Combined Methylmalonic Aciduria Research Articles

Analysis of clinical phenotypes and MMACHC gene variants in 65 children with Methylmalonic acidemia and homocysteinemia

To carry out Sanger sequencing for MMACHC gene variants among 65 Chinese pedigrees affected with combined methylmalonic aciduria and homocysteinemia, and summarize their genetic and clinical characteristics and prognosis. Clinical characteristics of the 65 children identified with Methylmalonic acidemia and homocysteinemia at the Children's Hospital Affiliated to Zhengzhou University (Zhengzhou Children's Hospital) from April 2017 to April 2022 were selected as the study subjects. Potential variants of the MMACHC gene were detected by direct sequencing of the PCR products. The median age of the 65 children was 3 months (14 days to 17 years old). These included 28 cases (43.08%) from neonatal screening, 11 cases (16.92%) with a history of jaundice, and 9 cases (13.85%) with various degrees of anemia. The main clinical symptoms included development delay, slow growth, epilepsy, hydrocephalus, lethargy, feeding difficulty, regression or decline in motor ability, recurrent respiratory infections, anemia, jaundice, respiratory and heart failures, hydrocephalus, limb weakness, and hypertension. Blood and urine tandem mass spectrometry screening has revealed increase of methylmalonic acid, propionyl carnitine, propionyl carnitine/acetylcarnitine ratio, and propionyl carnitine/free carnitine ratio to various extents, and blood homocysteine was increased in all patients. The detection rate of genetic variants was 98.46% (128/130), and in total 22 types of MMACHC gene variants were detected. The most common ones have included c.609G>A (W203X) (58/128), c.658-660del (K220del) (19/128), and c.80A>G (Q27A) (16/128). Two novel variants have been identified, namely c.565C>T (p.R189C) and c.624_ 625delTG (p.A208Afs), which were respectively predicted as likely pathogenic (PM2_Supporting+PM3+PP2+PP3) and pathogenic (PVS1+PM2_Supporting+PM3+PP2) based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). Exon 4 had the highest frequency for the detection. Identification of MMACHC gene variants has confirmed the diagnosis in the children, among which the c.609G>A variant has the highest frequency. Discovery of the new variants has enriched the mutational spectrum of the MMACHC gene.

A teenager with combined methylmalonic aciduria and homocystinuria (CblC type) presenting with neurological symptoms and congenital heart diseases: a case report

ABSTRACT Combined methylmalonic acidemia and homocystinuria, is a rare autosomal recessive disorder due to defective intracellular cobalamin metabolism. We report an 18-year-old Chinese male who presented with hypermyotonia, seizures, and congenital heart diseases. Mutation analysis revealed c.365A>T and c.482 G>A mutations in the MMACHC gene, diagnosed with methylmalonic aciduria and homocystinuria (CblC type). After treatment with vitamin B12, L-carnitine, betaine, and folate, which resulted in an improvement in his clinical symptoms and laboratory values. This case emphasizes that inborn errors of metabolism should be considered for a teenager presenting with challenging or neurologic symptoms, especially when combined with unexplained heart diseases.

Comparing amniotic fluid mass spectrometry assays and amniocyte gene analyses for the prenatal diagnosis of methylmalonic aciduria

BackgroundMethylmalonic aciduria (MMA), a rare inherited disorder, is the most common organic aciduria in China, and prenatal diagnosis has contributed to its prevention. However, the prenatal diagnosis of MMA using cultured amniocytes or chorionic villi to detect gene mutations is exclusively applicable to families with a definite genetic diagnosis. To evaluate the reliability of mass spectrometry assays for the prenatal diagnosis of MMA, we conducted a retrospective study of our 10 years’ experience.Materials and methodsThis retrospective compare study reviewed the medical records for maternal and fetuses data for 287 mothers with a family history of MMA from June 2010 to December 2020. Methylmalonate and propionylcarnitine in cell-free amniotic fluid were measured using a stable isotope dilution method (GC/MS) and MS/MS-based method (LC/MS/MS). Total homocysteine (tHcy) was measured by fluorescence polarization immunoassay. Depending on the presence of disease-causing gene mutations in probands, gene studies on amniocytes from 222 pregnant women were performed.ResultsFor 222 fetuses of the families with definite genetic diagnosis, gene analyses were performed using cultured amniocytes. 52 fetuses were affected by MMA, whereas 170 were “unaffected”. For GC/MS and LC/MS/MS, the specificity was 96.5% and 95.9%, sensitivity was 71.2% and 84.6%, respectively. The positive and negative predictive values were 86.0% and 91.6% and 86.3% and 95.3%, respectively. Propionylcarnitine/butyrylcarnitine ratio showed the highest accuracy and could thus serve as a sensitive indicator to identify those at a risk for MMA. When GC/MS and LC/MS/MS were performed in parallel, the specificity was 92.5% and sensitivity was 95.6%. When evaluating tHcy, the positive and negative predictive values were 95.0% and 96.1%, respectively. In 65 fetuses without family genetic diagnosis, 11 were finally confirmed to have MMA and 54 were “unaffected” by amniotic fluid biochemical assays. The 54 children showed normal urine organic acids and healthy development after birth.ConclusionsAmniotic fluid biochemical assays using GC/MS and LC/MS/MS in parallel increased the accuracy of prenatal diagnosis of MMA. Propionylcarnitine is a more reliable marker than methylmalonic acid in amniotic fluid. Further, tHcy is recommended for the prenatal diagnosis of combined MMA and homocysteinemia.

Inherited defects of cobalamin metabolism.

Cobalamin (vitamin B12) is required for activity of the enzymes methylmalonyl-CoA mutase and methionine synthase in human cells. Inborn errors affecting cobalamin uptake or metabolism are characterized by accumulation of the substrates for these enzymes, methylmalonic acid and homocysteine, in blood and urine. Inborn errors affecting synthesis of the adenosylcobalamin coenzyme required by methylmalonyl-CoA mutase (cblA and cblB) result in isolated methylmalonic aciduria; inborn errors affecting synthesis of the methylcobalamin coenzyme required by methionine synthase (cblE and cblG) result in isolated homocystinuria. Combined methylmalonic aciduria and homocystinuria is seen in patients with impaired intestinal cobalamin absorption (intrinsic factor deficiency, Imerslund-Gräsbeck syndrome) and with defects affecting synthesis of both cobalamin coenzymes (cblC, cblD, cblF and cblJ). A series of disorders caused by pathogenic variant mutations affecting gene regulators (transcription factors) of the MMACHC gene have recently been described (HCFC1 [cblX disorder] and deficiencies of THAP11, and ZNF143 [the cblK disorder]).

Outcomes of patients with cobalamin C deficiency: A single center experience.

Biallelic variants in MMACHC results in the combined methylmalonic aciduria and homocystinuria, called cobalamin (cbl) C (cblC) deficiency. We report 26 patients with cblC deficiency with their phenotypes, genotypes, biochemical parameters, and treatment outcomes, who were diagnosed and treated at our center. We divided all cblC patients into two groups: group 1: SX group: identified after manifestations of symptoms (n = 11) and group 2: NB group: identified during the asymptomatic period via newborn screening (NBS) or positive family history of cblC deficiency (n = 15). All patients in the SX group had global developmental delay and/or cognitive dysfunction at the time of the diagnosis and at the last assessment. Seizure, stroke, retinopathy, anemia, cerebral atrophy, and thin corpus callosum in brain magnetic resonance imaging (MRI) were common in patients in the SX group. Global developmental delay and cognitive dysfunction was present in nine patients in the NB group at the last assessment. Retinopathy, anemia, and cerebral atrophy and thin corpus callosum in brain MRI were less frequent. We report favorable outcomes in patients identified in the neonatal period and treated pre‐symptomatically. Identification of cblC deficiency by NBS is crucial to improve neurodevelopmental outcomes.

Prospective evaluation of pregnancy outcome in an Italian woman with late-onset combined homocystinuria and methylmalonic aciduria

BackgroundCobalamin metabolism disorders are rare, inherited diseases which cause megaloblastic anaemia and other clinical manifestations. Early diagnosis of these conditions is essential, in order to allow appropriate treatment as early as possible.Case presentationHere we report the case of a patient who was apparently healthy until the age of 20, when she presented with impaired renal function and normocytic anaemia. At the age of 34, when her first pregnancy resulted in an intrauterine death of a morphologically normal growth-restricted foetus, she was diagnosed with homocystinuria and methylmalonic aciduria due to cyanocobalamin C (cblC) defect, which was confirmed by molecular investigation. Consequently, hydroxocobalamin was administered to correct homocysteine plasma levels. This treatment was efficacious in lowering homocysteine plasma levels and restored anaemia and renal function. During a second pregnancy, the patient was also administered a prophylactic dose of low molecular -weight heparin. The pregnancy concluded with a full-term delivery of a healthy male.ConclusionsThis case emphasises the importance of awareness and appropriate management of rare metabolic diseases during pregnancy. We suggest that women with late-onset cblC defect can have a positive pregnancy outcome if this metabolic disease is treated adequately.

Molecular genetic characterization of cblC defects in 126 pedigrees and prenatal genetic diagnosis of pedigrees with combined methylmalonic aciduria and homocystinuria

BackgroundWe sought to analyse MMACHC variants among 126 pedigrees with cobalamin (cbl) C deficiency and combined methylmalonic aciduria and homocystinuria by Sanger sequencing, characterize the spectrum of MMACHC gene variants, and perform prenatal genetic diagnosis by chorionic villus sampling among these pedigrees.MethodsPeripheral blood was collected from 126 probands and their parents who visited the Genetic Counseling Clinic at our hospital between January 2014 and December 2017, and DNA was extracted from the blood. Then, we amplified the coding sequence and splicing regions of the MMACHC gene by PCR, and the PCR products were further sequenced to detect the variants in each pedigree. In 62 families, pregnant women were subjected to chorionic villus sampling for prenatal genetic diagnosis.ResultsIn total, 31 distinct variants were detected in the 126 pedigrees, and the most frequent variants were c.609G > A (p.Trp203Ter), c.658_660delAAG (p.Lys220del), c.567dupT (p.Ile190Tyrfs*13) and c.80A > G (p.Gln27Arg). Two of these variants have not been previously reported in the literature. One variant [c.463_465delGGG (p.Gly155del)] is a small-scale deletion, and the other variant [c.637G>T(p.Glu213Ter)] is a nonsense mutation. Among the 62 pedigrees who received a prenatal diagnosis, 16 foetuses were normal, 34 foetuses were carriers of heterozygous variants, and the remaining 12 foetuses harboured compound heterozygous variants or homozygous variants. Couples whose foetuses were normal or carriers continued the pregnancy, whereas couples whose foetuses harboured compound heterozygous variants or homozygous variants decided to terminate the pregnancy. The follow-up results were consistent with the prenatal diagnosis.ConclusionsTwo novel MMACHC variants were identified, and prenatal genetic diagnosis is an accurate and convenient method that helps avoid the delivery of combined methylmalonic aciduria and homocystinuria patients.

Methylmalonic acidemia: Current status and research priorities.

Methylmalonic acidemia (MMA) is a lethal, severe heterogeneous disorder of methylmalonate and cobalamin (cbl; vitamin B12) metabolism with poor prognosis. Two main forms of the disease have been identified, isolated methylmalonic acidurias and combined methylmalonic aciduria and homocystinuria, which is respectively caused by different gene mutations. Here, we review the improvement of pathogenesis, diagnosis and treatment in MMA. Importantly, the reported epidemiological data of MMA patients in China and the hot mutation sites in Chinese patients are listed, which will aid in improving healthcare of Chinese patients in the future. c.729_730insTT was the most common mutation in Chinese isolated MMA patients, while c.609G>A and c.658_660delAAG were in Chinese cblC type patients according to unrelated studies. The estimated newborn screening incidence was reported to be 1:26,000, 1:3,920, 1:11,160, 1:6,032 respectively in Beijing and Shanghai, Shandong province, Taian district, and Henan province of China. Alternatively, when patients with suspected inherited metabolic diseases were used as the screened sample, the relatively high incidence 0.3% and 1.32% were respectively obtained in southern China and throughout all the provinces of mainland China and Macao with the exception of five provinces (Hainan, Neimenggu, Tibet, Ningxia, and Hong Kong).

Cobalamin D Deficiency Identified Through Newborn Screening.

Cobalamin D deficiency (cblD) is one of the least common cobalamin metabolism disorders. It may result in isolated homocystinuria, isolated methylmalonic aciduria, or combined methylmalonic aciduria and homocystinuria (cblD-combined). Only seven cases of the combined cblD form have been reported to date. Due to the rarity of this disorder, the presentation and symptoms are not well described. We present an eighth case of the cblD-combined subtype, who had a positive newborn screen (NBS) on day of life 3. She was symptomatic and developed lethargy and poor oral intake at 8days of life. She was treated with 10% dextrose, folinic acid, intramuscular hydroxocobalamin, and betaine. Despite the early initiation of treatment, she developed complications of the disease and was found to have abnormal brain imaging findings at 17days of age and macular atrophy at 3months of age and has global developmental delay. We provide detailed description of her presentation, her treatment, and her complications to aid in the understanding of this rare disorder, which is very similar to the more common cobalamin C disorder (cblC).

P-076: A successful pregnancy in a woman with late-onset combined homocystinuria and methylmalonic aciduria

P-076: A successful pregnancy in a woman with late-onset combined homocystinuria and methylmalonic aciduria

Combined methylmalonic aciduria and homocystinuria

Combined methylmalonic aciduria and homocystinuria is a very rare disease caused by a defect in the synthesis of two cofactors in cobalamin dependent reactions, i.e. adenosylcobalamin and methylcobalamin, active forms of vitamin B12. Early onset disease consists of neurological, hematological and gastrointestinal abnormalities seen in the first year of life and some minor face abnormalities (long philtrum, wide forehead, big and low set ears, long face). Here we report a 4-month-old male with failure to thrive, pancytopenia and neurological disturbance.

Prenatal diagnosis using genetic sequencing and identification of a novel mutation in MMACHC.

BackgroundCombined methylmalonic aciduria and homocystinuria, cobalamin(cbl)C deficiency, is a rare disorder of intracellular vitamin B12(cbl) metabolism caused by mutations in the MMACHC gene. Both genetic and biochemical approach have been established to diagnose children and fetuses with cblC deficiency, while in China there is no report of prenatal genetic diagnosis of cblC deficiency. The aim of the present study was to characterize the mutational spectrum of cblC deficiency and investigate the feasibility of genetic-sequencing-based prenatal diagnosis for cblC deficiency.Methods10 pedigrees were recruited in this study with the probands clinically and biochemically confirmed combined methymalonic aciduria and homocystinuria. Peripheral blood samples were collected for MMACHC genetic test from the probands and their parents (4 probands had already dead) and 50 control subjects. The entire coding region and adjacent splice sites of MMACHC were sequenced. After the genotypes of the pedigrees were identified, chorionic villi sampling were performed for 3 high-risk pregnant women for prenatal genetic diagnosis.ResultsA total of 7 mutations were identified: c.217C > T (R73X), c.394C > T (R132X), c.463G > C (G155R), c.609G > A (W203X), c.616C > T (R206W), c.658-660delAAG (220delK), and c.567dupT (I190YfsX13), as well as 2 polymophsims: c.321G > A(V107V), c.-302G > T. And G155R is a novel mutation that haven’t been reported in the literatures. All the 6 probands identified with compound heterozygous mutations or homozygous mutations of MMACHC gene, and all the parents of the probands were found to have one MMACHC mutation at a heterozygous level. Prenatal diagnosis of fetuses from 3 families with a child affected cblC deficiency showed that one fetus had the same compound heterozygous mutations as the proband, one did not have MMACHC mutation, and the third fetus had a mutation at a heterozygous level of MMACHC gene. Results from the follow-ups were consistent with the prenatal diagnosis.ConclusionA novel mutation p.G155R of the MMACHC gene is identified. Genetic diagonsis is an accurate and convenient method for prenatal diagnosis and early intervention of combined methylmalonic aciduria and homocystinuria.

P84 – 2801: Two novel, disease-causing mutations in the ACSF3 gene in a patient with combined malonic and methylmalonic aciduria

Background Combined malonic and methylmalonic aciduria (CMAMMA) is a rare inborn error of metabolism with elevated excretion of malonic acid (MA) and methylmalonic acid (MMA) in urine commonly caused by mutations in the MLYCD gene leading to malonyl-CoA decarboxylase (MCD) deficiency. Clinical features are metabolic acidosis, seizures, axial hypotonia, developmental delay, failure to thrive, microcephaly, cardiomyopathy, elevated transaminases and gastrointestinal symptoms. Benign clinical courses and asymptomatic patients summarized as non-classical phenotype with mostly normal MCD activity are described. In these patients mutations in the ACSF3 gene encoding a methylmalonyl-CoA and malonyl-CoA synthetase are frequently detected. Case report We describe a 3-year-old boy diagnosed with CMAMMA at the age of 26 months. After an uneventful pregnancy a boy of unrelated Afghan parents was born at term with a birth weight of 2955 g. At three weeks he presented with two generalized seizures and metabolic acidosis. Plasma lactat level was normal. Initially moderately elevated levels of MMA were found in urine. Plasma amino acids, carnitine levels, acylcarnitine profile, homocysteine, folate, vitamin B12, liver and renal function parameters were normal. Initial electroencephalogram (EEG) was pathologic and anticonvulsive therapy with phenobarbital was initiated. No further seizures occurred, EEG normalized at the age of three months and antiepileptic medication was stopped. Besides microcephaly the development is age-appropriate up to now. Asymptomatic cholelithiasis persisted despite therapy with ursodeoxycholic acid. Echocardiography was normal. Genetic analysis revealed two novel, disease-causing mutations in the ACSF3 gene and led to the diagnosis auf CMAMMA. Excretion of MMA in urine decreased spontaneously. A mild protein restricted diet was started. A younger sister also presented generalized seizure and elevated MMA in urine at the age of four weeks. Her genetic analysis is in progress. Conclusion We found two novel mutations in the ACSF3 gene which we assume to cause a non-classic CMAMMA phenotype.

First Chinese case of successful pregnancy with combined methylmalonic aciduria and homocystinuria, cblC type

Combined methylmalonic aciduria (MMA) and homocystinuria, cblC type, is the most common MMA in Mainland China. Its clinical spectrum varies from severe neonatal-onset forms with brain injury and high mortality to milder forms with late onset. Timely diagnosis and adequate treatment greatly improve the prognosis. In the past 15 years, many Chinese patients with this condition have achieved favorable treatment outcomes, and some of them have reached childbearing age. Here, we report the first case of Chinese woman with cblC undergoing a successful pregnancy and delivering a healthy boy. A patient of late-onset cblC, who achieved successful pregnancy and delivery of a healthy boy, was enrolled in our studies. The patient and her disease characteristics were analyzed, including age at onset, age at diagnosis, clinical presentation, clinical classification, family history, laboratory findings and MMACHC gene mutation. Our patient presented mild neurological symptoms at the age of 15 years. She had the compound heterozygous mutations, c.315C>G and c.482G>A, on the MMACHC gene. After long-term treatment with cobalamin, calcium folinate, l-carnitine and betaine, along with normal diet, she recovered completely. At age 23, she visited us for genetic counseling and fetal evaluation at 15 weeks of gestation. Her general condition and the fetal growth were normal. At 20 weeks of gestation, intramuscular cobalamin was changed to pure hydroxocobalamin (1 mg, every other day) because of markedly elevated urine MMA and plasma total homocysteine. In addition, the dosages of l-carnitine (1 g, p.o., tds), folic acid (5 mg, p.o., tds) and betaine (1 g, p.o., tds) were increased. Protein intake was not restricted. This is the first report of a Chinese woman with cblC, undergoing a successful pregnancy and delivering a healthy baby at term. The favorable outcome of the patient and her fetus should owe much to the mild phenotype of her disease.

Characterization of functional domains of the cblD (MMADHC) gene product.

In humans vitamin B12 (cobalamin, Cbl) must be converted into two coenzyme forms, methylcobalamin (MeCbl) and adenosylcobalamin (AdoCbl), in order to maintain intracellular homeostasis of homocysteine and methylmalonic acid, respectively. Previously we have shown that in cblD patients three types of MMADHC mutations exist: 1) null mutations N-terminal to Met116 cause isolated methylmalonic aciduria (cblD-MMA) due to AdoCbl deficiency; 2) null mutations across the C-terminus (p.Y140-R250) cause combined methylmalonic aciduria and homocystinuria (cblD-MMA/HC) due to AdoCbl and MeCbl deficiency; 3) missense mutations in a conserved C-terminal region (p.D246-L259) cause isolated homocystinuria (cblD-HC) due to MeCbl deficiency. To better understand the domain boundaries related to MeCbl formation, we made selected point mutations and C-terminal truncations in MMADHC and tested rescue of MeCbl and AdoCbl synthesis in immortalized cblD-MMA/HC patient fibroblasts. Testing 20 mutations (15 missense and five C-terminal truncations) across p.P154-S287 revealed the presence of a region (p.R197-D226) responsible for MeCbl synthesis, which gave a similar cellular phenotype as cblD-HC. Further, mutation of the polypeptide stretch between the new and patient defined regions (p.D226-D246) and directly C-terminal to the patient region (p.L259-R266), gave cellular phenotypes intermediate to those of cblD-HC and cblD-MMA/HC. Finally, C-terminal truncation of more than 20 amino acids resulted in a cblD-MMA/HC like cellular phenotype, while truncation of between ten and 20 amino acids resulted in a cblD-HC like cellular phenotype. These data suggest that specific regions of MMADHC are involved in differential regulation of AdoCbl and MeCbl synthesis and help better define the boundaries of these regions.

Clinical, Biochemical, and Molecular Presentation in a Patient with the cblD-Homocystinuria Inborn Error of Cobalamin Metabolism.

Disorders of intracellular cobalamin (vitamin B12) metabolism result from deficient synthesis of the coenzymes derived from vitamin B12: adenosylcobalamin and methylcobalamin. Disturbances of cobalamin-cofactor synthesis result in elevated levels of homocysteine and/or methylmalonic acid. Nine defects of intracellular cobalamin metabolism have been defined. The most common of these disorders is cblC (combined methylmalonic aciduria and homocystinuria). The cblD disorder is rare with fewer than twenty cases reported in the literature. Some cblD patients have combined methylmalonic aciduria and homocystinuria (referred to as "cblD original," "cblD-combined," or herein "cblD-MMA/HC"); some have isolated homocystinuria (referred to as "cblD-variant 1" or herein "cblD-HC"); and others have isolated methylmalonic aciduria (called "cblD-variant 2" or herein "cblD-MMA"). Only six cases of cblD-HC have been defined thus far. We report the 7th case of cblD-HC. The clinical manifestations, biochemical profile, genetic mutation, and plausible ancestry are discussed.

Neurologic and neurodevelopmental phenotypes in young children with early-treated combined methylmalonic acidemia and homocystinuria, cobalamin C type

Abnormal neurodevelopment has been widely reported in combined methylmalonic aciduria (MMA) and homocystinuria, cblC type (cblC disease), but neurodevelopmental phenotypes in cblC have not previously been systematically studied. We sought to further characterize developmental neurology in children with molecularly-confirmed cblC. Thirteen children at our center with cblC, born since implementation of expanded newborn screening in New York State, undertook standard-of-care evaluations with a pediatric neurologist and pediatric ophthalmologist. At most recent follow-up (mean age 50months, range 9–84months), of twelve children with early-onset cblC, three (25%) had a history of clinical seizures and two (17%) meet criteria for microcephaly. A majority of children had hypotonia and nystagmus. Twelve out of thirteen (92%) underwent neurodevelopmental evaluation (mean age 41months; range 9–76months), each child tested with standardized parental interviews and, where possible, age- and disability-appropriate neuropsychological batteries. All patients showed evidence of developmental delay with the exception of one patient with a genotype predictive of attenuated disease and near-normal biochemical parameters. Neurodevelopmental deficits were noted most prominently in motor skills, with relative preservation of socialization and communication skills. Nine children with early-onset cblC underwent magnetic resonance imaging and spectroscopy (MRI/MRS) at mean age of 47months (range 6–81months); common abnormalities included callosal thinning, craniocaudally short pons, and increased T2 FLAIR signal in periventricular and periatrial white matter. Our study further characterizes variable neurodevelopmental phenotypes in treated cblC, and provides insights into the etiopathogenesis of disordered neurodevelopment frequently encountered in cblC. Plasma homocysteine and MMA, routinely measured at clinical follow-up, may be poor predictors for neurodevelopmental outcomes. Additional data from large, prospective, multi-center natural history studies are required to more accurately define the role of these metabolites and others, as well as that of other genetic and environmental factors in the etiopathogenesis of the neurologic components of this disorder.

Subcellular location of MMACHC and MMADHC, two human proteins central to intracellular vitamin B12 metabolism

MMACHC and MMADHC are the genes responsible for cblC and cblD defects of vitamin B12 metabolism, respectively. Patients with cblC and cblD defects present with various combinations of methylmalonic aciduria (MMA) and homocystinuria (HC). Those with cblC mutations have both MMA and HC whereas cblD patients can present with one of three distinct biochemical phenotypes: isolated MMA, isolated HC, or combined MMA and HC. Based on the subcellular localization of these enzymatic pathways it is thought that MMACHC functions in the cytoplasm while MMADHC functions downstream of MMACHC in both the cytoplasm and the mitochondrion. In this study we determined the subcellular location of MMACHC and MMADHC by immunofluorescence and subcellular fractionation. We show that MMACHC is cytoplasmic while MMADHC is both mitochondrial and cytoplasmic, consistent with the proposal that MMADHC acts as a branch point for vitamin B12 delivery to the cytoplasm and mitochondria. The factors that determine the distribution of MMADHC between the cytoplasm and mitochondria remain unknown. Functional complementation experiments showed that retroviral expression of the GFP tagged constructs rescued all biochemical defects in cblC and cblD fibroblasts except propionate incorporation in cblD-MMA cells, suggesting that the endogenous mutant protein interferes with the function of the transduced wild type construct.

Abnormal findings during newborn period of 160 patients with early-onset methylmalonic aciduria

Methylmalonic aciduria is the most common disorder of organic acidurias in the mainland of China. It is also the one of treatable metabolic disorders. The clinical spectrum of the patients varies from severe neonatal-onset forms with neonatal brain injury and high mortality to milder forms with adult-onset. The clinical manifestations of neonates with methylmalonic aciduria are non-specific. Early diagnosis and adequate treatment contribute a lot to improving the prognosis of the patients. In this study, the abnormal clinical and laboratory findings in neonatal period of 160 Chinese patients with early-onset methylmalonic aciduria were investigated. From 1996 to 2011, a total of 398 patients with methylmalonic aciduria were diagnosed in our hospital; 286 (71.9%) patients had early-onset before 1 year of age. Among 286 patients, 160 (55.9%) presented symptoms in neonatal period. Their urine organic acids were analyzed by gas chromatography-mass spectrometry. Blood amino acids and acylcarnitine profiles were determined by liquid chromatography tandem mass spectrometry. Serum and urine total homocysteine were measured using a fluorescence polarization immunoassay. In some patients, gene analysis was performed. Based on the disease types and general condition, individual dietary and medical interventions were started soon after diagnosis. Out of the 160 patients, 131 (81.9%) had combined methylmalonic aciduria and homocysteinemia. Isolated methylmalonic aciduria was found in 29 cases (18.1%). The common presentations in neonatal period were feeding difficulty, seizures, lethargy and dyspnea. Megaloblastic anemia, liver dysfunction, hyperammonemia and metabolic acidosis were the frequent findings in the routine laboratory test. The most common initial clinical diagnosis was suspected hypoxic-ischemic encephalopathy. Even in 36 cases with abnormal family history, only 3 patients were admitted with suspected inborn errors of metabolism. Five cases (3.1%) were diagnosed by postmortem metabolic examination; 7 cases (4.4%) were detected by newborn screening. In 148 cases (92.5%), the diagnosis was much delayed to the age of one month to 8 years and 5 months (mean 13 months). Methylmalonic aciduria combined with homocysteinemia (MMACHC) gene analyses were performed in 31 cases with combined methylmalonic aciduria. CblC defect was confirmed. The patients with isolated methylmalonic aciduria were treated with protein-restricted diet, cobalamin and L-carnitine. The patients of methylmalonic aciduria combined with homocysteinemia were treated with cobalamin, L-carnitine, calcium folinate, betaine and common diet. Seven patients died without treatment. Clinical improvement was observed in 153 patients. Only 2 patients detected by newborn screening had normal mental and physical development. Mild to severe psychomotor retardation was observed in 151 cases. High mortality and disability rates were observed in the patients with early-onset methylmalonic aciduria. Combined methylmalonic aciduria and homocysteinemia is the common type of methylmalonic aciduria. The clinical manifestation in neonatal period of the patients with early-onset methylmalonic aciduria is complex. Feeding difficulty, seizures, lethargy and dyspnea are the common symptoms in neonatal period of the patients. Megaloblastic anemia, liver dysfunction, hyperammonemia and metabolic acidosis were the frequent laboratory findings.

A clinical and gene analysis of late-onset combined methylmalonic aciduria and homocystinuria, cblC type, in China

BackgroundCombined methylmalonic aciduria and homocystinuria, cblC type (cblC disease), is the most common inborn disorder of cobalamin metabolism. This disorder is caused by MMACHC gene mutations, and it is usually diagnosed in the early neonatal period. Late-onset cblC is rare and difficult to recognize due to a wide diversity of symptoms. MethodsThree cases with late-onset combined methylmalonic aciduria and homocystinuria, cblC type, are reported; patients' clinical presentation, imaging and MMACHC gene mutations were analyzed. ResultsThe age of onset in the three patients was 22years, 40years and 7years of age. Two of the patients had MMACHC gene mutations heterozygous for c.609G>A and c.482G>A (case 1 and case 3). The other patient (case 2) presented with gene mutations heterozygous for c.609G>A and c.1A>G. The three patients presented with a heterogeneous clinical picture, including cognitive impairment, epilepsy, ataxia, pyramidal and peripheral nerve symptoms. Cerebral atrophy and bilateral hyperintensity in the deep white matter were visible in MRI scans of the patients' brains; those were significant findings in the three patients with late-onset cblC disease. In contrast with previous reports, bilateral cerebellar cortex abnormalities were also found in one patient (case 2). ConclusionAlthough its occurrence is rare, late-onset combined methylmalonic aciduria and homocystinuria, cblC type, should be considered in making a differential diagnosis in patients who present with neurological symptoms that are not consistent with common neurological diseases, especially when cognition, the pyramidal tract and peripheral nerves are involved.