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Home , Fumarase, Fumarase deficiency, Fumaric acid, Glutamate dehydrogenase

J Inherit Metab Dis (2010) 33:411–419 DOI 10.1007/s10545-010-9134-2

ORIGINAL ARTICLE

Fumaric aciduria: an overview and the first Brazilian case report

Gabriella Allegri & Marcia J. Fernandes & Fernanda B. Scalco & Patricia Correia & Ruth E. Simoni & Juan C. Llerena Jr & Maria L. Costa de Oliveira

Received: 18 January 2010 /Revised: 5 May 2010 /Accepted: 12 May 2010 /Published online: 15 June 2010 # SSIEM and Springer 2010

Abstract Fumaric aciduria is a rare metabolic disease, Abbreviations with 40 cases reported so far. deficiency leads AICAR 5-aminoimidazole-4-carboxamide ribotide mainly to brain abnormalities, developmental delay, and AMP great accumulation of in urine. This work AS argininosuccinase presents the first case of fumaric aciduria described in ASL adenylosuccinate Brazil, which presented with some interesting clinical ASS argininosuccinic synthetase and biochemical findings such as colpocephaly, hepatic AST aspartate aminotransferase alterations, and marked metabolic acidosis since birth. CPSI carbamoylphosphate synthetase I. Common findings were ventriculomegaly, , and FAH fumarylacetoacetate microcephaly. Biochemically, besides the high urinary FAICAR 5-formaminoimidazole-4-carboxamide ribotide fumaric acid excretion, atypical elevation of plasma FH fumarate hydratase , and levels were also GD glutamate observed. In order to show all features and variants of 4-HPPD 4-hydroxy-phenylpyruvate dioxygenase fumaric aciduria, literature data of 40 patients was IMP inosine monophosphate reviewed and compared with the case reported here. MAI maleylacetoacetate Findings in all these patients demonstrate that this ME malic disorder does not yet have its phenotype completely MDH defined; it is important that more patients be described. OT transcarbamoylase PC PDHC complex SAICAR 5-aminoimidazole-(N-succinylcarboxiamide) Communicated by: Jan Smeitink ribotide SDH Competing interest: None declared. TAT tyrosine aminotransferase G. Allegri (*) : M. J. Fernandes : F. B. Scalco : R. E. Simoni : M. L. C. de Oliveira Laboratório de Erros Inatos do Metabolismo (LABEIM), Departamento de Bioquímica, Instituto de Química, Introduction Universidade Federal do Rio de Janeiro (UFRJ), Cidade Universitária, Ilha do Fundão, Centro de Tecnologia, Fumaric aciduria, an autosomal recessive disorder, is bloco A, 536 C, 21941 900 Rio de Janeiro, Brasil caused by deficiency of the enzyme fumarate hydratase e-mail: [email protected] [fumarase (FH)]. The first patient with : (OMIM 136850), presenting with lethargy, microcephaly, P. Correia J. C. Llerena Jr and hypotonia combined with high urinary fumaric acid Centro de Genética Médica, Instituto Fernandes Figueira, FIOCRUZ, excretion, was described by Zinn et al. (1986). Fumaric Rio de Janeiro, Brasil aciduria, however, had already been reported in 1983 by 412 J Inherit Metab Dis (2010) 33:411–419

Whelan et al in a couple of siblings (Whelan et al 1983). sutures, hooked nose, micrognathia, camptodactyly of the Fumaric aciduria is characterized by massive excretion of fifth finger, single transverse palmar crease on the left hand fumaric acid in urine. Other metabolites may also be and postaxial polydactyly on the right hand. Immediately altered in body fluids: Krebs cycle intermediates, other after birth and during the following 17 days, she presented dicarboxylic acids (suberic, adipic), and succinylpurine with hypoglycemic episodes difficult to control (: derivatives. The disease is considered rare, since in the 20–40 mg/dl). Laboratory exams, performed during one of last 27 years, only 40 cases have been reported in the these episodes, showed lactic acidemia (4.0 µmol/ml; N: literature. However, a higher frequency can be observed 0.7–2.5 µmol/ml), mild ,– and marked - at the border of northern Arizona and southern Utah, metabolic acidosis (pH 7.20; pCO2 25.1 mmHg; HCO3 USA. This is probably due to a founder effect in closed, 9.6 mEq/L; BE -17.5 mEq/L). Neurologic exam, per- religious polygamist communities living in these regions, formed at 4 days of life, showed incomplete Moro reflex, among which marriage between relatives seems to be posture of limbs flexion, reduced axial tonus, and frequent (Kerrigan et al 2000,Szep2007). Fumaric slightly elevated appendicular tonus. At 6 days of life, aciduria is classified into the group of disorders that antibiotics were administered for one week due to involve metabolic energy. In this group are included suspicion of sepsis. On the seventh and 25th days, two mitochondrial disorders that may interfere in embryophe- samples of urine and one of plasma were collected and tal development, leading to developmental delay, , sent for inborn errors of (IEM) screening. motor dysfunction, and alterations in other organ systems, analyses were carried out on a Biochrom 20 especially liver, kidney, and blood cells (Saudubray et al (Biochrom Ltd., Cambridge, UK) amino acid analyzer. 2006; Filiano 2006). Sample preparation and analysis were performed accord- FH (E.C. 4.2.1.2) catalyzes the stereospecific and ingtothemanufacturer’s standard protocols. General reversible hydration/dehydration of fumarate to S-malate. was observed and some plasma amino This enzyme is homotetrameric, iron-dependent, and acids were also altered (Table 1). Urinary organic acid thermostable. It exists in two isoforms, mitochondrial and analyses by gas chromatography mass spectrometry (GC- cytosolic, both being encoded by the same located on MS) (according to Tanaka et al 1980) showed massive 1q.42.1 (Estevez et al 2002, Remes et al excretion of fumaric acid (> 232 mmol/ mol , 2004). They only differ at the aminoterminal residue: normal range < 45 mmol/mol creatinine). The same result pyroglutamic acid on the mitochondrial isoform and N- was presented by a new urine sample collected at 40 days acetylalanine on the cytosolic one (Akiba et al 1984). In of life. fumarase deficiency, both isoforms are affected. It is At 1 month of age, despite breast feeding and normal supposed that the cytosolic isoform processes fumaric acid sucking, the baby had extreme difficulty gaining weight, from the cycle, amino acid, and , remaining undernourished due to persistent vomiting and whereas the mitochondrial isoform originates from the occasional diarrhea. Cholestatic jaundice and hepatomegaly Krebs cycle (Lehtonen 2006). were noted since birth. At 40 days of life, the following The aim of this study was to report the first case of hepatic were altered: [aspartate aminotransferase fumaric aciduria in Brazil and to review data from patients (AST) 137 U/L, N 10–34 U/L; aminotransferase found in the literature. (ALT) 78 U/L, N 5–35 U/L; alkaline leukocyte phosphatase (ALP) 267 U/L, N 20–140 U/L, and gamma- glutamyltranspeptidase (γ–GT) 505 U/L, N 0–51 U/L]. Case report Further exams at this period revealed colpocephaly [cranial computed tomography (CT)], infantile hypertrophic pyloric A girl, the fourth child of consanguineous parents (first stenosis [abdominal ultrasound (US)] and interatrial com- ), was born at 38 weeks of gestation after a munication (ECO). At one-and-a-half months of age, pregnancy complicated by polyhydramnios. Birth weight pyloromyotomy was performed, but vomiting episodes was 2180 g (< 3rd percentile), length 47 cm (∼10th persisted. At this time, she still presented with lactic percentile), head circumference 30 cm (< 3rd percentile), acidemia (5.0 µmol/ml), metabolic acidosis, and mild and APGAR score 8/9. One brother, born at term, and one hyperammonemia. Hemotransfusion was performed due to premature sister, had both died 6 h(presumed perinatal anemia. At 2 months and 4 days, hemoculture was positive asphyxia) and two days (cause not reported), respectively, for Staphylococcus epidermidis, and she was treated with after birth. Ultrasound images of the fourth child showed antibiotics for 14 days. There was a small weight gain, but agenesis of the corpus callosum and mild bilateral pyelo- hepatic enzymes remained altered. At 3 months and 5 days, caliceal dilatation. At physical examination, dysmorphic the girl was discharged but continued to gain weight slowly, features were noted, such as hypertelorism, overriding although being fed with breast milk and supplementary J Inherit Metab Dis (2010) 33:411–419 413

Table 1 Urinary and plasma amino acid analysis by ion-exchange chromatography of the patient here reported

Amino acid Urine 1 (nmol/mg creatinine) Urine 2 (nmol/mg creatinine) Reference valuesa (0 - 1m) Plasma 1 Reference valuesa (nmol/mg creatinine) (nmol/ml) (0-1m) (nmol/ml)

Threonine 10600 2262 124–1118 313 90–329 7350 3416 248–2898 263 99–395 Citrulline 5494 638 0–146 181 10–45 Methionine nd nd 104–875 70 10–60 4470 207 38–1373 99 29–132 5120 1540 81–923 430 376–709 Cystine 1359 1564 47–200 nd 17–98 539 177.5 28–402 66 48–160 Tyrosine 3690 351.5 133–1490 257 55–147 Ornithine 1238 925.7 15–411 77 48–211 3652 2683 218–1795 107 92–325 6427 2467 365–2267 130 30–138 nd not detected a Metabolic Laboratory, Children’s Medical Center, Dallas, Texas, USA formula. Vomiting episodes twice a day and occasional dehydration. Besides hepatosplenomegaly, she presented diarrhea persisted. At 4 months and 9 days, she was with severe metabolic acidosis (pH 6.45; pCO2 32 mmHg; – readmitted owing to vomiting with traces of blood, HCO3 2 mEq/L; BE –32 mEq/L) and bradycardia, which hyperpyrexia, intense crying and pallor, dyspnea, and evolved to cardiac arrest and death. It was not possible to

Table 2 Clinical features found in 36 patients with fumaric aciduria described in the literature and in the case presented here

Clinical findings Patients % Present Case References

Global developmental delay 34/36 94 + 1–20 Cerebral abnormalities 33/36 92 + 1–19 Hypotonia 26/36 72 + 2, 4–10, 12, 14–19 Dysmorphism 16/36 44 + 3, 9, 11–13, 16, 18, 19 Convulsive disorders a 16/36 44 – 4, 7–9, 13, 18, 19, 20 Microcephaly 13/36 36 + 2, 4–6, 8, 9, 15–17, 19 Feeding difficulties 12/36 33 – 2, 3, 6–10, 12, 14, 17 Failure to thrive 11/36 31 + 2, 3, 5–7, 9, 12, 14–16 Do not follow or fix 10/36 28 – 2, 8, 11, 14, 16 Hematologic alterations b 9/36 25 + 6, 11, 12, 16 Macrocephaly c 9/36 25 – 11, 18 Vomiting 8/36 22 + 1– 4, 6, 7, 10, 14 Lethargy 4/36 11 – 2, 7, 14 Gastrointestinal abnormalities d 4/36 +f 2, 11, 16 Gestational history Prematurity (33–36 weeks) 10/36 28 – 1, 4, 6, 7, 9, 13,15, 18 Polyhydramnios 5/36 14 + 3, 6, 13, 17 Others e 6/36 17 – 6, 7, 9, 16, 19

1: Whelan et al 1983; 2: Zinn et al 1986; 3: Walker et al 1989; 4: Gellera et al 1990; 5: Elpeleg et al 1992; 6: Bourgeron et al 1994; 7: Narayanan et al 1996; 8: Bonioli et al 1998; 9: Coughlin et al 1998; 10: Manning et al 2000; 11: Kerrigan et al 2000; 12: Zeman et al 2000; 13: Kimonis et al 2000; 14: Remes et al 1992 and 2004; 15: Loeffen et al 2005; 16: Phillips et al 2006; 17: Deschauer et al 2006; 18: Zeng et al 2006; 19: Maradin et al 2006; 20: De Meirleir et al 2006 a All types of convulsions included and two patients were not responsive to drug treatment, b Neonatal in five patients from the same family, c Only two families; d malrotation of the bowel and esophageal dysmotility; e reduced fetal movements, excessive fetal weight, bleeding, intrauterine growth retardation, and leakage of amniotic fluid, f infantile hypertrophic pyloric stenosis 414 J Inherit Metab Dis (2010) 33:411–419

Table 3 Cerebral abnormalities found in 33 patients with reported neuroimaging and in the case presented here

Cerebral abnormalities Patients Percent This case References

Ventriculomegaly/ Hydrocephaly 27/33 82 + 1–9, 11–16, 18, 19 Cerebral Atrophies 24/33 73 - 2–5, 7, 9–11, 13, 14, 16–19 Open Sylvian Operculum a 10/33 30 - 11, 16 Polymicrogyria a 9/33 27 - 11, 18 Small brain Stem b 8/33 24 - 11, 16 Agenesis/ Hypoplasia of corpus callosum 6/33 18 + 3, 7,9, 16, 19 Brain cysts c 6/33 18 - 3, 11, 15 Hypomyelination 5/33 15 - 4, 7, 8, 16 Others d 2/33 6 - 9, 12

Author’s references according Table 2. a : 8 from the same family; b : 6 from the same family; c : 3 choroid plexus cysts; 1 arachnoid cyst in the temporal fossa; 1 periventricular cysts; 1 cyst in occipital horn. d : Dandy-Walker malformation and lissencephaly measure enzymatic activity and perform molecular analysis, Clinical findings since no laboratory in Brazil executed these exams. Postmortem exam was not allowed by the parents. The literature group consisted of 40 patients from 27 families. Clinical and biochemical data were described in 36 patients, enzymatic activity determined in 31, and mutational data in Overview: clinical, laboratorial, and mutational features 26. Of these 36 patients, 20 were females and 16 males, and in fumarase-deficient patients their main clinical features and cerebral abnormalities are shown in Tables 2 and 3,respectively. Data from the literature were compiled, compared with the Cerebral abnormalities were observed in 33 patients and case here reported, and discussed. It is important to stress the three remaining were mentally impaired (Whelan et al that not all patients had complete description of clinical, 1983; Bourgeron et al 1994; De Meirleir et al 2006). These biochemical, and mutational data. malformations in fumaric aciduria could be a result of

Table 4 Dysmorphic features described in 15 of 16 patients Dysmorphic Features Patients Percent Percent total This case References and in the case presented here Depressed nasal bridge 12/15 80 33 – 11, 16, 18, 19 Frontal bossing 10/15 67 28 – 11, 12, 18 Hypertelorism 9/15 60 25 + 11, 19 Notched/anteverted naresa 5/15 33 14 – 11 Low-set/anteverted/prominent ears 4/15 27 11 – 12, 13, 18, 19 High-arched palatea 3/15 20 8 – 11 Bitemporal narrowing 3/15 20 8 – 13, 16 Overridging sutures 2/15 13 6 + 16 Metopic ridgingb 2/15 13 6 – 16 Triangular lower fasciesb 2/15 13 6 – 16 Brachycephaly b 2/15 13 6 – 16 Dolichocephaly 1/15 7 3 – 13 Micrognathia 1/15 7 3 + 12 Hypotelorism 1/15 7 3 – 13 Pointed chin 1/15 7 3 – 13 Camptodactyly 1/15 7 3 + 10 References according Table 2. Polydactyly ––– + – a Described in one family with Colpocephaly ––– + – eight affected members (Kerrigan Hooked nose ––– + – et al 2000), b monozygotic twins J Inherit Metab Dis (2010) 33:411–419 415

Table 5 Altered substances in physiological fluids reported in patients with fumaric aciduria

Altered organic acid Number of patients This case References

Urine Plasmaa CSFa

Fumaric 36/36 5 1 + 1–20 Succinic 6/36 –– 2–4, 7, 9, 12, 13 2-ketoglutaric 6/36 1 – 3, 4, 6, 13, 15 Adipic 2/36 –– 2, 4 Citric 2/36 –– 2, 7 Latic 3/36 4 5 +b 2–4, 6, 7, 8, 12, 17 Malic 3/36 –– 4, 8, 13 Pyruvic 2/36 3 2 4, 6–8, 15 Suberic 2/36 –– 2, 4 3-OH-butyric 1/36 –– 7 4-OH-phenylacetic 1/36 –– 7 Acetoacetic 1/36 2 – 6, 7 Methylmalonic 1/36 1 – 15

References according to Table 2 CSF cerebral spinal fluid a Organic acid analyses were not performed in all patients b plasma aerobic energy loss during early embryogenesis (Phillips et (Whelan et al 1983; Maradin et al 2006; Coughlin et al al 2006). Global developmental delay was present in almost 1998; Kerrigan et al 2000; De Meirleir et al 2006; Zeng et all patients reviewed in the literature. From the two al 2006). remaining patients, one had only seizures and the other Abnormal posture was seen in seven hypotonic patients, one died at 17 h of life (Coughlin et al 1998; Manning et al five showing opisthotonus. Two patients presented with 2000). Speech impairment was observed in 14 patients areflexia associated with hypotonia. (Narayanan et al 1996; (from six different families), some being unable to speak, Manning et al 2000). Other neurologic findings were some being dysarthric, and others restricted to few words bilateral Babinski sign, spasticity, choreoathetoid move-

ASL Fig. 1 Metabolic pathways SAICAR AICAR FAICAR IMP involving fumarate ASPARTATE + GTP

GDP + Pi MALEYLACETO- MAI FUMARYL ACETATE ACETOACETATE ASL AMP ADENYLOSUCCINATE

HGO FAH O2 ACETO- HOMOGENTISATE ACETATE SCS CO2 FH S-MALATE FUMARATE SUCCINATE SUCCINYL-CoA 4HPPD CO ME SDH 2

O2 2-CGDH PYRUVATE p–OH–PHENYLPYRUVATE -KETOGLUTARATE + CO2 FUMARATE CO2 ICDH GLUCOSE TAT LACTATE FH KREBS AS ISOCITRATE AC TYROSINE ARGININO CYCLE SUCCINATE CITRATE PYRUVATE - PDHC ATP + HCO 3 UREA S-MALATE PC H2O CS ASS ACETIL-CoA ADP + Pi CYCLE ME MDH UREA OXALOACETATE ASPARTATE PYRUVATE CITRULLINE ORNITHINE + CO2 OT -KETOGLUTARATE AMINO ACID

AST GD AMINOTRANSFERASE

GLUTAMATE -KETO ACID

NH 3

CARBAMOYL - HCO 3 + ATP PHOSPHATE CPSI ADP 416 J Inherit Metab Dis (2010) 33:411–419 ments, ataxia, and dystonic tetraplegia (Walker et al 1989; dramnios are a consequence of disorders that affect Narayanan et al 1996; Kerrigan et al 2000; Manning et al gastrointestinal, musculoskeletal and the central nervous 2000; Remes et al 2004; Loeffen et al 2005; Maradin et al system (CNS) (Kyle et al. 1997). Angulation of the frontal 2006). Of 11 patients with reported EEG, ten showed horns and optic nerve hypoplasia/pallor were described in a epileptiform features (Zinn et al 1986; Gellera et al 1990; consanguineous family that had eight affected members; Elpeleg et al 1992; Remes et al 1992; Bonioli et al 1998; these features are unique in study group (Kerrigan et al Kimonis et al. 2000; Loeffen et al 2005; Deschauer et al 2000). By the time of their respective reports, ten patients 2006; Phillips et al 2006). Sixteen patients from eight had died early (28%), seven of them being younger than different families had some degree of dysmorphia, and 1 year, such as the case here reported. The cause of craniofacial dysmorphic features were present in all of them deathwasdescribedonlyinsevenpatientsandingeneral (Table 4). It must be observed that in disorders with was due to infections (3), cardiac arrest (3), or prolonged abnormalities in energy production, dysmorphic features apnea (1). may occur. However, the underlying mechanism in fumaric aciduria is unknown (Kerrigan et al 2000; Peters and Kahler Laboratorial findings 2003). Prematurity was a relevant gestational feature, have Metabolite alterations in physiological fluids reported in the being associated with polyhydramnios in two cases. In literature in patients with fumaric aciduria are described in general, the main fetal anomalies associated with polyhy- Table 5.

Table 6 Mutations found in 25 patients with fumarase deficiency (adapted from Bayley et al 2008)

Original Original mutation description DNA change* Protein mutation* Number reference of patients DNA Protein

1, 3, 8, 9, 11 1302insAAA,1433insAAA, 434insLys, 435insLys, c.1431_1433dup p.Lys477dup 7/26 AAAins435 477insLys 5, 7, 10 998A>C, 1127A>C Gln13Pro, Gln333Pro, c.1127A>C p.Gln376Pro 5/26 a,b Gln376Pro 6, 9, 12 392C>G, 492C>G Pro131Arg c.521C>G p.Pro174Arg 3/26 e 10 1078C>T His360Cys c.1207C>T p.His402Cys 2/26 a 2 955G>C Glu319Gln c.1084G>C p.Glu362Gln 2/26 c 8, 9 698G>A Arg190His, Arg233His c.698G>A p.Arg233His 2/26 3 1148A>T Asp383Val c.1274A>T p.Asp425Val 2/26 d 3, 9 560A>G Lys187Arg c.689A>G p.Lys230Arg 2/26 6 779T>C Leu260Ser – p.Leu303Ser 1/26 9 2-slice exon 2 – Exon 2 splice variant – 1/26 3 66del74 – c.195_268del p.Lys66fs 1/26 3 793G>A Ala265Thr c.922G>A p.Ala308Thr 1/26 3 806T>G Phe269Cys c.935T>G p.Phe312Cys 1/26 11 824A>T His275Leu c.953A>T p.His318Leu 1/26 13 976C>T Pro326Ser c.1105C>T p.Pro369Ser 1/26 9 – Arg58X c.301C>T p.Arg101X 1/26 1 – Arg190Cys c.697C>T p.Arg233Cys 1/26 13 1029_1031delAGT Gln343His/ 344delVal c.1158_1060delAGT p.Gln386_ 1/26 Val387delinsHis 3 1373G>A Trp458X c.1499G>A p.Trp500X 1/26 14 1078C>T – c.1204C>T p.His402Tyr 1/26 14 Intronic mutation c.1391-269A>G –––1/26

1: Gellera et al 1994; 2: Bourgeron et al 1994; 3: Coughlin et al 1998; 4: Manning et al 2000; 5: Zeman et al 2000; 6: Kimonis et al 2000; 7: Remes et al 2004; 8: Loeffen et al 2005; 9: Pollard et al 2005; 10: Phillips et al 2006; 11: Deschauer et al 2006; 12: Zeng et al 2006; 13: Maradin et al 2006; 14: De Meirleir et al 2006 * Some of the reported FH mutations were traditionally based on the mature protein rather than the primary translation product, which also contains the signal peptide of 43 amino acids, for mitochondrial signaling. They were corrected for the mitochondrial isoform, considered the canonic form,a monozygotic twins with compound heterozygous mutation; b,c,d siblings; e one case of paternal partial isodisomy J Inherit Metab Dis (2010) 33:411–419 417

The massive excretion of fumaric acid in urine is amount of free adenosine monophosphate (AMP) (Toth and common to all patients (15- to 1,000-fold higher when Yeates 2000;Tsaietal2007). In purine de novo , compared with normal range). In one patient, fumaric acid adenylosuccinate lyase (ASL, E.C. 4.3.2.2) catalyzes two elevation was not observed with the first symptoms but only similar reactions producing 5-aminoimidazole-4-carboxamide appeared later (Narayanan et al 1996). Some considerations ribotide (AICAR), AMP,and fumaric acid (Fig. 1). ASL could regarding other metabolite changes in patients with fumaric be affected by the increased amount of fumaric acid, which is aciduria are presented below. Other organic acids were also a weak and noncompetitive inhibitor of this enzyme (Barnes shown to be altered in fumaric aciduria, with urinary α- and Bishop 1975). In one patient, two succinylpurine ketoglutaric and succinic acids being the most relevant derivatives, 5-aminoimidazole-(N-succinylcarboxiamide) besides fumaric acid (Table 5). These findings were probably ribotide (SAICAR) and adenylosuccinate, were found in due to secondary enzymatic inhibition of succinate dehydro- cerebral spinal fluid (CSF). In urine of two patients, these genase (SDH). This enzyme belongs to complex II and is derivatives were not detected, as such metabolites do not part of the respiratory chain and Krebs cycle; it catalyzes the cross the blood-brain barrier (Elpeleg et al 1992, Zeman conversion of to fumaric acid (Fig. 1). The et al 2000). Zeman and coworkers considered that the reverse reaction is barely detectable in human cells and accumulation of these succinylpurine derivatives could tissues in normal conditions (Brière et al 2005). have a profound effect on impairment of the CNS in An unexpected finding from the literature was the fumaric aciduria (Zeman et al 2000). SAICAR, which is a presence of in three patients. This could be substrate for ASL, was considered a potential neurotoxic explained by the accumulation of acetyl- agent to pyramidal neurons of rat hippocampus (Stone et (CoA) in mitochondria that inhibits pyruvate dehydroge- al 1997). nase complex (PDHC) and stimulates pyruvate carboxylase From 12 literature cases in which amino acid analysis (PC). The result is increased production of , was performed, three patients, two being siblings, showed which is converted to malic acid by malate dehydrogenase altered results. Bourgeron and coworkers reported that (MDH) (Gellera et al 1990). Another alternative pathway both siblings had slightly elevated phosphoethanolamine, would be the mitochondrial and cytosolic malic enzyme , alanine, , serine, and in blood and converting to malic acid. This bidirectional urine. Aspartate and asparagine were low in these two enzyme is highly stimulated when the mitochondrial levels patients’ blood and elevated in urine of one of them of fumaric and succinic acids are increased (Taroni et al (Bourgeron et al 1994). In the third patient, only urinary 1988) (Fig. 1). Accumulation of fumaric acid due to FH amino acids were altered, but these were not discriminated deficiency may affect several other metabolic pathways. (Narayanan et al 1996). The patient here reported showed The purine nucleotide cycle, necessary for cellular replication, an increased urinary excretion of several amino acids aids in tight regulation of cellular metabolism by controlling (Table 1). Another interesting finding was the elevated the amounts of available cycle intermediates and the levels of tyrosine, methionine, and citrulline in plasma not

Fig. 2 Mutations observed on 10 20 30 40 50 60 fumarate hydratase enzyme MYRALRLLAR SRPLVRAPAA ALASAPGLGG AAVPSFWPPN AARMASQNSF RIEYDTFGEL (mitochondrial isoform) 70 80 90 100 110 120 KVPNDKYYGA QTVRSTMNFK IGGVTERMPT PVIKAFGILK RAAAEVNQDY GLDPKIANAI

130 140 150 160 170 180 MKAADEVAEG KLNDHFPLVV WQTGSGTQTN MNVNEVISNR AIEMLGGELG SKIPVHPNDH

190 200 210 220 230 240 VNKSQSSNDT FPTAMHIAAA IEVHEVLLPG LQKLHDALDA KSKEFAQIIK IGRTHTQDAV

250 260 270 280 290 300 PLTLGQEFSG YVQQVKYAMT RIKAAMPRIY ELAAGGTAVG TGLNTRIGFA EKVAAKVAAL

310 320 330 340 350 360 TGLPFVTAPN KFEALAAHDA LVELSGAMNT TACSLMKIAN DIRFLGSGPR SGLGELILPE

370 380 390 400 410 420 NEPGSSIMPG KVNPTQCEAM TMVAAQVMGN HVAVTVGGSN GHFELNVFKP MMIKNVLHSA

430 440 450 460 470 480 RLLGDASVSF TENCVVGIQA NTERINKLMN ESLMLVTALN PHIGYDKAAK IAKTAHKNGS

490 500 510 TLKETAIELG YLTAEQFDEW VKPKDMLGPK 418 J Inherit Metab Dis (2010) 33:411–419 previously reported in the literature. Elevated tyrosine and Final considerations methionine levels could be due to the secondary inhibition of the last enzyme of the tyrosine catabolic pathway, It must be considered that some of the unusual findings fumarylacetoacetate hydrolase (FAH, E.C.3.7.1.2.), by of the patient here described could also be due to another increased levels of fumaric acid (Fig. 1). This inhibition genetic defect and not exclusively to fumaric aciduria, could also result in elevation of several hepato- and since the parents are consanguineous. Fumaric aciduria neurotoxic metabolites, such as observed in has a broad spectrum of clinical manifestations, although type I. The main features of this disease, renal Fanconi the great majority of patients present with global syndrome, and hepatic problems (hepatomegaly, jaundice, developmental delay and cerebral abnormalities. Bio- and cholestasis) (Mitchell et al 2001), were also presented chemically, however, elevated excretion of fumaric acid by the patient here reported. Hepatomegaly was described has been found in all patients, being therefore considered in two literature patients, and one of them also presented a hallmark of this disorder. The significant accumulation with jaundice and cholestasis (Walker et al 1989; Zeman of fumaric acid due to FH deficiency may affect several et al 2000). Another consequence of fumaric acid metabolic pathways, including de novo purine nucleotide accumulation could be a diminished activity of arginino- biosynthesis, tyrosine catabolic pathway, , and succinase (AS, E.C. 4.3.2.1), which would explain the . Some possible explanations, regarding significant plasma citrulline elevation (Fig. 1). Some clinical features and abnormal metabolite levels in fumaric symptoms, such as hyperammonemia and vomiting, aciduria patients, are presented. common to urea cycle defects, were presented by the An interesting finding observed in the patient here patient. Hyperammonemia wasreportedonlyintwo reported was the alteration of several plasma and patients from the literature (Zinn et al 1986,Walkeretal urinary amino acid levels. Such changes allowed a few 1989). Severe metabolic acidosis was present in the pertinent considerations that may expand the current patientdescribedherefrombirthandinonlytwo knowledge of this rare disease. Although organic acid literature patients (Gellera et al. 1990, Narayanan et al analysis is the first and main step to be carried out in 1996). The measured residual FH activity in the literature patients suspected of having fumaric aciduria, it is patients (n=31) ranged from not detectable to 36% of suggested that amino acid analysis in body fluids be control mean (average ∼5%). Heterozygous parents had also performed. Moreover, it would be interesting to the activity ranging from 30% to 74% of the control mean. consider the hepatic picture owing to the signs de- However, it has been observed that fumaric acid excretion scribedinthecasepresentedhere. is not correlated with residual enzyme activity or severity of the clinical picture (Bonioli et al 1998). Acknowledgments We thank Francisco Radler de Aquino Neto, Luis Nelson L F Gomes, DLE (Diagnósticos Laboratoriais Especializados), Mutational findings Markéta Tesařová and Jean-Pierre Bayley for their contribution to the study. Human FH gene consists of ten exons encoding 510 amino acids. The first exon (exon 0) encodes a mitochondrial localization signal peptide of 43 amino References acids (UniProt, 2009). The mutations related to FH deficiency reported in the literature so far are shown in Akiba T, Hiraga K, Tuboi S (1984) Intracellular distribution of fumarase in various animals. J Biochem 26:189–195 Table 6 (modified from Bayley et al 2008). It can be Barnes LB, Bishop SH (1975) Adenylosuccinate lyase from human observed that FH genotype is very heterogeneous, and erythrocytes. Int J Biochem 6:497–503 most mutations are concentrated at the C-terminus of the Bayley JP, Launonen V, Tomlinson IPM (2008) The FH mutation FH enzyme (Fig. 2). Twenty-one different mutations database: an online database of fumarate hydratase mutations involved in the MCUL (HLRCC) tumor syndrome and congenital related with congenital fumarase deficiency were de- fumarase deficiency. BMC Med Genet 9:20 scribed. In two patients, only one allele mutation was Bonioli E, Di Stefano A, Peri V et al (1998) Fumarate hydratase found at the time of their reports (Coughlin et al 1998). deficiency. J Inherit Metab Dis 21:435–436 Fourteen mutations were missense. The remaining muta- Bourgeron T, Chretien D, Poggi-Bach J et al (1994) Mutation of fumarase gene in two sibilings with progressive tions were nonsense, duplication, frameshift, splice site, and fumarase deficiency. J Clin Invest 93:2514–2518 and deletion/insertion. Twelve patients were homozygous Brière JJ, Favier J, El Ghouzzi V, Djouadi F et al (2005) Succinate for the mutations Gln376Pro, Pro174Arg, Glu362Gln, dehydrogenase deficiency in human. CMLS, Cell Mol Life Sci – Asp425Val, and Lys230Arg. Mutation p.Lys477dup was 62:2317 2324 Coughlin EM, Christensen E, Kunz PL et al (1998) Molecular the most frequent, being observed in seven heterozygous Analysis and prenatal diagnosis of human fumarase deficiency. nonrelated patients (27%). 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