Andrea M. Atherton

Diagnosis of mitochondrial disorders by concomitant next-generation sequencing of the exome and mitochondrial genome.

Mitochondrial diseases are notoriously difficult to diagnose due to extreme locus and allelic heterogeneity, with both nuclear and mitochondrial genomes potentially liable. Using exome sequencing we demonstrate the ability to rapidly and cost effectively evaluate both the nuclear and mitochondrial genomes to obtain a molecular diagnosis for four patients with three distinct mitochondrial disorders. One patient was found to have Leigh syndrome due to a mutation in MT-ATP6, two affected siblings were discovered to be compound heterozygous for mutations in the NDUFV1 gene, which causes mitochondrial complex I deficiency, and one patient was found to have coenzyme Q10 deficiency due to compound heterozygous mutations in COQ2. In all cases conventional diagnostic testing failed to identify a molecular diagnosis. We suggest that additional studies should be conducted to evaluate exome sequencing as a primary diagnostic test for mitochondrial diseases, including those due to mtDNA mutations.

Fabry disease in infancy and early childhood: a systematic literature review

Purpose:Fabry disease is a pan-ethnic, progressive, X-linked genetic disorder that commonly presents in childhood and is caused by deficient activity of the lysosomal enzyme alpha-galactosidaseA (α-gal A). Symptoms of Fabry disease in the pediatric population are well described for patients over five years of age; however, data are limited for infancy and early childhood. The purpose of this article is to delineate the age of detection for specific Fabry symptoms in early childhood.Methods:A systematic retrospective analysis of PubMed indexed, peer-reviewed publications and case reports in the pediatric Fabry population was performed to review symptoms in patients reported before 5 years of age.Results:The most frequently reported symptom in all age groups under 5 years was acroparesthesias/neuropathic pain, reported in 9 children, ranging in age from 2.0-4.0 years. Also notable is the frequency of gastrointestinal issues reported in 6 children aged 1.0-4.1 years of age.Conclusion:This article finds clear evidence that symptoms can occur in early childhood, before age 5 years. Given early presenting symptoms and the ability to monitor these disease hallmarks, a timely referral to a medical geneticist or other specialty clinician experienced in managing children with Fabry disease is strongly indicated.Genet Med 17 5, 323–330.

Enzyme Replacement Therapy in Mucopolysaccharidosis II Patients Under 1 Year of Age

Mucopolysaccharidosis (MPS) II, or Hunter syndrome, is a lysosomal storage disease characterized by multi-systemic involvement and a progressive clinical course. Enzyme replacement therapy with idursulfase has been approved in more than 50 countries worldwide; however, safety and efficacy data from clinical studies are currently only available for patients 1.4 years of age and older. Sibling case studies of infants with MPS I, II, and VI who initiated ERT in the first weeks or months of life have reported no new safety concerns and a more favorable clinical course for the sibling treated in infancy than for the later-treated sibling. Here we describe our experiences with a case series of eight MPS II patients for whom idursulfase treatment was initiated at under 1 year of age. The majority of the patients were diagnosed because of a family history of disease. All of the infants displayed abnormalities consistent with MPS II at diagnosis. The youngest age at treatment start was 10 days and the oldest was 6.5 months, with duration of treatment varying between 6 weeks and 5.5 years. No new safety concerns were observed, and none of the patients experienced an infusion-related reaction. All of the patients treated for more than 6 weeks showed improvements and/or stabilization of some somatic manifestations while on treatment. In some cases, caregivers made comparisons with other affected family members and reported that the early-treated patients experienced a less severe clinical course, although a lack of medical records for many family members precluded a rigorous comparison.

exome sequencing Reveals De novo Germline Mutation of the Mammalian Target of Rapamycin (MTOR) in a patient with Megalencephaly and Intractable seizures

A de novo somatic mutation in the mammalian target of rapamycin (MTOR) has previously been described in one patient with hemimegalencephaly and epilepsy. Here, we present a case of a young girl with megalencephaly and intractable seizures who was found to have an MTOR mutation in multiple cell lineages (p.Cys1483Phe) and, therefore, presumed to be of germline origin. The mutation was detected in peripheral blood DNA by exome sequencing of the patient and her parents, substantiating the utility of this approach for detection of clinically relevant de novo variations.

CLPB variants associated with autosomal-recessive mitochondrial disorder with cataract, neutropenia, epilepsy, and methylglutaconic aciduria.

3-methylglutaconic aciduria (3-MGA-uria) is a nonspecific finding associated with mitochondrial dysfunction, including defects of oxidative phosphorylation. 3-MGA-uria is classified into five groups, of which one, type IV, is genetically heterogeneous. Here we report five children with a form of type IV 3-MGA-uria characterized by cataracts, severe psychomotor regression during febrile episodes, epilepsy, neutropenia with frequent infections, and death in early childhood. Four of the individuals were of Greenlandic descent, and one was North American, of Northern European and Asian descent. Through a combination of homozygosity mapping in the Greenlandic individuals and exome sequencing in the North American, we identified biallelic variants in the caseinolytic peptidase B homolog (CLPB). The causative variants included one missense variant, c.803C>T (p.Thr268Met), and two nonsense variants, c.961A>T (p.Lys321*) and c.1249C>T (p.Arg417*). The level of CLPB protein was markedly decreased in fibroblasts and liver of affected individuals. CLPB is proposed to function as a mitochondrial chaperone involved in disaggregation of misfolded proteins, resulting from stress such as heat denaturation.

Mucopolysaccharidosis Type I Newborn Screening: Best Practices for Diagnosis and Management

The mucopolysaccharidoses (MPS) are a group of rare progressive genetic disorders of glycosaminoglycan (GAG) metabolism caused by deficiency of enzymes responsible for lysosomal GAG degradation. The accumulation of partially degraded GAG and the resulting disturbance of cellular homeostasis leads to progressive cellular and tissue damage ultimately resulting inmultiorgan system involvement. Mucopolysaccharidosis type 1 (MPS I) results from deficiency of the lysosomal enzyme a-L-iduronidase (IDUA) because of pathogenic variants in the IDUA gene. MPS I presents clinically as a disease spectrum spanning early onset, progressive severe disease with cognitive impairment (Hurler syndrome), to later onset progressive disease with highly variable and later onset central nervous system (CNS) involvement (attenuated MPS I). Attenuated MPS I encompasses a spectrum previously referred to by the eponyms Hurler-Scheie and Scheie syndromes. Untreated patients with the most severe form of MPS I usually die in the first decade. In contrast, life expectancy for untreated patients with attenuated disease ranges from mortality in the second or third decade to full life expectancy, albeit with considerable morbidity. The birth incidence of MPS I is ~1 in 100 000 live births estimated from a number of population studies. Although the disease pathophysiology is not well understood, most of the clinical manifestations of MPS I are secondary deleterious effects because of disturbed GAG metabolism. Organs involved include the brain, musculoskeletal system, heart, lungs, and eyes. Many symptoms and complications are difficult or impossible to reverse. Thus, initiation of treatment early in the natural history of disease is thought to be a key factor in achieving optimal outcome. Available disease modifying therapies include hematopoietic stem cell transplantation (HSCT) and enzyme replacement therapy (ERT) with laronidase. Because early intervention with HSCT has been demonstrated to stabilize neurocognitive function in MPS I, it is currently recommended as standard for patients who are predicted to have severe disease. ERT with laronidase is used for treating nondirect CNS manifestations of MPS I. Clinical trials and follow-up studies of ERT in MPS I have demonstrated improvements in some somatic manifestations and functional outcomes in patients with attenuated MPS I. Laronidase has also been found to be useful in the peritransplant period for patients with severe MPS I. The use of ERT in the peritransplant period has been shown to be safe and has led to clinical improvements particularly in patients with significant cardiopulmonary disease before transplantation. In addition, ERT before transplantation has been reported to alleviate symptoms that may have influenced the conditioning regime or candidacy for transplantation of some patients. Because MPS I is a progressive disorder, the success of both HSCTandERTdepends on early initiation of treatment.Therefore, early identification of patients is critical. Because many of the early disease manifestations represent common childhood symptoms (eg, inguinal/umbilical hernia and recurrent upper respiratory tract infections), diagnosis based on early symptom recognition is challenging and has been met with limited success. Newborn screening (NBS) strategies should be more effective in this regard.MPS I NBS via determination of IDUA activity in dried blood spot (DBS)-derived samples is currently underway in the US and in pilot programs in Taiwan, Italy, Austria, andHungary. TheUSDepartment of Health andHuman Services recommended uniform screening panel provides the list of core and secondary disorders that should be included in every NBS program.A proposal for

A Systematic Approach to Implementing Monogenic Genomic Medicine: Genotype-Driven Diagnosis of Genetic Diseases

Genomic medicine is an emerging paradigm for disease diagnosis and management that incorporates individual genome sequence information based on and identified by next-generation sequencing. Here we report on the initial experience in implementing genomic medicine for inherited diseases in a large children’s hospital. In two families, next-generation sequencing identified molecular diagnoses that had not been disclosed by years of traditional diagnostic tests. Two sisters with progressive ataxia were found to have a mutation in aprataxin gene (APTX c.717G > A, p.Trp239X) and were treated with oral Coenzyme Q10. Two brothers with intellectual disability, dysmorphic features, doughy skin, and truncal obesity were found to have autosomal recessive cutis laxa caused by mutations in pyrroline-5-carboxylate reductase, type 1 (PYCR1 c.120_121delCA). Pediatric genomic medicine appears to enable early diagnosis of inherited diseases that feature clinical or genetic heterogeneity and it may allow for targeted treatment. We discuss several bottlenecks to improving care though genomic medicine, as well as potential solutions.

Maternal serum screening and 22q11.2 deletion syndrome

Second trimester maternal serum screening (MSS) is performed by measuring concentrations of alpha fetoprotein (AFP), unconjugated estriol (uE3), beta human gonadotropin (bhCG) and inhibin-A. This technology is commonly used to identify pregnancies at increased risk for trisomy 21, trisomy 18, open neural tube defects and most recently, steroid sulfatase deficiency ichthyosis, and the Smith–Lemli–Opitz Syndrome (Table I). We report here on, two children with the 22q11.2 deletion syndrome whose mothers both had MSS results consistent with an increased risk for trisomy 18. The first patient was a female infant transferred to our hospital for evaluation and management of multiple congenital anomalies. The child was born to a 24-year-old G II P II who had MSS with a 1 in 23 risk that the infant had trisomy 18. Following a normal level II ultrasound exam amniocentesis was declined. The child was born at 36 weeks gestation at less than the 10th centile for height, weight, and head circumference, with the following anomalies: overriding sutures, sclerocorneas, microphthalmia with colobomas, posteriorly rotated ears, bulbous nasal tip, imperforate anus, long fingers and postaxial polydactyly of the right hand and right foot. FISH probes documented the 22q11.2 deletion. The second childwasborn at 29weeks gestation to a gravida V para IV SAB I, 36-year-old woman who had MSS with a 1 in 40 risk of trisomy 18. An ultrasound at 20 weeks gestation was normal. A follow-up ultrasound 4 weeks later identified a congenital heart defect and an amniocentesis was done with normal results (46,XY). At birth he presented with an interrupted aortic arch type B, VSD, hypospadias, posteriorly rotated ears, and asymmetric crying facies. 22q11.2 FISH probe established the diagnosis. Approximately 1 in 200 women who have MSS will be positive for trisomy 18 [Yankowitz et al., 1998]. The incidence of 22q11.2 deletion has been reported to be between 1/4,000 [Wilson et al., 1994] and 1/ 6,395 [Devriendt et al., 1998]. If MSS for trisomy 18 was not also identifying fetuses at risk for 22q11.2 deletion syndrome, then the expected incidence of these two independent events occurring together would be between 1/800,000 and 1/1,279,000 screening tests. Given the small probability that these events would occur by chance, we suggest that MSS positive for trisomy 18 may also be identifying fetuses at risk for the 22q11.2 deletion. We therefore suggest that all pregnancies with

HSP and deafness: Neurocristopathy caused by a novel mosaic SOX10 mutation.

Objective: To identify the underlying genetic cause in 2 sisters affected with progressive lower extremity spasticity, neuropathy, and early-onset deafness. Methods: Whole-exome sequencing was performed, and segregation testing of variants was investigated using targeted Sanger sequencing. An inherited paternal mosaic mutation was further evaluated through quantitative analysis of the ratio of mutant vs wild-type allele in genomic DNA from various tissues, including blood, dermal fibroblasts, and saliva. Results: A novel heterozygous nonsense mutation (c.1140C>A; p.Y380X) in SOX10 was identified in the affected sisters. Paternal mosaicism was suspected based on a small chromatogram peak, which was less than the heterozygous peak of the mutated allele. Consistent with mosaicism, the mosaic paternal samples had notable variability in the ratio of mutant vs wild-type allele in various tissues (compared with the fully heterozygous daughter), with the highest paternal mutant levels in saliva (32.7%) and lowest in dermal fibroblasts (13.9%). Targeted clinical re-examination of the father revealed a sensorimotor neuropathy that was previously clinically unrecognized. Conclusions: These findings expand the phenotypic spectrum of SOX10-related neurocristopathy. Mutations in SOX10 should be considered in patients presenting with a complicated form of hereditary spastic paraplegia that includes neuropathy and deafness. Diagnostic workup may be complicated, as SOX10 mutations can present in a mosaic state, with a mild clinical manifestation.

Ethical Considerations When Including Lysosomal Storage Disorders in Newborn Screening Programs

There is essentially unanimous agreement that newborn screening saves lives. Nevertheless, newborn screening—rated as one of the top 10 successes in public health in the first decade of the twenty-first century by the Centers for Disease Control and Prevention (Koppaka in JAMA 306(5):484–487, 2003)—faces multiple challenges, including questions about how conditions are added to the list of disorders for which screening is done in each state, about the need for parental consent for use of dried blood spots after screening, and the roles of parents in expansion of newborn screening. Such questions figure prominently in controversy surrounding Pennsylvania’s recent passage of House Bill1654, also called Hannah’s Law, which institutes newborn screening and follow-up for 6 lysosomal storage disorders including Krabbe, Fabry, Pompe, Niemann–Pick, types A and B, Gaucher diseases, and mucopolysaccharidosis type I, also called Hurler disease. This review article provides an historical perspective on newborn screening including discussion of how conditions came to be added to the Recommended Uniform Screening Panel, and an overview of current issues and concerns for key stakeholders including parents, healthcare providers, laboratorians, and legislators.