A (ASA) activity and characterized by neurological involvement that results in severe disability and premature death. We examined the safety and tolerability of intrathecally delivered recombinant human ASA (rhASA; SHP611, now TAK-611) in children with MLD (NCT01510028). Secondary endpoints included change in cerebrospinal fluid (CSF) sulfatide and lysosulfatide levels, and motor function (assessed by Gross Motor Function Measure-88 total score).

Methods: Twenty-four children with MLD who experienced symptom onset aged ≤ 30 months were enrolled. Patients received rhASA every other week (EOW) for 38 weeks at 10, 30, or 100 mg (cohorts 1-3; n = 6 per cohort), or 100 mg manufactured using a revised process (cohort 4; n = 6).

Results: No rhASA-related serious adverse events (SAEs) were observed; 25% of patients experienced an SAE related to the intrathecal device or drug delivery method. Mean CSF sulfatide and lysosulfatide levels fell to within normal ranges in both 100 mg cohorts following treatment. Although there was a general decline in motor function over time, there was a tendency towards a less pronounced decline in patients receiving 100 mg.

Conclusion: Intrathecal rhASA was generally well tolerated at doses up to 100 mg EOW. These preliminary data support further development of rhASA as a therapy for patients with MLD.
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http://dx.doi.org/10.1016/j.ymgme.2020.07.002DOI Listing
July 2020

Intra-CSF AAV9 and AAVrh10 Administration in Nonhuman Primates: Promising Routes and Vectors for Which Neurological Diseases?

Mol Ther Methods Clin Dev 2020 Jun 11;17:771-784. Epub 2020 Apr 11.

UMR PAnTher 703 INRA/Oniris Animal Pathophysiology and Bio Therapy for Muscle and Nervous System Diseases, Oniris, Nantes-Atlantic College of Veterinary Medicine Food Sciences and Engineering, 44307 Nantes Cedex 03, France.

The identification of the most efficient method for whole central nervous system targeting that is translatable to humans and the safest route of adeno-associated virus (AAV) administration is a major concern for future applications in clinics. Additionally, as many AAV serotypes were identified for gene introduction into the brain and the spinal cord, another key to human gene-therapy success is to determine the most efficient serotype. In this study, we compared lumbar intrathecal administration through catheter implantation and intracerebroventricular administration in the cynomolgus macaque. We also evaluated and compared two AAV serotypes that are currently used in clinical trials: AAV9 and AAVrh10. We demonstrated that AAV9 lumbar intrathecal delivery using a catheter achieved consistent transgene expression in the motor neurons of the spinal cord and in the neurons/glial cells of several brain regions, whereas AAV9 intracerebroventricular delivery led to a consistent transgene expression in the brain. In contrast, AAVrh10 lumbar intrathecal delivery led to rare motor neuron targeting. Finally, we found that AAV9 efficiently targets respiratory and skeletal muscles after injection into the cerebrospinal fluid (CSF), which represents an outstanding new property that can be useful for the treatment of diseases affecting both the central nervous system and muscle.
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http://dx.doi.org/10.1016/j.omtm.2020.04.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7184633PMC
June 2020

Mild Encephalitis/Encephalopathy with reversible splenial lesion syndrome: An unusual presentation of anti-GFAP astrocytopathy.

Eur J Paediatr Neurol 2020 May 18;26:89-91. Epub 2020 Mar 18.

Assistance Publique-Hôpitaux de Paris, Hôpitaux Universitaires Paris-Sud, Hôpital Bicêtre, Pediatric Neurology Department, National Referral Center for Rare Inflammatory Brain and Spinal Diseases, France; Université Paris-Sud, UMR 1184-CEA-IDMIT, Center for Immunology of Viral Infections and Autoimmune Diseases, 94275, Le Kremlin Bicêtre, France. Electronic address:

Autoimmune glial fibrillary acidic protein (GFAP) astrocytopathy is a rare recently defined antibody-mediated encephalitis. Meningo-encephalomyelitis presentation is frequent with lymphocytic pleiocytosis in the cerebro-spinal fluid and brain MRI classically demonstrates in 50% of cases, a linear perivascular enhancement extending radially from the ventricles. Here, we describe 2 cases of pediatric autoimmune GFAP astrocytopathy with limbic encephalitis presentation and peculiar MRI characteristics: one with normal MRI and the second suggestive of Mild Encephalitis/Encephalopathy with reversible splenial lesion syndrome (MERS). These two cases illustrate that anti-GFAP antibodies should be sought in children presenting limbic encephalitis with a normal and/or MERS suggestive MRI, as treatment strategies may differ.
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http://dx.doi.org/10.1016/j.ejpn.2020.03.002DOI Listing
May 2020

Fatigue, depression, and quality of life in children with multiple sclerosis: a comparative study with other demyelinating diseases.

Dev Med Child Neurol 2020 02 11;62(2):241-244. Epub 2019 Apr 11.

Pediatric Neurology Department, National Referral Center for Rare Inflammatory Brain and Spinal Diseases, Assistance Publique-Hôpitaux de Paris, Hôpital Bicêtre, Hôpitaux Universitaires Paris-Sud, Le Kremlin Bicêtre, France.

Aim: To evaluate fatigue, depression, and quality of life (QoL) of children with multiple sclerosis and compare to other acute demyelinating syndromes (ADS).

Method: Children followed in the National Referral Centre of rare inflammatory brain and spinal diseases were included in this study. The Expanded Disability Status Scale, the fatigue severity scale, the Multiscore Depression Inventory for Children, and the Pediatric Quality of Life Inventory were used for evaluation.

Results: Thirty-seven children (23 females, 14 males) were included in this study. Multiple sclerosis was diagnosed in 26 children and ADS in 11 children. Although not significant, severe fatigue was less frequently reported by patients with multiple sclerosis than children with ADS (44% vs 63%, p=0.2). Depression was reported more often in the multiple sclerosis group compared to the ADS group (24% vs 18%, p=0.6). Concerning the QoL in patients with multiple sclerosis, both parents and children reported poor emotional and school functioning. Physical and social functioning were rated as being good in both groups, and was significantly higher in the children's group (p=0.007).

Interpretation: This study highlights the importance of fatigue and depression in children with ADS and particularly in paediatric onset multiple sclerosis. Moreover, difficulties in school and emotional functioning were the main concerns for parents and children in the multiple sclerosis group which need to be taken in account during their care and treatment proposal.

What This Paper Adds: Invisible signs such as fatigue and depression affect all forms of acute demyelinating syndromes (ADS) in children. Depression seems to be higher in children with multiple sclerosis than with other forms of ADS. Fatigue seems to be lower in children with multiple sclerosis than with other forms of ADS. Children with multiple sclerosis and their parents are most concerned with emotional and academic functioning.
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http://dx.doi.org/10.1111/dmcn.14242DOI Listing
February 2020

Intrathecal heparan-N-sulfatase in patients with Sanfilippo syndrome type A: A phase IIb randomized trial.

Mol Genet Metab 2019 02 24;126(2):121-130. Epub 2018 Oct 24.

Shire, Lexington, MA, USA. Electronic address:

Background: Sanfilippo syndrome type A (mucopolysaccharidosis type IIIA) is a lysosomal disorder wherein deficient heparan-N-sulfatase (HNS) activity results in the accumulation of heparan sulfate in the central nervous system and is associated with progressive neurodegeneration in early childhood. We report on the efficacy, pharmacokinetics, safety, and tolerability of intrathecal (IT) administration of recombinant human HNS (rhHNS) from a phase IIb randomized open-label trial.

Methods: Twenty-one patients, randomized 1:1:1 to rhHNS IT 45 mg administered every 2 weeks (Q2W), every 4 weeks (Q4W), or no treatment, were assessed for amelioration in neurocognitive decline as determined by the Bayley Scales of Infant and Toddler Development®, Third Edition. The primary efficacy goal was defined as ≤10-point decline (responder) in at least three patients in a dosing cohort after 48 weeks. Other efficacy assessments included adaptive behavioral function, assessments of cortical gray matter volume, and glycosaminoglycan (GAG) levels in urine.

Results: A clinical response to rhHNS IT was observed in three treated patients (two in the Q2W group, one in the Q4W group). Cerebrospinal fluid heparan sulfate and urine GAG levels were reduced in all treated patients. However, most secondary efficacy assessments were similar between treated patients (n = 14; age, 17.8-47.8 months) and untreated controls (n = 7; age, 12.6-45.0 months). Treatment-emergent adverse events that occurred with rhHNS IT were mostly mild, none led to study discontinuation, and there were no deaths.

Conclusion: rhHNS IT treatment reduced heparan sulfate and GAG levels in treated patients. Though the primary neurocognitive endpoint was not met, important lessons in the design and endpoints for evaluation of cognitive and behavioral diseases resulted.

Trial Registration: ClinicalTrials.govNCT02060526; EudraCT 2013-003450-24.
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http://dx.doi.org/10.1016/j.ymgme.2018.10.006DOI Listing
February 2019

Hematopoietic Stem-Cell Gene Therapy for Cerebral Adrenoleukodystrophy.

N Engl J Med 2017 10 4;377(17):1630-1638. Epub 2017 Oct 4.

From Massachusetts General Hospital and Harvard Medical School (F.E., P.L.M.), Dana-Farber and Boston Children's Cancer and Blood Disorders Center (C. Duncan, M.A., C. Dansereau, D.A.W.), and Boston Children's Hospital, Harvard Medical School, and Harvard Stem-Cell Institute (D.A.W.), Boston, and Bluebird Bio, Cambridge (A.M.P., E.S., T.O., D.D.) - all in Massachusetts; University of Minnesota Children's Hospital, Minneapolis (P.J.O., T.C.L., W.P.M., G.V.R.); University of California, Los Angeles, Los Angeles (S.D.O., R.S., A.J.S.); University College London Great Ormond Street Hospital Institute of Child Health and Great Ormond Street Hospital NHS Trust, London (A.J.T., H.B.G., P.G.); Pediatric Neurology Department, Hôpital Bicêtre-Hôpitaux Universitaires Paris Sud, Le Kremlin Bicêtre, France (C.S., P.A.); Fundacion Investigar, Buenos Aires (H.A.); and Women's and Children's Hospital, North Adelaide, SA, Australia (D.B., N.J.C.S.).

Background: In X-linked adrenoleukodystrophy, mutations in ABCD1 lead to loss of function of the ALD protein. Cerebral adrenoleukodystrophy is characterized by demyelination and neurodegeneration. Disease progression, which leads to loss of neurologic function and death, can be halted only with allogeneic hematopoietic stem-cell transplantation.

Methods: We enrolled boys with cerebral adrenoleukodystrophy in a single-group, open-label, phase 2-3 safety and efficacy study. Patients were required to have early-stage disease and gadolinium enhancement on magnetic resonance imaging (MRI) at screening. The investigational therapy involved infusion of autologous CD34+ cells transduced with the elivaldogene tavalentivec (Lenti-D) lentiviral vector. In this interim analysis, patients were assessed for the occurrence of graft-versus-host disease, death, and major functional disabilities, as well as changes in neurologic function and in the extent of lesions on MRI. The primary end point was being alive and having no major functional disability at 24 months after infusion.

Results: A total of 17 boys received Lenti-D gene therapy. At the time of the interim analysis, the median follow-up was 29.4 months (range, 21.6 to 42.0). All the patients had gene-marked cells after engraftment, with no evidence of preferential integration near known oncogenes or clonal outgrowth. Measurable ALD protein was observed in all the patients. No treatment-related death or graft-versus-host disease had been reported; 15 of the 17 patients (88%) were alive and free of major functional disability, with minimal clinical symptoms. One patient, who had had rapid neurologic deterioration, had died from disease progression. Another patient, who had had evidence of disease progression on MRI, had withdrawn from the study to undergo allogeneic stem-cell transplantation and later died from transplantation-related complications.

Conclusions: Early results of this study suggest that Lenti-D gene therapy may be a safe and effective alternative to allogeneic stem-cell transplantation in boys with early-stage cerebral adrenoleukodystrophy. Additional follow-up is needed to fully assess the duration of response and long-term safety. (Funded by Bluebird Bio and others; STARBEAM ClinicalTrials.gov number, NCT01896102 ; ClinicalTrialsRegister.eu number, 2011-001953-10 .).
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http://dx.doi.org/10.1056/NEJMoa1700554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5708849PMC
October 2017

Intracerebral Gene Therapy Using AAVrh.10-hARSA Recombinant Vector to Treat Patients with Early-Onset Forms of Metachromatic Leukodystrophy: Preclinical Feasibility and Safety Assessments in Nonhuman Primates.

Hum Gene Ther Clin Dev 2015 Jun 28;26(2):113-24. Epub 2015 Apr 28.

1 Inserm U986, 94275 Le Kremlin Bicêtre , France .

No treatment is available for early-onset forms of metachromatic leukodystrophy (MLD), a lysosomal storage disease caused by autosomal recessive defect in arylsulfatase A (ARSA) gene causing severe demyelination in central and peripheral nervous systems. We have developed a gene therapy approach, based on intracerebral administration of AAVrh.10-hARSA vector, coding for human ARSA enzyme. We have previously demonstrated potency of this approach in MLD mice lacking ARSA expression. We describe herein the preclinical efficacy, safety, and biodistribution profile of intracerebral administration of AAVrh.10-hARSA to nonhuman primates (NHPs). NHPs received either the dose planned for patients adjusted to the brain volume ratio between child and NHP (1×dose, 1.1×10(11) vg/hemisphere, unilateral or bilateral injection) or 5-fold this dose (5×dose, 5.5×10(11) vg/hemisphere, bilateral injection). NHPs were subjected to clinical, biological, and brain imaging observations and were euthanized 7 or 90 days after injection. There was no toxicity based on clinical and biological parameters, nor treatment-related histological findings in peripheral organs. A neuroinflammatory process correlating with brain MRI T2 hypersignals was observed in the brain 90 days after administration of the 5×dose, but was absent or minimal after administration of the 1×dose. Antibody response to AAVrh.10 and hARSA was detected, without correlation with brain lesions. After injection of the 1×dose, AAVrh.10-hARSA vector was detected in a large part of the injected hemisphere, while ARSA activity exceeded the normal endogenous activity level by 14-31%. Consistently with other reports, vector genome was detected in off-target organs such as liver, spleen, lymph nodes, or blood, but not in gonads. Importantly, AAVrh.10-hARSA vector was no longer detectable in urine at day 7. Our data demonstrate requisite safe and effective profile for intracerebral AAVrh.10-hARSA delivery in NHPs, supporting its clinical use in children affected with MLD.
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http://dx.doi.org/10.1089/humc.2014.139DOI Listing
June 2015

Comparative efficacy and safety of multiple routes of direct CNS administration of adeno-associated virus gene transfer vector serotype rh.10 expressing the human arylsulfatase A cDNA to nonhuman primates.

Hum Gene Ther Clin Dev 2014 Sep 21;25(3):164-77. Epub 2014 Aug 21.

1 Department of Genetic Medicine, Weill Medical College of Cornell University , New York, NY 10065.

Metachromatic leukodystrophy (MLD), a fatal disorder caused by deficiency of the lysosomal enzyme arylsulfatase A (ARSA), is associated with an accumulation of sulfatides, causing widespread demyelination in both central and peripheral nervous systems. On the basis of prior studies demonstrating that adeno-associated virus AAVrh.10 can mediate widespread distribution in the CNS of a secreted lysosomal transgene, and as a prelude to human trials, we comparatively assessed the optimal CNS delivery route of an AAVrh.10 vector encoding human ARSA in a large animal model for broadest distribution of ARSA enzyme. Five routes were tested (each total dose, 1.5 × 10(12) genome copies of AAVrh.10hARSA-FLAG): (1) delivery to white matter centrum ovale; (2) deep gray matter delivery (putamen, thalamus, and caudate) plus overlying white matter; (3) convection-enhanced delivery to same deep gray matter locations; (4) lateral cerebral ventricle; and (5) intraarterial delivery with hyperosmotic mannitol to the middle cerebral artery. After 13 weeks, the distribution of ARSA activity subsequent to each of the three direct intraparenchymal administration routes was significantly higher than in phosphate-buffered saline-administered controls, but administration by the intraventricular and intraarterial routes failed to demonstrate measurable levels above controls. Immunohistochemical staining in the cortex, white matter, deep gray matter of the striatum, thalamus, choroid plexus, and spinal cord dorsal root ganglions confirmed these results. Of the five routes studied, administration to the white matter generated the broadest distribution of ARSA, with 80% of the brain displaying more than a therapeutic (10%) increase in ARSA activity above PBS controls. No significant toxicity was observed with any delivery route as measured by safety parameters, although some inflammatory changes were seen by histopathology. We conclude that AAVrh.10-mediated delivery of ARSA via CNS administration into the white matter is likely to be safe and yields the widest distribution of ARSA, making it the most suitable route of vector delivery.
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http://dx.doi.org/10.1089/humc.2013.239DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4227442PMC
September 2014

Correction of brain oligodendrocytes by AAVrh.10 intracerebral gene therapy in metachromatic leukodystrophy mice.

Hum Gene Ther 2012 Aug 23;23(8):903-14. Epub 2012 Jul 23.

INSERM U745, University Paris-Descartes, 75005, Paris, France.

Metachromatic leukodystrophy (MLD) is a lysosomal storage disorder characterized by accumulation of sulfatides in glial cells and neurons, the result of an inherited deficiency of arylsulfatase A (ARSA; EC 3.1.6.8) and myelin degeneration in the central and peripheral nervous systems. No effective treatment is currently available for the most frequent late infantile (LI) form of MLD, which results in rapid neurological degradation and early death after the onset of clinical manifestations. To potentially arrest or reverse disease progression, ARSA enzyme must be rapidly delivered to brain oligodendrocytes of patients with LI MLD. We previously showed that brain gene therapy with adeno-associated virus serotype 5 (AAV5) driving the expression of human ARSA cDNA under the control of the murine phosphoglycerate kinase (PGK) promoter alleviated most long-term disease manifestations in MLD mice. Herein, we evaluated the short-term effects of AAVrh.10 driving the expression of human ARSA cDNA under the control of the cytomegalovirus/β-actin hybrid (CAG/cu) promoter in 8-month-old MLD mice that already show marked sulfatide accumulation and brain pathology. Within 2 months, and in contrast to results with the AAV5-PGK-ARSA vector, a single intrastriatal injection of AAVrh.10cuARSA resulted in correction of brain sulfatide storage, accumulation of specific sulfatide species in oligodendrocytes, and associated brain pathology in the injected hemisphere. Better potency of the AAVrh.10cuARSA vector was mediated by higher neuronal and oligodendrocyte transduction, axonal transport of the AAVrh.10 vector and ARSA enzyme, as well as higher CAG/cu promoter driven expression of ARSA enzyme. These results strongly support the use of AAVrh.10cuARSA vector for intracerebral gene therapy in rapidly progressing early-onset forms of MLD.
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http://dx.doi.org/10.1089/hum.2012.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3413898PMC
August 2012

Autosomal recessive cerebellar ataxia caused by mutations in the PEX2 gene.

Orphanet J Rare Dis 2011 Mar 10;6. Epub 2011 Mar 10.

Pediatric Neurology and Endocrinology, Hôpital St Vincent de Paul, Paris, France.

Objective: To expand the spectrum of genetic causes of autosomal recessive cerebellar ataxia (ARCA).

Case Report: Two brothers are described who developed progressive cerebellar ataxia at 3 1/2 and 18 years, respectively. After ruling out known common genetic causes of ARCA, analysis of blood peroxisomal markers strongly suggested a peroxisomal biogenesis disorder. Sequencing of candidate PEX genes revealed a homozygous c.865_866insA mutation in the PEX2 gene leading to a frameshift 17 codons upstream of the stop codon. PEX gene mutations usually result in a severe neurological phenotype (Zellweger spectrum disorders).

Conclusions: Genetic screening of PEX2 and other PEX genes involved in peroxisomal biogenesis is warranted in children and adults with ARCA.
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http://dx.doi.org/10.1186/1750-1172-6-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3064617PMC
March 2011

Efficient intracerebral delivery of AAV5 vector encoding human ARSA in non-human primate.

Hum Mol Genet 2010 Jan;19(1):147-58

UMR INRA 703, Ecole Vétérinaire de Nantes, Nantes, France.

Metachromatic leukodystrophy (MLD) is a lethal neurodegenerative disease caused by a deficiency in the lysosomal arylsulfatase A (ARSA) enzyme leading to the accumulation of sulfatides in glial and neuronal cells. We previously demonstrated in ARSA-deficient mice that intracerebral injection of a serotype 5 adeno-associated vector (AAV) encoding human ARSA corrects the biochemical, neuropathological and behavioral abnormalities. However, before considering a potential clinical application, scaling-up issues should be addressed in large animals. Therefore, we performed intracerebral injection of the same AAV vector (total dose of 3.8 x 10(11) or 1.9 x 10(12) vector genome, three sites of injection in the right hemisphere, two deposits per site of injection) into three selected areas of the centrum semiovale white matter, or in the deep gray matter nuclei (caudate nucleus, putamen, thalamus) of six non-human primates to evaluate vector distribution, as well as expression and activity of human ARSA. The procedure was perfectly tolerated, without any adverse effect or change in neurobehavioral examination. AAV vector was detected in a brain volume of 12-15 cm(3) that corresponded to 37-46% of the injected hemisphere. ARSA enzyme was expressed in multiple interconnected brain areas over a distance of 22-33 mm. ARSA activity was increased by 12-38% in a brain volume that corresponded to 50-65% of injected hemisphere. These data provide substantial evidence for potential benefits of brain gene therapy in patients with MLD.
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http://dx.doi.org/10.1093/hmg/ddp475DOI Listing
January 2010

Intracerebral adeno-associated virus-mediated gene transfer in rapidly progressive forms of metachromatic leukodystrophy.

Hum Mol Genet 2006 Jan 25;15(1):53-64. Epub 2005 Nov 25.

Institut National de la Santé et de la Recherche Médicale U561, Université Paris V, 75014 Paris, France.

Metachromatic leukodystrophy (MLD) is a neurodegenerative lysosomal disease caused by a defect of the enzyme arylsulfatase A (ARSA) that disrupts the degradation of sulfatides (Sulf) in neurons and glial cells. Therapy for MLD requires active production of ARSA in the brain to prevent demyelination and neuronal damage, and efficient delivery of ARSA to act faster than disease progression, particularly in the rapidly progressive late infantile form. We used an adeno-associated virus serotype 5 (AAV5) vector to express the human ARSA gene in the brain of MLD mouse model. We achieved rapid, extensive and long-lasting expression of the recombinant ARSA in the brain, cerebellum and brainstem from at least 3 to 15 months post-injection. Analysis of the vector genome and ARSA distribution gave evidence for in vivo cross-correction of many untransduced neurons and astrocytes. ARSA delivery rapidly reversed Sulf storage and prevented neuropathological abnormalities and neuromotor impairment. We believe that AAV5-mediated brain delivery of ARSA is a potentially efficacious therapeutic strategy for MLD patients, especially for those with rapidly progressive form of the disease.
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http://dx.doi.org/10.1093/hmg/ddi425DOI Listing
January 2006

Intralysosomal cystine accumulation in mice lacking cystinosin, the protein defective in cystinosis.

Mol Cell Biol 2002 Nov;22(21):7622-32

INSERM U423, Hôpital Necker-Enfants Malades, Paris, France.

Cystinosis is an autosomal recessive disorder characterized by an accumulation of intralysosomal cystine. The causative gene, CTNS, encodes cystinosin, a seven-transmembrane-domain protein, which we recently showed to be a lysosomal cystine transporter. The most severe and frequent form of cystinosis, the infantile form, appears around 6 to 12 months, with a proximal tubulopathy (de Toni-Debré-Fanconi syndrome) and ocular damage. End-stage renal failure is reached by 10 years of age. Accumulation of cystine in all tissues eventually leads to multisystemic disease. Treatment with cysteamine, which reduces the concentration of intracellular cystine, delays disease progression but has undesirable side effects. We report the first Ctns knockout mouse model generated using a promoter trap approach. We replaced the last four Ctns exons by an internal ribosome entry site-betagal-neo cassette and showed that the truncated protein was mislocalized and nonfunctional. Ctns(-/-) mice accumulated cystine in all organs tested, and cystine crystals, pathognomonic of cystinosis, were observed. Ctns(-/-) mice developed ocular changes similar to those observed in affected individuals, bone defects and behavioral anomalies. Interestingly, Ctns(-/-) mice did not develop signs of a proximal tubulopathy, or renal failure. A preliminary therapeutic trial using an oral administration of cysteamine was carried out and demonstrated the efficiency of this treatment for cystine clearance in Ctns(-/-) mice. This animal model will prove an invaluable and unique tool for testing emerging therapeutics for cystinosis.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC135682PMC
http://dx.doi.org/10.1128/mcb.22.21.7622-7632.2002DOI Listing
November 2002

Heterogeneity of persistent hyperinsulinaemic hypoglycaemia. A series of 175 cases.

Eur J Pediatr 2002 Jan;161(1):37-48

Federation de Pédiatrie, Hôpital Necker-Enfants-Malades, Paris, France.

Unlabelled: Hyperinsulinism is a heterogeneous disorder characterised by severe hypoglycaemia due to an inappropriate oversecretion of insulin. In a personal series of 175 patients investigated for hyperinsulinaemic hypoglycaemia over the last 20 years, we review clinical presentations, molecular studies and therapeutic management of hyperinsulinism. There were 98 neonatal-onset patients, including 86 permanent hyperinsulinism and 12 transient forms, 68 with infancy-onset and nine with childhood-onset. Hyperammonaemia was found in 12 out of 69 patients tested, 4 neonates and 8 infants. Neonates were clinically more severely affected than infants. Diagnosis of infancy-onset hyperinsulinism was often delayed because of less profound hypoglycaemia and better tolerance to hypoglycaemia. Neonates required higher rates of i.v. glucose than infants to maintain normal plasma glucose levels (16 mg/kg per min versus 12 mg/kg per min). Only 16% of neonates were diazoxide-sensitive compared to 66% of the infants. Neonates with hyperammonaemia or transient hyperinsulinism were diazoxide-sensitive. Most neonates were pancreatectomised whereas 65% of the infants were treated medically. Among surgically-treated patients, 47% had a focal adenomatous hyperplasia (31 neonates and 13 infants) and 53% a diffuse form of hyperinsulinism (39 neonates and 11 infants). Diazoxide-responsiveness in the focal and diffuse forms did not differ in both neonates and infants; it depended only upon the age of onset of hypoglycaemia. One or two mutations, SUR1 or KIR6.2, were found in 41 of 73 neonates who were investigated and in 13/38 infants using polymerase chain reaction-single strand conformational polymorphism analysis of both genes. Almost all patients with SUR1 (38/41) or KIR6.2 (5/7) mutations were resistant to diazoxide. Ten patients with hyperinsulinism-hyperammonaemia syndrome had a mutation in the glutamate dehydrogenase gene (three neonates and seven infants) after reverse transcriptase-polymerase chain reaction and sequence analysis of cDNA. No mutation was found by polymerase chain reaction-single strand conformational polymorphism in the glucokinase gene. Eight of nine patients with childhood onset hyperinsulinism were treated surgically and histological examination confirmed an adenoma in each case.

Conclusion: the clinical severity of hyperinsulinism varies mainly with age at onset of hypoglycaemia. The heterogeneity of hyperinsulinism has major consequences in terms of therapeutic outcome and genetic counselling.
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http://dx.doi.org/10.1007/s004310100847DOI Listing
January 2002