A compound heterozygote case of glutaric aciduria type II in a patient carrying a novel candidate variant in ETFDH gene: A case report and literature review on compound heterozygote cases.

BACKGROUND: Glutaric aciduria type II (GA2) is a rare genetic disorder inherited in an autosomal recessive manner. Double dosage mutations in GA2 corresponding genes, ETFDH, ETFA, and ETFB, lead to defects in the catabolism of fatty acids, and amino acids lead to broad-spectrum phenotypes, including muscle weakness, developmental delay, and seizures. product of these three genes have crucial role in transferring electrons to the electron transport chain (ETC), but are not directly involve in ETC complexes. METHODS: Here, by using exome sequencing, the cause of periodic cryptic gastrointestinal complications in a 19-year-old girl was resolved after years of diagnostic odyssey. Protein modeling for the novel variant served as another line of validation for it. RESULTS: Exome Sequencing (ES) identified two variants in ETFDH: ETFDH:c.926T>G and ETFDH:c.1141G>C. These variants are likely contributing to the crisis in this case. To the best of our knowledge at the time of writing this manuscript, variant ETFDH:c.926T>G is reported here for the first time. Clinical manifestations of the case and pathological analysis are in consistent with molecular findings. Protein modeling provided another line of evidence proving the pathogenicity of the novel variant. ETFDH:c.926T>G is reported here for the first time in relation to the causation GA2. CONCLUSION: Given the milder symptoms in this case, a review of GA2 cases caused by compound heterozygous mutations was conducted, highlighting the range of symptoms observed in these patients, from mild fatigue to more severe outcomes. The results underscore the importance of comprehensive genetic analysis in elucidating the spectrum of clinical presentations in GA2 and guiding personalized treatment strategies.

The presence of white cell Jordan's anomaly in multiple Acyl-CoA dehydrogenase deficiency: A case report and implications for clinical practice.

BACKGROUND: Multiple Acyl-CoA Dehydrogenase Deficiency (MADD), also known as Glutaric Aciduria Type II, is an exceptionally rare autosomal recessive genetic disorder that disrupts the metabolism of fatty acids, amino acids, and choline. It presents with a wide range of clinical manifestations, from severe neonatal-onset forms to milder late-onset cases, with symptoms including metabolic disturbances and muscle weakness. Jordan's anomaly is a distinctive morphological feature found in peripheral blood white cells and is typically associated with Neutral Lipid Storage Disease (NLSD). CASE REPORT: In our case report, the patient initially presented with symptoms of vomiting, abdominal pain, and altered consciousness. The presence of white cell Jordan's anomaly was detected in the blood smear. Subsequent serum tests revealed elevated levels of transaminases, creatine kinase, uric acid, and multiple acylcarnitines, while blood glucose and free carnitine levels were notably reduced. High-throughput sequencing confirmed heterozygous pathogenic variants in the electron-transferring flavoprotein dehydrogenase (ETFDH) gene, leading to the conclusive diagnosis of MADD. Following a three-month treatment regimen involving high-dose vitamin B2, coenzyme Q10, and other supportive interventions, the patient exhibited significant clinical improvement, ultimately resulting in discharge. CONCLUSION: The identification of Jordan's anomaly in a pediatric patient with late-onset MADD sheds light on its broader implications within the realm of lipid storage myopathies. The significance of this finding extends beyond its conventional association with NLSD, challenging the notion of its exclusivity. This novel observation serves as a compelling reminder of the diagnostic significance this morphological abnormality holds, potentially revolutionizing diagnostic practices within the field.

Clinical, biochemical, and genetic spectrum of MADD in a South African cohort: an ICGNMD study.

BACKGROUND: Multiple acyl-CoA dehydrogenase deficiency (MADD) is an autosomal recessive disorder resulting from pathogenic variants in three distinct genes, with most of the variants occurring in the electron transfer flavoprotein-ubiquinone oxidoreductase gene (ETFDH). Recent evidence of potential founder variants for MADD in the South African (SA) population, initiated this extensive investigation. As part of the International Centre for Genomic Medicine in Neuromuscular Diseases study, we recruited a cohort of patients diagnosed with MADD from academic medical centres across SA over a three-year period. The aim was to extensively profile the clinical, biochemical, and genomic characteristics of MADD in this understudied population. METHODS: Clinical evaluations and whole exome sequencing were conducted on each patient. Metabolic profiling was performed before and after treatment, where possible. The recessive inheritance and phase of the variants were established via segregation analyses using Sanger sequencing. Lastly, the haplotype and allele frequencies were determined for the two main variants in the four largest SA populations. RESULTS: Twelve unrelated families (ten of White SA and two of mixed ethnicity) with clinically heterogeneous presentations in 14 affected individuals were observed, and five pathogenic ETFDH variants were identified. Based on disease severity and treatment response, three distinct groups emerged. The most severe and fatal presentations were associated with the homozygous c.[1067G > A];c.[1067G > A] and compound heterozygous c.[976G > C];c.[1067G > A] genotypes, causing MADD types I and I/II, respectively. These, along with three less severe compound heterozygous genotypes (c.[1067G > A];c.[1448C > T], c.[740G > T];c.[1448C > T], and c.[287dupA*];c.[1448C > T]), resulting in MADD types II/III, presented before the age of five years, depending on the time and maintenance of intervention. By contrast, the homozygous c.[1448C > T];c.[1448C > T] genotype, which causes MADD type III, presented later in life. Except for the type I, I/II and II cases, urinary metabolic markers for MADD improved/normalised following treatment with riboflavin and L-carnitine. Furthermore, genetic analyses of the most frequent variants (c.[1067G > A] and c.[1448C > T]) revealed a shared haplotype in the region of ETFDH, with SA population-specific allele frequencies of < 0.00067-0.00084%. CONCLUSIONS: This study reveals the first extensive genotype-phenotype profile of a MADD patient cohort from the diverse and understudied SA population. The pathogenic variants and associated variable phenotypes were characterised, which will enable early screening, genetic counselling, and patient-specific treatment of MADD in this population.

Riboflavin-responsive lipid-storage myopathy in elderly patients.

There are scarce reports of riboflavin-responsive lipid storage myopathy in elderly patients with onset in their sixties. We describe three elderly patients with riboflavin-responsive lipid-storage myopathy. All three patients (aged 67-71 years on first examination) had subacute onset of neck extensors and proximal limb weakness progressing to inability to rise from a sitting position or to walk. Muscle biopsies showed vacuoles with lipid content, mainly in type 1 fibers. Genetic analysis failed to identify any pathogenic variant in one patient, identified a heterozygous variant of uncertain significance c.812 A > G; p.Tyr271Cys in the ETFDH gene in the second patient, and revealed a heterozygote likely pathogenic variant c.1286-2 A > C in the ETFDH gene predicted to cause abnormal splicing in the third patient. All patients responded to treatment with riboflavin and carnitine, and regained normal strength. This report emphasizes the importance of muscle biopsy in revealing treatable lipid storage myopathy in elderly patients with progressive myopathy of unidentifiable cause.

"Liver Failure in an Infant of Late-Onset Glutaric Aciduria Type II": Case Report.

Glutaric aciduria type II, also known as Multiple acyl-CoA Dehydrogenase Deficiency, results from a defect in the mitochondrial electron transport chain resulting in an inability to break down fatty-acids and amino acids. There are three phenotypes- type 1 and 2 are of neonatal onset and severe form, with and without congenital anomalies, respectively, and presents with acidosis, severe hypotonia, cardiomyopathy, hepatomegaly, and non-ketotic hypoglycemia. Type 3 or late-onset Multiple acyl-CoA Dehydrogenase Deficiency usually presents in the adolescent or adult age group with phenotype ranging from mild forms of myopathy and exercise intolerance to severe forms of acute metabolic decompensation on its chronic course. Type 3 Multiple acyl-CoA Dehydrogenase Deficiency rarely presents in infancy and in liver failure. We present a five-month-old developmentally normal female child with acute encephalopathy, hypotonia, non-ketotic hypoglycemia, metabolic acidosis, and liver failure, with a history of sibling death of suspected inborn error of metabolism. The blood acyl-carnitine levels in Tandem Mass Spectrometry and urinary organic acid analysis through Gas Chromatography-Mass Spectrometry were unremarkable. The patient initially responded to riboflavin, CoQ, and supportive management but ultimately succumbed to sepsis with shock and multi-organ dysfunction. The clinical exome sequencing reported a homozygous missense variation in exon 11 of the ETFDH gene (chr4:g.158706270C > T) that resulted in the amino acid substitution of Leucine for Proline at codon 456 (p.Pro456Leu) suggestive of Glutaric aciduria type IIc (OMIM#231,680).

Case report: Novel ETFDH compound heterozygous mutations identified in a patient with late-onset glutaric aciduria type II.

Glutaric aciduria type II (GA II) is an autosomal recessive metabolic disorder of fatty acid, amino acid, and choline metabolism. The late-onset form of this disorder is caused by a defect in the mitochondrial electron transfer flavoprotein dehydrogenase or the electron transfer flavoprotein dehydrogenase (ETFDH) gene. Thus far, the high clinical heterogeneity of late-onset GA II has brought a great challenge for its diagnosis. In this study, we reported a 21-year-old Chinese man with muscle weakness, vomiting, and severe pain. Muscle biopsy revealed myopathological patterns of lipid storage myopathy, and urine organic acid analyses showed a slight increase in glycolic acid. All the aforementioned results were consistent with GA II. Whole-exome sequencing (WES), followed by bioinformatics and structural analyses, revealed two compound heterozygous missense mutations: c.1034A > G (p.H345R) on exon 9 and c.1448C>A (p.P483Q) on exon 11, which were classified as "likely pathogenic" according to American College of Medical Genetics and Genomics (ACMG). In conclusion, this study described the phenotype and genotype of a patient with late-onset GA II. The two novel mutations in ETFDH were found in this case, which further expands the list of mutations found in patients with GA II. Because of the treatability of this disease, GA II should be considered in all patients with muscular symptoms and acute metabolism decompensation such as hypoglycemia and acidosis.

Multiple Acyl-Coenzyme A Dehydrogenase Deficiency Leading to Severe Metabolic Acidosis in a Young Adult.

Background: Multiple acyl-coenzyme A dehydrogenase deficiency (MADD) is a rare metabolic disorder affecting fatty acid oxidation. Incidence at birth is estimated at 1:250 000, but type III presents in adults. It is characterized by nonspecific symptoms but if undiagnosed may cause ketoacidosis and rhabdomyolysis. A review of 350 patients found less than one third presented with metabolic crises. Our objective is to describe an adult with weakness after carbohydrate restriction that developed a pulmonary embolism and ketoacidosis, and was diagnosed with MADD type III. Case Report: A 27-year-old woman with obesity presented to the hospital with fatigue and weakness worsening over months causing falls and decreased intake. She presented earlier to clinic with milder symptoms starting months after initiating a low carbohydrate diet. Testing revealed mild hypothyroidism and she started Levothyroxine for presumed hypothyroid myopathy but progressed. Muscle biopsy suggested a lipid storage myopathy. Genetic testing revealed a mutation in the ETFDH (electron transfer flavoprotein dehydrogenase) gene likely pathogenic for MADD; however, before this was available she developed severe ketoacidosis and rhabdomyolysis. She empirically started a low-fat diet, carnitine, cyanocobalamin, and coenzyme Q10 supplementation with improvement. Over months her energy and strength normalized. Discussion: MADD may cause ketoacidosis and rhabdomyolysis but this is rare in adults. Diagnosis requires clinical suspicion followed by biochemical and genetic testing. It should be considered when patients present with weakness or fasting intolerance. Treatment includes high carbohydrate, low-fat diets, supplementation, and avoiding fasting. Conclusion: There should be greater awareness to consider MADD in adults presenting with neuromuscular symptoms, if untreated it may cause severe metabolic derangements.

Clinical and molecular investigation of 37 Japanese patients with multiple acyl-CoA dehydrogenase deficiency: p.Y507D in ETFDH, a common Japanese variant, causes a mortal phenotype.

Multiple acyl-CoA dehydrogenase deficiency (MADD) is an inherited metabolic disease caused by a defect in electron transfer flavoprotein alpha (ETFA), ETF beta (ETFB), or ETF dehydrogenase (ETFDH), and riboflavin metabolism disorders have recently been reported to present as mimicking MADD. MADD is roughly classified into neonatal (type 1 or 2) and later-onset (type 3) forms. To identify clinicogenetic characteristics in Japan, we investigated 37 Japanese patients with MADD diagnosed from 1997 to 2020. The causes of MADD were ETFDH deficiency in 26 patients, ETFA deficiency in four, ETFB deficiency in six, and riboflavin metabolism disorder in one. All 15 patients with the neonatal-onset type died by 2 years of age, while five of 22 patients with the later-onset form died by 3 years of age. Furthermore, 8 of 15 patients with the later-onset form of ETFDH deficiency treated with riboflavin were riboflavin non-responders. p.Y507D in ETFDH was identified as the most common variant (9 of 48 alleles, 18.8%). Of two patients with a homozygous p.Y507D variant, one experienced disease onset and died in the neonatal period, while the other experienced disease onset at two months of age and died at two years old, suggesting that the p.Y507D variant results in fatal outcomes. Our study concluded that more than half of Japanese patients with MADD died by three years old, and more than half of patients with the later-onset form had poor responsiveness to riboflavin, partly due to the unique Japanese p.Y507D variant in ETFDH.

The clinical, pathological, and genetic characteristics of lipid storage myopathy in northern China.

BACKGROUND: The lipid storage myopathy (LSM) diagnosis is based on the patient's clinical manifestations and muscle pathology. However, when genetic testing is lacking, there is a high rate of misdiagnosis of the disease. This study aimed to investigate the clinical and pathological features of genetically diagnosed LSM in northern China, analyze genetic mutations' characteristics, and improve the LSM diagnostic rate. METHODS: Twenty patients with LSM diagnosed were collected; meanwhile, the clinical data, muscle samples, and routine pathological staining of muscle specimens were collected. The morphological changes of muscle fibers were observed under an optical microscope. RESULTS: Among the included patients, 18 cases had ETFDH (HGNC ID: 3483) mutations, and two had PNPLA2 mutations. Family pedigree verification was performed on three patients with heterozygous mutations in the ETFDH gene complex. Histopathological staining showed that all patients had fine vacuoles in the muscle fibers, and some of them merged to form fissures, and the lipid droplets increased in cells. After therapy, 18 patients were associated with a favorable prognosis, and two patients were ineffective with the treatment of neutral lipid storage myopathy (NLSDM) caused by PNPLA2 mutation. DISCUSSION: The clinical manifestations of LSM are complex and diverse, mainly manifested by proximal muscle weakness and exercise intolerance in the extremities. The pathological images of LSM muscles are abnormal storage of lipid droplets in muscle fibers, primarily involving type I fibers. The LSM patients were mainly multiple acyl-CoA dehydrogenase deficiency (MADD) caused by the ETFDH gene mutation. It is necessary to perform an accurate typing diagnosis of LSM.

A fatal case of neonatal onset multiple acyl-CoA dehydrogenase deficiency caused by novel mutation of ETFDH gene: case report.

BACKGROUND: Multiple acyl-CoA dehydrogenase deficiency (MADD) or glutaric aciduria type II is an extremely rare autosomal recessive inborn error of fatty acid beta oxidation and branched-chain amino acids, secondary to mutations in the genes encoding the electron transfer flavoproteins A and B (ETFs; ETFA or ETFB) or ETF dehydrogenase (ETFDH). The clinical manifestation of MADD are heterogeneous, from severe neonatal forms to mild late-onset forms. CASE PRESENTATION: We report the case of a preterm newborn who died a few days after birth for a severe picture of untreatable metabolic acidosis. The diagnosis of neonatal onset MADD was suggested on the basis of clinical features displaying congenital abnormalities and confirmed by the results of expanded newborn screening, which arrived the day the newborn died. Molecular genetic test revealed a homozygous indel variant c.606 + 1 _606 + 2insT in the ETFDH gene, localized in a canonical splite site. This variant, segregated from the two heterozygous parents, is not present in the general population frequency database and has never been reported in the literature. DISCUSSION AND CONCLUSION: Recently introduced Expanded Newborn Screening is very important for a timely diagnosis of Inherited Metabolic Disorders like MADD. In some cases which are the most severe, diagnosis may arrive after symptoms are already present or may be the neonate already died. This stress the importance of collecting all possible samples to give parents a proper diagnosis and a genetic counselling for future pregnacies.

Combined isobutyryl-CoA and multiple acyl-CoA dehydrogenase deficiency in a boy with altered riboflavin homeostasis.

In this report, we describe the case of an 11-year-old boy, who came to our attention for myalgia and muscle weakness, associated with inappetence and vomiting. Hypertransaminasemia was also noted, with ultrasound evidence of hepatomegaly. Biochemical investigations revealed acylcarnitine and organic acid profiles resembling those seen in MADD, that is, multiple acyl-CoA dehydrogenase deficiencies (OMIM #231680) a rare inherited disorder of fatty acids, amino acids, and choline metabolism. The patient carried a single pathogenetic variant in the ETFDH gene (c.524G>A, p.Arg175His) and no pathogenetic variant in the riboflavin (Rf) homeostasis related genes (SLC52A1, SLC52A2, SLC52A3, SLC25A32, FLAD1). Instead, compound heterozygosity was found in the ACAD8 gene (c.512C>G, p.Ser171Cys; c.822C>A, p.Asn274Lys), coding for isobutyryl-CoA dehydrogenase (IBD), whose pathogenic variants are associated to IBD deficiency (OMIM #611283), a rare autosomal recessive disorder of valine catabolism. The c.822C>A was never previously described in a patient. Subsequent further analyses of Rf homeostasis showed reduced levels of flavins in plasma and altered FAD-dependent enzymatic activities in erythrocytes, as well as a significant reduction in the level of the plasma membrane Rf transporter 2 in erythrocytes. The observed Rf/flavin scarcity in this patient, possibly associated with a decreased ETF:QO efficiency might be responsible for the observed MADD-like phenotype. The patient's clinical picture improved after supplementation of Rf, l-carnitine, Coenzyme Q10, and also 3OH-butyrate. This report demonstrates that, even in the absence of genetic defects in genes involved in Rf homeostasis, further targeted molecular analysis may reveal secondary and possibly treatable biochemical alterations in this pattern.

Novel variant of ETFDH leading to multiple acyl-CoA dehydrogenase deficiency by promoting protein degradation via ubiquitin proteasome pathway.

Multiple acyl-CoA dehydrogenase deficiency (MADD) is a rare autosomal recessive metabolic disease. Patients present with metabolic decompensation, muscle weakness, respiratory failure, and cardiomyopathy. Late-onset MADD is primarily caused by mutations in the ETFDH gene. Here, we report a patient who has been diagnosed with Down syndrome after birth following karyotype analysis and simultaneously carrying compound heterozygous variants of ETFDH (c.3G > C (p. M1?); c.725C > T (p. T242I), which is novel). Further molecular analyses revealed that the novel c.725C > T (p. T242I) mutation enhances the degradation of electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) via the ubiquitin proteasome pathway. Five ubiquitin E3 ligases (STUB1, RNF40, UBE3C, CUL3, and CUL1) and one ubiquitin modification site (Cystein, C101) of the ETF-QO were reported in this study. Our study not only expanded the pathogenic variant spectrum of ETFDH gene but also proved that the c.725C > T (p. T242I) will promote protein degradation through ubiquitin proteasome pathway.

Diagnostic Challenges in Late Onset Multiple Acyl-CoA Dehydrogenase Deficiency: Clinical, Morphological, and Genetic Aspects.

Background: Multiple acyl-CoA dehydrogenase deficiency (MADD) is an autosomal recessive disorder of fatty acid oxidation due to deficiency of the mitochondrial electron transfer chain. The late-onset form is characterized by exercise intolerance, muscle weakness, and lipid storage in myofibers. Most MADD patients greatly benefit from riboflavin supplementation. Patients and methods: A retrospective study was conducted on patients with a diagnosis of vacuolar myopathy with lipid storage followed in our neuromuscular unit in the last 20 years. We selected 10 unrelated patients with the diagnosis of MADD according to clinical, morphological, and biochemical aspects. Clinical features, blood tests including serum acylcarnitines, EMG, and ENG were revised. Muscle biopsy was performed in all, and one individual underwent also a sural nerve biopsy. Gene sequencing of ETFA, ETFB, and ETFDH was performed as a first-tier genetic analysis followed by next-generation sequencing of an hyperCKemia gene panel in patients with undefined genotypes. Results: Clinical evaluation at onset in all our patients showed fatigue and muscle weakness; four patients showed difficulties in chewing, three patients complained of dysphagia, two patients had a dropped head, and a patient had an unexpected ataxia with numbness and dysesthesia. Laboratory blood tests revealed a variable increase in serum CK (266-6,500) and LDH levels (500-2,000). Plasma acylcarnitine profile evidenced increased levels of different chains intermediates. EMG was either normal or showed myogenic or neurogenic patterns. NCS demonstrated sensory neuropathy in two patients. Muscle biopsies showed a vacuolar myopathy with a variable increase in lipid content. Nerve biopsy evidenced an axonal degeneration with the loss of myelinated fibers. ETFDH genetic analysis identifies 14 pathogenic variants. Patients were treated with high doses of riboflavin (400 mg/die). All of them showed a rapid muscle strength improvement and normalization of abnormal values in laboratory tests. Neuropathic symptoms did not improve. Conclusion: Our data confirmed that clinical features in MADD patients are extremely variable in terms of disease onset and symptoms making diagnosis difficult. Laboratory investigations, such as serum acylcarnitine profile and muscle biopsy evaluation, may strongly address to a correct diagnosis. The favorable response to riboflavin supplementation strengthens the importance of an early diagnosis of these disorders among the spectrum of metabolic myopathies.

Case report: A novel c.1842_1845dup mutation of ETFDH in two Chinese siblings with multiple acyl-CoA dehydrogenase deficiency.

This article reports the characterization of two siblings diagnosed with late-onset multiple Acyl-CoA dehydrogenase deficiency (MADD) caused by mutations in electron transfer flavoprotein(ETF)-ubiquinone oxidoreductase (ETF-QO) (ETFDH) gene. Whole exome sequencing (WES) was performed in the proband's pedigree. Clinical phenotypes of Proband 1 (acidosis, hypoglycemia, hypotonia, muscle weakness, vomiting, hypoglycemia, hepatomegaly, glutaric acidemia, and glutaric aciduria) were consistent with symptoms of MADD caused by the ETFDH mutation. However, Proband 2 presented with only a short stature. The patients (exhibiting Probands 1 and 2) showed identical elevations of C6, C8, C10, C12, and C14:1. c.1842_1845 (exon13)dup, and c.250 (exon3) G > A of the ETFDH gene were compound heterozygous variants in both patients. The novel variant c.1842_1845dup was rated as likely pathogenic according to the American College of Medical Genetics and Genomics guidelines (ACMG). This is the first report on the c.1842_1845dup mutation of the ETFDH gene in patients with late-onset MADD, and the data described herein may help expand the mutation spectrum of ETFDH.

Lipid storage myopathy due to late-onset multiple Acyl-CoA dehydrogenase deficiency with novel mutations in ETFDH: A case report.

Background: Lipid storage myopathy (LSM) is an autosomal recessive inherited lipid and amino metabolic disorder with great clinical heterogeneity. Variations in the electron transfer flavoprotein dehydrogenase (ETFDH) gene cause multiple acyl-CoA dehydrogenase deficiency (MADD), and have a manifestation of LSM. Muscle biopsy helps clarify the diagnosis of LSM, and next-generation sequencing (NGS) can be useful in identifying genomic mutation sites. The diagnosis of MADD contributes to targeted therapy. Case presentation: We report on a teenager who appeared to have muscle weakness and exercise intolerance at the onset. Before the referral to our hospital, he was unsuccessfully treated with glucocorticoid for suspected polymyositis. The next-generation sequencing of the proband and his parents revealed heterozygous variations, c.365G>A (p.G122D) inherited from the father, c.176-194_176-193del, and c.832-316C>T inherited from the mother in the ETFDH gene. The tandem mass spectrometry identified the mutations to be pathogenic. However, his parents and his younger sister who were detected with a mutation of c.365G>A presented no clinical symptoms. This indicates that the combination of the three compound heterozygous mutations in ETFDH is significant. After MADD was diagnosed, a dramatic clinical recovery and biochemical improvement presented as riboflavin was given to the patient across a week, which further confirmed the diagnosis of MADD. Conclusion: Our observations extend the spectrum of ETFDH variants in Chinese the population and reinforce the role of NGS in diagnosis of MADD. Early diagnosis and appropriate treatment of LSM lead to great clinical efficacy and avoid some lethal complications.

Clinical，pathological and gene mutation characteristics of 18 cases of riboflavin-responsive lipid storage myopathy caused by ETFDH gene mutation / 中风与神经疾病杂志

@#To investigate the clinical，pathological and ETFDH gene mutation characteristics of riboflavinresponsive lipid storage myopathy（RRLSM） caused by ETFDH gene mutation.Methods The clinical and pathological data of 18 RR-LSM patients who were admitted to our hospital from January 2009 to December 2020 and confirmed by muscle biopsy pathology and gene testing were retrospectively analyzed. The Illumina NovaSeq highthroughput sequencing of peripheral blood DNA was performed for data reading and bioinformatics analysis.Results Among the 18 patients，there were 9 males and 9 females. The age of onset ranged from 9 to 60 years old（mean 29.83±13.44 years）. The course of disease ranged from 1 month to 22 years（mean 4.5 years）. The main clinical manifestations were proximal limb weakness and movement intolerance，accompanied by cervical extensor weakness in 14 cases，masticatory muscle weakness in 9 cases，dysphagia in 5 cases，nausea and poor appetite in 5 cases，and muscle pain and dyspnea in a few patients. Muscle pathology showed a large amount of lipid deposition in muscle fibers and a small amount of muscle fiber necrosis in 5 cases. Immunohistochemical staining indicated that the necrotic muscle fibers were mainly infiltrated by CD68（+） lymphophagocytes. All 18 patients were treated with riboflavin and had good efficacy. ETFDH gene mutation was detected in all cases in this study，including 15 cases（83.3%） with complex heterozygous mutation，2 cases（11.1%） with single heterozygous mutation，and 1 case（5.6%） with homozygous mutation. A total of 20 mutation sites were found，among which the most frequent mutation site was c.770A>G，accounting for 19.4%（7/36） of the allele，followed by c.1454C>G，accounting for 8.3%（3/36） of the allele. Conclusion RRLSM patients caused by ETFDH gene mutation are characterized by trunk axial muscle and masticatory muscle involvement. Muscle pathology found that there are a large number of lipid deposits in muscle fibers is an important basis for diagnosis. c.770A>G and c.1454C>G are the most common mutation sites of ETFDH gene in this grou.

Mimicking human riboflavin responsive neuromuscular disorders by silencing flad-1 gene in C. elegans: Alteration of vitamin transport and cholinergic transmission.

Riboflavin (Rf), or vitamin B2, is the precursor of FMN and FAD, redox cofactors of several dehydrogenases involved in energy metabolism, redox balance and other cell regulatory processes. FAD synthase, coded by FLAD1 gene in humans, is the last enzyme in the pathway converting Rf into FAD. Mutations in FLAD1 gene are responsible for neuromuscular disorders, in some cases treatable with Rf. In order to mimic these disorders, the Caenorhabditis elegans (C. elegans) gene orthologue of FLAD1 (flad-1) was silenced in a model strain hypersensitive to RNA interference in nervous system. Silencing flad-1 resulted in a significant decrease in total flavin content, paralleled by a decrease in the level of the FAD-dependent ETFDH protein and by a secondary transcriptional down-regulation of the Rf transporter 1 (rft-1) possibly responsible for the total flavin content decrease. Conversely an increased ETFDH mRNA content was found. These biochemical changes were accompanied by significant phenotypical changes, including impairments of fertility and locomotion due to altered cholinergic transmission, as indicated by the increased sensitivity to aldicarb. A proposal is made that neuronal acetylcholine production/release is affected by alteration of Rf homeostasis. Rf supplementation restored flavin content, increased rft-1 transcript levels and eliminated locomotion defects. In this aspect, C. elegans could provide a low-cost animal model to elucidate the molecular rationale for Rf therapy in human Rf responsive neuromuscular disorders and to screen other molecules with therapeutic potential.

Clinical Presentations and Genetic Characteristics of Late-Onset MADD Due to ETFDH Mutations in Five Patients: A Case Series.

Background: Late-onset multiple acyl-CoA dehydrogenase deficiency (LO-MADD) describes a curable autosomal recessive genetic disease caused by ETFDH mutations that result in defects in ETF-ubiquinone oxidoreductase. Almost all patients are responsive to riboflavin. This study describes the clinical presentations and genetic characteristics of five LO-MADD patients. Methods: From 2018 to 2021, we collected clinical and genetic data on five patients diagnosed with LO-MADD at our hospital and retrospectively analyzed their clinical characteristics, laboratory examination, electromyography, muscle biopsy, genetic analysis, and outcome data. Results: This study included three males and two females with mean onset age of 37.8 years. Fluctuating exercise intolerance was the most common presentation. Serum creatine kinase (CK) levels were significantly elevated in all patients, and plasma acylcarnitine profiles revealed an increase in long-chain acylcarnitine species in three cases. The urinary organic acid study revealed a high level of hydroxyglutaric acid in all patients. Electrophysiology demonstrated myogenic impairment. Muscle biopsies revealed lipid storage myopathy. Molecular analysis identified nine mutations (three novels and six reported) in ETFDH. Exercise intolerance and muscle weakness were dramatically improved in all patients treated with riboflavin (100 mg) daily following diagnosis. Conclusions: LO-MADD is caused by ETFDH variants and responds well to riboflavin. Three novel ETFDH pathogenic variants were identified, expanding their spectrum in the Chinese population and facilitating future interpretation and analysis of ETFDH mutations.

Multiple Acyl-CoA Dehydrogenase Deficiency with Variable Presentation Due to a Homozygous Mutation in a Bedouin Tribe.

Multiple acyl-CoA dehydrogenase deficiency (MADD) is a fatty acid and amino acid oxidation defect caused by a deficiency of the electron-transfer flavoprotein (ETF) or the electron-transfer flavoprotein dehydrogenase (ETFDH). There are three phenotypes of the disease, two neonatal forms and one late-onset. Previous studies have suggested that there is a phenotype-genotype correlation. We report on six patients from a single Bedouin tribe, five of whom were sequenced and found to be homozygous to the same variant in the ETFDH gene, with variable severity and age of presentation. The variant, NM_004453.3 (ETFDH): c.524G>A, p.(R175H), was previously recognized as pathogenic, although it has not been reported in the literature in a homozygous state before. R175H is located near the FAD binding site, likely affecting the affinity of FAD for EFT:QO. The single homozygous ETFDH pathogenic variant was found to be causing MADD in this cohort with an unexpectedly variable severity of presentation. The difference in severity could partly be explained by early diagnosis via newborn screening and early treatment with the FAD precursor riboflavin, highlighting the importance of early detection by newborn screening.

Role of RNA in Molecular Diagnosis of MADD Patients.

The electron-transfer flavoprotein dehydrogenase gene (ETFDH) encodes the ETF-ubiquinone oxidoreductase (ETF-QO) and has been reported to be the major cause of multiple acyl-CoA dehydrogenase deficiency (MADD). In this study, we present the clinical and molecular diagnostic challenges, at the DNA and RNA levels, involved in establishing the genotype of four MADD patients with novel ETFDH variants: a missense variant, two deep intronic variants and a gross deletion. RNA sequencing allowed the identification of the second causative allele in all studied patients. Simultaneous DNA and RNA investigation can increase the number of MADD patients that can be confirmed following the suggestive data results of an expanded newborn screening program. In clinical practice, accurate identification of pathogenic mutations is fundamental, particularly with regard to diagnostic, prognostic, therapeutic and ethical issues. Our study highlights the importance of RNA studies for a definitive molecular diagnosis of MADD patients, expands the background of ETFDH mutations and will be important in providing an accurate genetic counseling and a prenatal diagnosis for the affected families.