T. Andrew Burrow

A Phase 3 Trial of Sebelipase Alfa in Lysosomal Acid Lipase Deficiency

BACKGROUND Lysosomal acid lipase is an essential lipid-metabolizing enzyme that breaks down endocytosed lipid particles and regulates lipid metabolism. We conducted a phase 3 trial of enzyme-replacement therapy in children and adults with lysosomal acid lipase deficiency, an underappreciated cause of cirrhosis and severe dyslipidemia. METHODS In this multicenter, randomized, double-blind, placebo-controlled study involving 66 patients, we evaluated the safety and effectiveness of enzyme-replacement therapy with sebelipase alfa (administered intravenously at a dose of 1 mg per kilogram of body weight every other week); the placebo-controlled phase of the study was 20 weeks long and was followed by open-label treatment for all patients. The primary end point was normalization of the alanine aminotransferase level. Secondary end points included additional disease-related efficacy assessments, safety, and side-effect profile. RESULTS Substantial disease burden at baseline included a very high level of low-density lipoprotein cholesterol (≥190 mg per deciliter) in 38 of 66 patients (58%) and cirrhosis in 10 of 32 patients (31%) who underwent biopsy. A total of 65 of the 66 patients who underwent randomization completed the double-blind portion of the trial and continued with open-label treatment. At 20 weeks, the alanine aminotransferase level was normal in 11 of 36 patients (31%) in the sebelipase alfa group and in 2 of 30 (7%) in the placebo group (P=0.03), with mean changes from baseline of -58 U per liter versus -7 U per liter (P<0.001). With respect to prespecified key secondary efficacy end points, we observed improvements in lipid levels and reduction in hepatic fat content (P<0.001 for all comparisons, except P=0.04 for triglycerides). The number of patients with adverse events was similar in the two groups; most events were mild and were considered by the investigator to be unrelated to treatment. CONCLUSIONS Sebelipase alfa therapy resulted in a reduction in multiple disease-related hepatic and lipid abnormalities in children and adults with lysosomal acid lipase deficiency. (Funded by Synageva BioPharma and others; ARISE ClinicalTrials.gov number, NCT01757184.).

Characterization of congenital anomalies in individuals with choanal atresia.

OBJECTIVE To review a tertiary care pediatric hospitals experience with choanal atresia and stenosis (CA/S) related to associated congenital anomalies (birth defects, including minor abnormalities) and genetic disorders. DESIGN Retrospective case series. SETTING Tertiary care pediatric hospital. PATIENTS Individuals with CA/S. MAIN OUTCOME MEASURES Identification of congenital anomalies, neurologic abnormalities, and developmental disabilities in individuals with CA/S. RESULTS One hundred twenty-nine individuals with CA/S were evaluated between July 1, 1997, and July 1, 2007. Choanal atresia and stenosis was an isolated finding in 34 patients (26.4%) and was associated with other anomalies in 95 patients (73.6%). Specific conditions were diagnosed in 66 patients (51.2%); CHARGE (coloboma, heart defect, atresia choanae, retarded growth, genitourinary abnormalities, and ear anomalies) syndrome was the most common diagnosis (33 patients [25.6%]). Numerous conditions were seen, including chromosomal abnormalities, single-gene defects, deformations, and those caused by teratogens. Choanal atresia and stenosis was unilateral in 62 patients (48.1%) and was bilateral in 60 patients (46.5%). Unilateral cases were more likely to be isolated (30 patients [53.2%]). Bilateral cases were more likely to be associated with specific disorders or multiple congenital anomalies (60 patients [98.4%]). There was no difference in laterality among unilateral cases. CONCLUSIONS Choanal atresia and stenosis is associated with a wide range of disorders. Congenital anomalies, neurologic abnormalities, and developmental disabilities are commonly identified in affected individuals. Bilateral CA/S is more commonly seen in patients in whom specific diagnoses or other congenital anomalies are identified. Unilateral CA/S occurs more frequently in isolated cases. A comprehensive evaluation is recommended in individuals with CA/S to evaluate for other congenital anomalies, neurologic abnormalities, developmental delays, and evidence of a specific underlying disorder.

Enzyme reconstitution/replacement therapy for lysosomal storage diseases.

Purpose of review Over the past 15 years, the lysosomal storage diseases have become paradigms for the specific treatment of monogenic disorders, particularly those affecting children. This review summarizes the phenotypes and recent literature regarding enzyme reconstitution (replacement) therapy and outcomes for such treatable lysosomal storage diseases: Gaucher disease, Fabry disease, Pompe disease and the mucopolysaccharidoses. Recent findings Recent clinical trials have shown that enzyme reconstitution therapy effectively treats many of the manifestations of the lysosomal storage diseases. When initiated early in the disease course, enzyme reconstitution therapy can reverse some disease manifestations, but may not completely alleviate the disease progression. Enzyme reconstitution therapy is generally well tolerated. Many adverse events are antibody-related, but can be managed without requiring cessation of enzyme reconstitution therapy. Documented IgE reactions, i.e. anaphylactoid, are quite rare (fewer than 1%). Summary Enzyme reconstitution therapy is a safe and effective treatment modality available for several of the lysosomal storage diseases. Owing to the short history of enzyme reconstitution therapy, the long-term outcomes of enzyme reconstitution therapy-treated individuals are unknown and require further investigation. Medical professionals must learn to identify patients likely to benefit from these life-changing therapies so as to prevent many of the devastating, irreversible complications of the lysosomal storage diseases.

Acute Progression of Neuromuscular Findings in Infantile Pompe Disease

A 2-year-old girl with Pompe disease developed an acute worsening of muscle weakness during a hospitalization, and required intubation for an upper respiratory infection. Electromyography and nerve conduction studies produced results consistent with a severe chronic motor axonal peripheral polyneuropathy, with no evidence of reinnervation. Magnetic resonance imaging of the brain demonstrated generalized hypomyelination and parenchymal volume loss, whereas magnetic resonance spectroscopy suggested neuronal injury and hypomyelination. This case provides compelling evidence for a slowly progressive neurodegenerative process in patients with infantile Pompe disease, affecting the motor neurons. Routine electromyography, nerve conduction studies, and cranial magnetic resonance imaging should be considered to delineate the presence of a neurodegenerative process in infantile-onset Pompe disease.

Somatic HRAS p.G12S mutation causes woolly hair and epidermal nevi.

TO THE EDITOR Woolly hair nevus (WHN) is a mosaic disorder characterized by distinct patterns of tightly curled scalp hair which can appear concurrently with epidermal nevi (EN) at other sites (Peteiro et al., 1989; Venugopal et al., 2012). Woolly hair is also found in congenital disorders resulting from mutations affecting diverse cellular components including intermediate filament, adherens junction, and signal transduction proteins (Harel and Christiano, 2012). Embryonic somatic mutation causes mosaic disorders which appear in patterns of ectodermal progenitor dorsovental migration. Somatic mutations causing mosaic disorders including Proteus syndrome (Lindhurst et al., 2011), port-wine stains (Shirley et al., 2013), and EN (Levinsohn et al., 2013; Sun et al., 2013) have been found using exome sequencing. Recognizing that exome sequencing would permit identification of mutations causing WHN, we ascertained two cases. Our first (WHN100, Figure 1a-d) was a 10 year-old girl without history of developmental delay who had regions of slightly curly hair over her occipital scalp from infancy which progressively curled with no scalp surface change and lie alongside areas of straight hair. She has hyperpigmented patches on her neck, trunk, and arms, with more keratotic lesions on her distal extremities, and acanthosis nigricans in both axillae. There was linear palmar keratoderma (PPK) and hyperkeratosis over most metacarpophalangeal and some proximal interphalangeal joints. Given concurrent PPK and woolly hair, clinical concern for Naxos or Carvajal syndromes led to regular cardiology evaluations that found no abnormalities. Figure 1 Clinical features of index cases with woolly hair nevi. On the scalp, woolly hair nevus presents with a portion of the scalp exhibiting patches of curly, thin, hair intermixed with regions of normal, straight hair, as observed in WHN100 and WHN101. On ... Our second case (WHN101, Figure 1e-h) was a 6 year-old girl whose hair developed at age one and consisted of a mixture of poker-straight hair and curly, thin hair. In infancy, she developed linear dyspigmentation on the right arm and trunk, which became more raised and scaly on the distal extremities over time. She had normal development, with no cardiac or ophthalmic abnormalities found on routine physical examination, cardiac MRI and serial electrocardiograms. Clinical suspicion of mosaic Naxos or Caravajal syndrome motivated clinical sequencing of DSP, DSC1, DSG1, JUP, PKP2, and TMEM43; no mutations were found. To determine the genetic basis of WHN, we performed paired whole exome sequencing of DNA isolated from affected tissue and blood in both cases (Supplementary Figure 2). Data was analyzed to identify somatic single nucleotide variants (SNVs), deletions and insertions (Supplementary Methods). A somatic heterozygous HRAS c.34G>A, p.G12S substitution was found in each (Figure 2a). There was no evidence of loss of heterozygosity (LOH) (Supplementary Figure 3) or secondary mutation somatic mutation, suggesting that HRAS mutation alone is sufficient to cause WHN. Sanger sequencing confirmed mutation presence in affected tissue (Figure 2b, c). To determine if this mutation causes woolly hair, we prepared DNA from hair bulbs of straight and curly hair obtained from affected individual WHN101, finding the HRAS p.G12S mutation in curly hair only (Figure 2d, Supplementary Figure 1). Figure 2 Somatic HRAS p.G12S mutation causes WHN. (a) In WHN100 and WHN101, exome sequencing of affected tissue and blood was performed. Tissue-specific SNVs are annotated bychromosome, position, base change, protein consequence, and numbers of reference and non-reference ... Consistent with somatic mosaicism in an epidermal progenitor, prior cases of WHN have been reported with concurrent keratinocytic epidermal nevi (KEN). KEN result from somatic mutations in HRAS, KRAS, PIK3CA, FGFR3, and NRAS (Hafner et al., 2012) including the HRAS p.G12S mutation found in WHN (Hafner et al., 2011). Furthermore, Costello syndrome (CS), in which patients present with developmental delay, high birth weight, feeding difficulties, failure to thrive, cardiac anomalies, and curly hair, results from germline heterozygous HRAS mutations, including p.G12S (Gripp and Lin, 2012; Siegel et al., 2012). The timing of somatic mutation during embryonic development determines extent of cutaneous involvement and presence of other systemic abnormalities (Moss et al., 1993). Notably, somatic activating HRAS mutations are found in most cases of nevus sebaceus (NS), a mosaic lesion which typically appears on the scalp and features alopecia, papillomatosis, and marked sebaceus hyperplasia (Groesser et al., 2012; Levinsohn et al., 2013; Sun et al., 2013). These features contrast with those of WHN in which hair is present but curly, and sebaceous hyperplasia is absent. Given that WHN and NS are both caused by somatic HRAS mutations, we hypothesize that their phenotypic divergence may derive from relative potency of the mutant allele with respect to MAP kinase activation. HRAS mutations in WHN and NS fall within the finger loop of HRAS, replacing glycine residues with larger amino acids which prevent GTP hydrolysis (Malumbres and Barbacid, 2003). Though comparison of the WHN p.G12S mutation and the common NS p.G13R mutation has not been performed, HRAS codon 12 serine substitutions have been shown to be less activating than arginine, aspartic acid or valine substitutions (Fasano et al., 1984). To evaluate the frequency of HRAS mutation in NS, we screened 116 archival scalp NS lesions for HRAS and KRAS mutation. We found 88 HRAS and 9 KRAS mutations. HRAS p.G13R was present in 85 NS and p.G12S was not found (Supplementary Table 2). In prior reports, 64 additional samples were screened, and HRAS p.G12S mutations were not found (Levinsohn et al., 2013; Sun et al., 2013). In one report, 3 specimens with HRAS p.G12S mutations were identified; in 2 there was a concurrent HRAS p.G13R mutation, and in one, the lesion was on the ear, a site at which it could be difficult to distinguish EN and NS (Groesser et al., 2012). These data combined with evidence from CS suggest that more strongly activating RAS mutations may cause the alopecia and sebaceous hyperplasia found in NS, and the more mildly activating p.G12S mutation causes woolly hair phenotypes. In summary, we find somatic HRAS c.34G>A, p.G12S mutation in affected tissue from two cases with mosaic woolly hair and EN. Consistent with reports of WHN and in KEN, the identified p.G12S mutation causes an EN phenotype on the body, but the finding of curly hair on the scalp suggests that WHN represents a mosaic RASopathy with phenotype determined by location, either due to distinct epidermal progenitor types or site-specific mesenchymal interactions. We hypothesize that in contrast to strongly activating RAS mutations found in NS which drive hair follicle progenitors toward sebocyte differentiation, the more weakly activating mutation found in WHN permits an intermediate phenotype with abnormal curly hair growth but without sebaceous hyperplasia.

Cardiac Disease in Methylmalonic Acidemia

Methylmalonic acidemia (MMA) is a heterogeneous disorder, with onset from infancy to adulthood and varying degrees of organ involvement and severity. Cardiac disease is a known lethal complication of other organic acidemias, but has not been associated with MMA. We identified 3 patients with MMA and cardiac disease.

Non‐lethal congenital hypotonia due to glycogen storage disease type IV

Glycogen storage disease type IV (GSD‐IV) is an autosomal recessive genetic disorder due to a deficiency in the activity of the glycogen branching enzyme (GBE). A deficiency in GBE activity results in the accumulation of glycogen with fewer branching points and long, unbranched outer chains. The disorder results in a variable phenotype, including musculoskeletal, cardiac, neurological, and hepatic involvement, alone or in continuum, which can be identified at any stage of life. The classic form of GSD‐IV is a hepatic presentation, which presents in the first 18 months of life with failure to thrive, hepatomegaly, and cirrhosis that progresses to liver failure, resulting in death by age 5 years. A severe congenital musculoskeletal phenotype with death in the neonatal period has also been described. We report an unusual case of congenital musculoskeletal presentation of GSD‐IV with stable congenital hypotonia, gross motor delay, and severe fibro‐fatty replacement of the musculature, but no hepatic or cardiac involvement. Molecular analysis revealed two novel missense mutations with amino acid changes in the GBE gene (Q236H and R262C), which may account for the mild phenotype.

Prevalence and management of Gaucher disease

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Stability is maintained in adults with Gaucher disease type 1 switched from velaglucerase alfa to eliglustat or imiglucerase: A sub-analysis of the eliglustat ENCORE trial

Gaucher disease type 1 is an autosomal recessive disorder caused by deficient activity of the lysosomal enzyme acid β-glucosidase resulting in accumulation of glucosylceramide and clinical manifestations of anemia, thrombocytopenia, hepatosplenomegaly, and skeletal disease. The historic standard of care is intravenous recombinant enzyme therapy with imiglucerase. Eliglustat, an oral substrate reduction therapy, is a first-line treatment for adults with Gaucher disease type 1 who have a compatible CYP2D6-metabolizer phenotype (≈ 95% of patients). The 12-month ENCORE trial (NCT00943111) found eliglustat non-inferior to imiglucerase in maintaining stability in adult Gaucher patients previously stabilized after ≥ 3 years of enzyme therapy (imiglucerase or velaglucerase alfa). This post-hoc analysis examined safety and efficacy in the 30 ENCORE patients who were receiving velaglucerase alfa at study entry and were randomized to eliglustat (n = 22) or imiglucerase (n = 8). Efficacy and safety in velaglucerase alfa-transitioned patients were consistent with the full ENCORE trial population; 90% of patients switched to eliglustat and 88% of patients switched to imiglucerase met the composite endpoint (stable hemoglobin concentration, platelet count, spleen volume, and liver volume). Clinical stability was maintained for 12 months in Gaucher disease type 1 patients in the ENCORE trial who switched from velaglucerase alfa to either eliglustat or imiglucerase.

Correlating liver stiffness with disease severity scoring system (DS3) values in Gaucher disease type 1 (GD1) patients

Gaucher disease (GD) is an autosomal-recessive lysosomal storage disease caused by a deficiency of the enzyme, glucocerebrocidase, resulting in accumulation of lipid-laden storage cells in multiple organs such as bone marrow, liver, spleen, and lungs. Type 1 Gaucher disease is the most common form of this condition in which the brain and spinal cord (the central nervous system) are not affected. The Gaucher disease severity scoring system (GD-DS3) is typically used to assess disease severity accounting for skeletal, hematologic, and visceral disease. In addition to being time consuming for the clinician to calculate the scores, some of the assessments are subjective and may falsely increase or decrease disease severity. The purpose of this study was to determine if there is a correlation between liver stiffness values obtained from MR elastography (MRE) and the GD-DS3 score. An IRB approved, HIPAA compliant retrospective study was performed. All patients with type 1 GD imaged with MRE between 2011 and 2016 were included in this study. Clinical and imaging data was collected. Two pediatric radiologists analyzed MR images from abdomen and thigh studies independently to determine bone marrow involvement using a semi-quantitative scoring system with one reviewer analyzing a subset of studies to determine inter-observer reliability. The collected data was used to calculate a GD-DS3 score for all patients. GD-DS3 scores were compared with liver MRE stiffness values. Clinical MRE scores were plotted against GD-DS3 severity scores for 31 patients (15 males, 16 females; median age 27years, age range: 4-67years). The median GD-DS3 score was 4 (range: 1-10.1) and median MRE value was 2.43kPa (range: 1.30-5.20kPa). A significant positive correlation was found between MRE and GD-DS3 scores; Pearsons correlation coefficient value of r=0.47, p<0.001 for all scores, r=0.68, p<0.001 for complete scores and r=0.46, p<0.07 for incomplete scores. The inter-observer variation of bone marrow burden showed only fair agreement with a Kappa coefficient of 0.26. There is a significant positive correlation between increasing liver stiffness and increasing composite GD-DS3 scores. This supports the use of MRE, a non-invasive reproducible quantitative test, as both an additional assessment and independent marker for monitoring disease severity and progression in GD.