Coenzyme Q– Responsive Leigh’s Encephalopathy in Two Sisters Lionel Van Maldergem, MD,1 Frans Trijbels, PhD,2 Salvatore DiMauro, MD,3 Pavel J. Sindelar, PhD,4 Olimpia Musumeci, MD,3 Antoon Janssen, MD,2 Xavier Delberghe, MD,5 Jean-Jacques Martin, MD, PhD,6 and Yves Gillerot, MD, PhD1 A 31-year-old woman had encephalopathy, growth retardation, infantilism, ataxia, deafness, lactic acidosis, and increased signals of caudate and putamen on brain magnetic resonance imaging. Muscle biochemistry showed succinate:cytochrome c oxidoreductase (complex II–III) deficiency. Both clinical and biochemical abnormalities improved remarkably with coenzyme Q10 supplementation. Clinically, when taking 300mg coenzyme Q10 per day, she resumed walking, gained weight, underwent puberty, and grew 20cm between 24 and 29 years of age. Coenzyme Q10 was markedly decreased in cerebrospinal fluid, muscle, lymphoblasts, and fibroblasts, suggesting the diagnosis of primary coenzyme Q10 deficiency. An older sister has similar clinical course and biochemical abnormalities. These findings suggest that coenzyme Q10 deficiency can present as adult Leigh’s syndrome. Ann Neurol 2002;52:750 –754 Leigh’s syndrome (LS; OMIM 256000) is a genetically heterogeneous condition characterized by developmental delay or regression, lactic acidosis, and symmetrical lesions in the basal ganglia,1 which are considered the neuroradiological or neuropathological signature of LS.2,3 Primary coenzyme Q10 (CoQ10) deficiency causes a mitochondrial encephalomyopathy that can be dominated by myopathic or cerebellar involvement and is responsive to CoQ10 administration.4 – 8 Here, we describe two sisters with LS and widespread CoQ10 deficiency, who responded drastically to CoQ supplementation. Patients and Methods Patient 1 The proposita was born at term after an uneventful pregnancy. There was no consanguinity and no history of neuromuscular disorders in the family. At 6 months, she was noted to have axial hypotonia and failure to thrive. At 19 months, her weight was 8,700gm (⫺2.5 standard deviation [SD]), and her height was 79cm (25th percentile). She had unsteady gait, bilateral ptosis, and dysmorphic features, including narrow bifrontal diameter, flat philtrum, and thin upper vermilion border. Visual contact was poor and funduscopy revealed tortuous retinal vessels. Plasma lactic acid was elevated (3.8mmol/L) whereas pyruvic acid was normal (0.47mmol/L). A diagnosis of LS was sug- From the 1Centre de Génétique Humaine, Institut de Pathologie et de Génétique, Loverval, Belgium; 2Center for Mitochondrial Disorders, Clinical Genetics Center, University of Nijmegen, Nijmegen, The Netherlands; 3Columbia University College of Physicians and Surgeons, Department of Neurology H. Houston Merritt Clinical Research Center, New York, NY; 4Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden; 5Institut Medico-Pedagogique Montfort, Herseaux, Belgium; and 6Department of Neuropathology, Born-Bunge Foundation, University of Antwerp, Antwerp, Edegem, Belgium. 750 © 2002 Wiley-Liss, Inc. gested. At age 4 years, she had marked muscle wasting and moderate spasticity, but she was still ambulant with the aid of orthopedic shoes for calcaneovalgus deformity. She became deaf and failed to undergo puberty. At age 19 years, she had ataxia with wide-base gait and slow, uncoordinated movements. Moderate spasticity of lower limbs, brisk knee jerks, atrophy of leg muscles, and distal contractures were suggestive of a pyramidal syndrome. A quadriceps muscle biopsy showed fiber-type disproportion but no lipid storage or ragged-red fibers. The activities of pyruvate dehydrogenase, PC, cytochrome oxydase (COX), succinate–cytochrome c reductase, and citrate synthase and lactate to pyruvate ratio were normal in cultured skin fibroblasts (B. H. Robinson, Hospital for Sick Children, Toronto, Ontario, Canada; C. Marsac, Paris, France, data not shown). The only abnormality in fibroblasts was a mild complex I deficiency (L. De Meirleir, University of Brussels, Belgium). No mtDNA point mutation was detected. A brain magnetic resonance imaging showed symmetrical bilateral T2 hypersignals in the caudate and putamen. In addition, there was a white matter hypersignal in the left frontal subcortical region (Fig 1). At age 24 years, her general condition worsened, she refused feeding, became less social, developed generalized muscle hypertonia and trismus, and lost 2kg in 3 months. She became bedridden. No infections or nutritional defects were detected to explain the global regression observed. She was given Received Aug 10, 2001, and in revised form Apr 29 and Jul 25, 2002. Accepted for publication Jul 27, 2002. Address correspondence to Dr Van Maldergem, Centre de Génétique Humaine, Institut de Pathologie et de Génétique, Allée des Templiers 41, B-6280 Loverval, Belgium. E-mail: firstname.lastname@example.org Table 1. Biochemical Measurements in a Frozen Muscle Specimen from Patient 1 before CoQ10 Supplementation Enzyme activities in 600gm supplementation (mU.mU COX⫺1) NADH: Q1 oxidoreductase (complex I) Decylubiquinol: cytochrome c oxidoreductase (complex III) Cytochrome c oxidase (complex IV ⫽ COX) (mU.mg protein⫺1) Succinate: cytochrome c oxidoreductase (SCC) (mU.mg protein⫺1) Succinate: cytochrome c oxidoreductase ⫹ decylubiquinone (mU.mg protein⫺1) SCC: stimulation by decylubiquinone: 8.0 ⫻ (n ⬍ 3) Patient 498 Control Range 101–389 n.d. 1,020–2,530 81.4 33–225 4.3 8–44 34.6 CoQ10 ⫽ coenzyme Q10; NADH ⫽ nicotinamide adenine dinucleotide; n.d. ⫽ not determined. Fig 1. Magnetic resonance imaging brain imaging of Patient 2 at 20 years. Putamen and caudate T2 hypersignals are evident. There is also a small frontal white matter hypersignal on the left. 300mg CoQ10 (Medgenix, Wevelgem, Belgium) per day. Within 15 days, both trismus and hypertonia decreased, and she resumed eating and recovered her ability to walk. She gained 7.6kg in 6 months and grew 20cm between 24 and 29 years of age. Puberty occurred at 26 years. A psychological assessment at age 29 years showed improved social interaction. She was better coordinated and fell less frequently. A muscle specimen taken before CoQ10 supplementation and stored at ⫺80°C then was analyzed for respiratory chain activities and for CoQ content. It showed decreased complex II ⫹ III (succinate–cytochrome c reductase) activity, which increased eightfold after in vitro addition of CoQ10. The content of CoQ10 was markedly decreased (Table 1). Patient 2 The older sister of our index patient was born after normal pregnancy and delivery, but at 1 month of age, she developed chronic diarrhea, axial hypotonia, limb hypertonia, and failure to thrive. At age 6 months, she had poor visual contact and upper limb dystonia. She started walking at 18 months but failed to acquire any language skills. At 3 years of age, she was small (height, 92.5cm; weight, 11.100gm) and had spasticity and choreoathetotic movements. Serum lactic acid was 8mmol/L (normal, ⬍2mmol/L) and pyruvic acid was 2mmol/L (normal, ⬍0.67mmol/L). Fundoscopy showed tortuous and enlarged retinal vessels. Electroencephalogram and alanine loading test were unremarkable. Pyruvate dehydrogenase and pyruvate carboxylase activities in a liver biopsy were normal (C. Marsac, Faculté de Medecine Necker, Paris, France). The clinical course was characterized by marked growth delay, severe mental retardation, ataxia, and deafness. At the age of 7 years, her bone-age was that of a 4-year-old child and she had persistent lactic acidosis. She was placed in an institution for the mentally retarded. Pyramidal and extrapyramidal symptoms worsened and she became wheelchair bound at age 15 years. At age 21 years, she had permanent dystonic movements, inappropriate laughing, and mild facial dysmorphism, with a narrow forehead and flat philtrum (Fig 2). Deep tendon reflexes were brisk and there was generalized muscle atrophy. Height was 131cm (⫺5 SD) and weight 29kg (⫺5 SD). Lactic acid was 6.47mmol/L in plasma and 3.9mmol/L in the cerebrospinal fluid (normal, ⬍2.5mmol/L). A quadriceps muscle biopsy showed selective hypotrophy of type 2 fibers, normal amounts of subsarcolemmal mitochondria (oxidative enzymes stainings), and no ragged-red fibers. Urinary organic acids showed large amounts of lactic, pyruvic, parahydroxyphenyllactic, and parahydroxyphenylpyruvic acids. There was a generalized increase in plasma amino acids, including alanine. Echocardiography and eye slit-lamp examinations were normal. Fundoscopy showed tortuous retinal vessels (Fig 3). At age 31 years, CoQ10 was started (300mg/day) but discontinued after 6 weeks for unknown reasons. A muscle biopsy showed fiber type disproportion without ragged-red fiber. Biochemical analysis showed a mild decrease in oxidation rates and in complexes I, III, and IV (Table 2). Respiratory chain complexes were normal in Epstein–Barr virus–transformed lymphoblasts (P. Rustin, INSERM, U393, Hospital Necker– Enfants Malades, Paris, France; data not shown). When her sister benefited from CoQ10 therapy, she also received CoQ10 (300mg/day) and improved in both growth parameters and behavior. Methods The activities of respiratory chain enzymes in muscle were measured by standard methods.9,10 CoQ10 in muscle and lymphoblasts was measured by the method of Lang and colleages11 and Podda.12 Van Maldergem et al: Leigh’s Encephalopathy 751 Fig 3. Eye fundus of Patient 2 at 28 years. There is marked tortuosity and enlargement of retinal vessels. Fig 2. (a) Facial appearances of the two patients (at the ages of 19 and 21 years): note gelastic facies, ptosis of the eyelids, thin vermilion border, and flat philtrum. (b) Patients 1 (right) and 2 (left) at the ages of 28 and 31 years. Note the growth of the younger sister after 4 years of coenzyme Q supplementation. Infantilism and short stature have regressed. 752 Annals of Neurology Vol 52 No 6 December 2002 Results Biochemical assays in the index patient were performed in a frozen muscle sample stored before CoQ supplementation. The activities of complex I and complex IV were normal, but the activity of succinate: cytochrome c oxidoreductase (a reaction including complex II, CoQ10, and complex III) was markedly decreased. Addition of exogenous decylubiquinone to the incubation mixture stimulated complex II ⫹ III activity eightfold (normal stimulation factor, 0.9 –1.3). This finding suggested CoQ10 deficiency. In the older sister, biochemical analyses were performed in muscle obtained during CoQ supplementation (see Table 2). The oxidation rates of several radiolabeled substrates (pyruvate, malate, and succinate) measured in a 600g supernatant from a fresh muscle specimen were moderately decreased. The production rate of ATP ⫹ PCr from pyruvate oxidation was decreased. Measurements of pyruvate dehydrogenase complex and of respiratory chain enzymes showed mildly reduced activities of complex I, II and IV. The concentration of CoQ10 in muscle obtained from the proposita before CoQ10 supplementation was approximately 5% of the normal mean (Table 3). CoQ10 content was decreased by 40% in Patient 2 Table 2. Biochemical Measurements in a Fresh Muscle Specimen from Patient 2 during CoQ10 Supplementation Rate Oxidation Rates (nmol 14CO2 䡠 hr⫺1 䡠 mU CS⫺1) [1-14C]pyruvate ⫹ malate [U-14C]malate ⫹ pyruvate ⫹ malonate [1,4-14C]succinate ⫹ acetylcarnitine Production rate of ATP ⫹ PCr from pyruvate (nmol 䡠 hr⫺1 䡠 mU CS⫺1) Enzyme activities in 600g supernatant (mU 䡠 mU CS⫺1) NADH: Q1 oxidoreductase (complex I) Decylubiquinol: cytochrome c oxidoreductase (complex III) Cytochrome c oxidase (complex IV) Succinate: cytochrome c oxidoreductase (SCC) Pyruvate dehydrogenase complex (PDHc) Citrate synthase (CS) (mU.mg protein⫺1) SCC: stimulation by decylubiquinone: 1.1⫻ (n ⬍ 3) even during CoQ10 supplementation. The concentration of ␣-tocopherol also was severely decreased (20% of control). The CoQ10 content in immortalized lymphoblasts from Patients 1 and 2 was decreased to 50% (see Table 3). Discussion LS is caused by a variety of inborn errors of oxidative metabolism resulting in congenital lactic acidosis and severe encephalopathy with symmetrical bilateral basal ganglia hypersignals on brain magnetic resonance imaging. Biochemical causes include pyruvate dehydrogenase, pyruvate carboxylase (PC), and complex I, II, IV, and V deficiencies, but it is likely that other biochemical defects may be involved. Recent advances in our understanding of complex IV (COX) deficiency LS illustrate this expanding spectrum of mitochondrial cytopathies. COX deficiency has been attributed to mutations in at least four genes controlling COX assembly, whereas mutations in nuclear genes encoding COX subunits have not yet been identified.13–16 Primary CoQ deficiency has been reported in a limited number of patients and is clinically heterogeneous, with predominant involvement of skeletal muscle4,5,8 or the central nervous system.6,7 Anecdotal benefit from CoQ10 administration has been reported in various mitochondrial cytopathies,4,17–24 although muscle CoQ10 was only mildly decreased in these disorders.25,26 The sisters presented here have unusual clinical presentations and remarkable responsiveness to oral CoQ supplementation. Although the index patient had the typical neuroradiological features of LS, her clinical phenotype was unusual because of long survival and coexistence of extrapyramidal and pyramidal signs. Other symptoms, including cerebellar ataxia, deafness, and ptosis, were more typical of mitochondrial encephalomyopathies. However, biochemical studies documented CoQ10 deficiency. Enzyme assays in frozen muscle extracts showed complex II ⫹ III deficiency, which was corrected by ad- Patient Control Range 1.86 1.85 1.13 25.1 3.61–7.48 (n⫽20) 4.68–9.62 (n⫽20) 2.54–6.39 (n⫽19) 42.1–81.2 (n⫽16) 0.05 2.17 0.56 0.43 0.06 52.7 0.07–0.25 (n⫽34) 2.50–6.61 (n⫽21) 0.81–3.12 (n⫽21) 0.30–0.97 (n⫽39) 0.034–0.122 (n⫽25) 37.4–162 (n⫽43) Table 3. Concentrations of Coenzyme Q10 and ␣-Tocopherol in Muscle and Cultured Lymphoid Cell Lines (values are the means of three determinations) Measurement Location Muscle Control Older sister Index patient Lymphoblastoid cell lines Control Older sister Index patient CoQ10 ␣-Tocopherol (g/gm protein) (g/gm protein) 793 493 43 122 25 NA 401 198 173 NA NA NA NA ⫽ not applicable. dition of decylubiquinone in vitro. Direct measurement of CoQ10 in muscle homogenate confirmed profound deficiency (5% of normal). In the older sister, biochemical studies of fresh muscle obtained during CoQ10 supplementation did not show decreased activity of complex II–III, which is CoQ10 dependent. Her muscle CoQ10 was approximately 60% of normal despite CoQ10 supplementation (300mg/day for 30 days). Because the patient had shown striking improvement with CoQ10 supplementation, it is plausible to assume that the CoQ content in muscle must have been even lower before treatment. In this context, it is of interest that immortalized lymphoblasts from both patients had approximately 50% decrease in CoQ10. Because these cells were grown in vitro for several passages, these CoQ10 levels could not have been influenced by CoQ10 administration in vivo. Patient 1 also showed a severe decrease in ␣-tocopherol. In vitro experiments showed that the lipid-soluble antioxidants CoQ10 and ␣-tocopherol act synergistically in protecting membranes from reactive oxygen species.27 Other studies have shown parallel changes in the concentrations of CoQ10 and ␣-tocopherol. The striking growth spurt in the index patient at the age of 24 years can be explained by her prepubertal status when CoQ10 supplementation was started. Van Maldergem et al: Leigh’s Encephalopathy 753 Of the 12 cases of primary CoQ10 deficiency reported in the literature, 10 were associated with involvement of the central nervous system. Of these, six had cerebellar ataxia,24 whereas three siblings had progressive encephalopathy (mental retardation, retinitis pigmentosa with visual deficiency and deafness), hypertrophic cardiomyopathy, and a nephrotic syndrome.23 Ataxia was also seen in our patients, but magnetic resonance imaging showed basal ganglia involvement rather than cerebellar atrophy, setting them aside from the patients with the cerebellar presentation of CoQ10 deficiency. The lack of retinitis pigmentosa, cardiopathy, and renal failure distinguishes also our patients from those reported by Rötig and colleagues.6 This wide variety of clinical presentations in primary CoQ deficiency is suggestive of genetic heterogeneity that is likely related to the many enzymatic steps involved in the biosynthesis of CoQ.27,28 Our results suggest that LS also may be caused by CoQ10 deficiency. The similar clinical presentation (and response to CoQ10) in two siblings is compatible with autosomal recessive inheritance. 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