Isolated Mitochondrial Myopathy Associated with Muscle Coenzyme Q10 Deficiency

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OBSERVATION Isolated Mitochondrial Myopathy Associated With Muscle Coenzyme Q10 Deficiency

Seema R. Lalani, MD; Georgirene D. Vladutiu, PhD; Katie Plunkett, MS; Timothy E. Lotze, MD; Adekunle M. Adesina, MD, PhD; Fernando Scaglia, MD

Background: Primary coenzyme Q10 (CoQ10) defi- partial reductions in complex I, I + III, and II + III ac- ciency is rare. The encephalomyopathic form, de- tivities with greater than 200% of normal citrate syn- scribed in few families, is characterized by exercise in- thase activity. The CoQ10 concentration in skeletal muscle tolerance, recurrent myoglobinuria, developmental delay, was 46% of the normal reference mean. The in vitro ad- ataxia, and seizures. dition of 50 µmol/L of coenzyme Q1 to the succinate cytochrome-c reductase assay of the patient’s skeletal muscle Objective: To report a rare manifestation of CoQ10 de- whole homogenate increased the succinate cyto- ficiency with isolated mitochondrial myopathy without chrome-c reductase activity 8-fold compared with 2.8- central nervous system involvement. fold in the normal control homogenates. Follow-up of the patient in 6 months demonstrated significant clini- Methods: The patient was evaluated for progressive cal improvement with normalization of creatine kinase muscle weakness. Comprehensive clinical evaluation and and lactate levels. muscle biopsy were performed for histopathologic analysis and mitochondrial DNA and respiratory chain en- Conclusions: The absence of central nervous system in- zyme studies. The patient began taking 150 mg/d of a volvement and recurrent myoglobinuria expands the clini- CoQ10 supplement. cal phenotype of this treatable mitochondrial disorder. The complete recovery of myopathy with exogenous Results: The elevated creatine kinase and lactate levels CoQ10 supplementation observed in this patient high- with abnormal urine organic acid and acylcarnitine pro- lights the importance of early identification and treat- files in this patient suggested a mitochondrial disorder. ment of this genetic disorder. Skeletal muscle histochemical evaluation revealed ragged red fibers, and respiratory chain enzyme analyses showed Arch Neurol. 2005;62:317-320

RIMARY COENZYME Q 10 ment and normalization of serum creatine (CoQ10) deficiency (Mende- kinase and lactate values. This report ex- lian Inheritance in Man tends the clinical spectrum of CoQ10 de- 607426) is rare and is char- ficiency to include isolated primary my- acterized by significant clini- opathy without ataxia, seizures, or cal heterogeneity. The clinical spectrum cognitive impairment. P 1-4 varies from encephalomyopathy, familial cerebellar ataxia,5 and Leigh encepha- lopathy6 to widespread multisystem dis- REPORT OF A CASE ease.7 The encephalomyopathic form, Author Affiliations: described in 4 families,1-4 is characterized The patient was initially evaluated at 11.5 Departments of Molecular and by exercise intolerance, recurrent myo- years of age for progressive muscle weak- Human Genetics (Drs Lalani globinuria, developmental delay, ataxia, ness. He was a previously healthy, devel- and Scaglia and Ms Plunkett), and seizures. Herein, we describe a pa- opmentally normal child, born at 34 weeks’ Neurology (Dr Lotze), and tient with exercise intolerance, ragged red gestation to healthy, nonconsanguine- Pathology (Dr Adesina), Baylor fibers, muscle CoQ deficiency, and as- ous parents. Insidious onset of exercise in- College of Medicine, Houston, 10 Tex; and Departments of sociated muscle carnitine deficiency with tolerance and proximal muscle weakness Pediatrics, Neurology, and no evidence of recurrent myoglobinuria or began 4 months prior to evaluation, mani- Pathology, State University central nervous system involvement. Treat- fested by difficulty ascending stairs and lift- of New York, Buffalo ment with CoQ10 supplementation re- ing heavy objects. This was preceded by (Dr Vladutiu). sulted in significant clinical improve- constitutional fatigue for several months.

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A skeletal muscle biopsy was performed at 11.5 years of age. Cryostat sections of flash-frozen muscle were stained with hematoxylin-eosin, the modified Gomori trichrome stain, succinate dehydrogenase, cytochrome-c oxidase, reduced nicotin- A B amide adenine dinucleotide dehydrogenase, and adenosine triphosphatase activities at pH 4.3 and 4.6. Muscle was fixed in 3% glutaraldehyde for electron microscopy. A blood speci- men for mitochondrial DNA mutation analysis was obtained from the proband. Mitochondrial DNA mutation analysis in the proband’s skeletal muscle and blood specimens were performed to analyze common point mutations (myopathy, encephalopathy, lactic acidosis, and strokelike episodes A3243G and T3271C; myoclonic epilepsy and ragged red fibers A8344G C D and T8356C; neuropathy, ataxia, and retinitis pigmentosa T8993G and T8993C; cardiomyopathy G8363A; and Leber he- redity optic neuropathy G11778A, G3460A, T14484C, and G14459A), deletions, and duplications. The respiratory chain (RC) enzyme analysis in skeletal muscle was performed using standard spectrophotometric analyses.8-11 The carnitine palmitoyltransferase activity was quantified using the isotope- exchange method of Norum.12 Fatty acid–oxidation enzyme activities were evaluated by spectrophotometric assays with ␤ chain-length specific substrates of -oxidation. The CoQ10 level was analyzed in skeletal muscle by using high- performance liquid chromatography with UV detection (275 13,14 nm) and using coenzyme Q9 as an internal standard.

E F RESULTS Figure. Histochemical and electron microscopy images. A, Mild variation in fiber size (hematoxylin-eosin, original magnification ϫ400). B, Modified Gomori trichrome (original magnification ϫ400) with slightly granular fibers Skeletal muscle histochemical evaluation revealed rare suggestive of ragged red fibers (arrows). C, Periodic acid–Schiff (original pale-staining myofibers with the cytochrome-c oxidase magnification ϫ400) with many fibers showing increased subsarcolemmal stain and scattered ragged red fibers with the Gomori tri- ϫ glycogen content (arrow). D, Sudan black (original magnification 400) with chrome stain (Figure). On electron microscopy, there increased lipid droplets. E, Electron microscopy (original magnification ϫ5600) with increased lipid droplets (thin arrows) and subsarcolemmal was an increase in the number of mitochondria, al- accumulation of glycogen (thick arrow). F, Subsarcolemmal aggregate of though no abnormally shaped or enlarged mitochon- mitochondria, some of which have electron dense bodies (arrows) (original ϫ dria were found. A prominent increase in lipid droplets magnification 22400). and subsarcolemmal and intermyofibrillar accumulation of free glycogen were found. In muscle, total and He lost weight over this interval and complained of lower free carnitine values were 2.3 SDs below the normal ref- extremity muscle cramps. There was no history of un- erence mean. Fatty acid–oxidation enzymes and carni- explained fever, rash, ataxia, hearing loss, or seizures. tine palmitoyltransferase activities in muscle were nor- Family history was unremarkable for neuromuscular dis- mal. Common mitochondrial DNA point mutations and orders. On physical examination he was noted to have deletions were not detected in skeletal muscle or lym- significantly reduced proximal muscle strength at the phocytes. The RC enzyme analyses showed partial re- shoulders and the hips, with mild wasting of the shoul- ductions in complex I, I + III, and II + III activities (Table) der muscles. The Gower maneuver was noted when aris- with greater than 200% of normal citrate synthase activ- ing from a seated position. There was no evidence of oph- ity, suggestive of increased mitochondrial content and thalmoplegia or ataxia. His creatine kinase level was corroborating histochemical findings. Results of mag- elevated at 359 U/L (reference range, 55-215 U/L), his netic resonance imaging and magnetic resonance spec- lactate level was 33.33 mg/dL (3.7 mmol/L) (reference troscopy of the brain were normal. An echocardiogram range, 1.80-18.01 mg/dL [0.2-2.0 mmol/L]), urinalysis demonstrated normal cardiac function. The patient was did not reveal myoglobinuria, and urine organic acid treated with 150 mg/d of a CoQ10 supplement and 100 analysis detected abnormal metabolites including ethyl- mg/kg per day of carnitine for 3 months. On a 3-month malonic acid, methylsuccinic acid, hexanoylglycine, and follow-up visit, a remarkable improvement in muscle lactic acid. The plasma acylcarnitine profile exhibited el- strength was noted with increased proximal muscle evations of butyrylcarnitine, pentanoylcarnitine, hex- strength and absent Gower sign. Based on his consider- anoylcarnitine, octanoylcarnitine, and decanoylcarni- able improvement with antioxidant therapy, the CoQ10 tine, with no evidence of plasma total carnitine depletion. concentration was analyzed by high-performance liquid Nerve conduction velocity measurements were normal, chromatography in the original skeletal muscle speci- but the electromyogram showed low-amplitude poly- men and found to be 46% of the normal reference mean. phasic units consistent with myopathy. The in vitro addition of 50 µmol/L of coenzyme Q1 to

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©2005 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/29/2021 the succinate cytochrome-c reductase assay of the patient’s skeletal muscle whole homogenate increased the Table. Respiratory Chain Analysis of Patient’s Skeletal succinate cytochrome-c reductase activity 8-fold com- Muscle Tissue pared with a 2.8-fold increase in the normal control ho- Enzyme Activity Reference, mogenates. Citrate synthase activity was not influenced Enzyme (µmol/min-1 per g-1)* Mean ± SD by the addition of coenzyme Q1 to the assay in patients or in controls (data not shown). The CoQ supplemen- NADH dehydrogenase (complex I) 5.1 (35)† 14.74 ± 4.48 10 Succinate dehydrogenase (complex II) 0.53 (61) 0.87 ± 0.21 tation was increased to 300 mg/d at the 3-month visit and NADH cytochrome-c reductase 0.35 (43)† 0.81 ± 0.20 within 6 months of therapy, both the creatine kinase (complex I + III) (140 U/L) and lactate (5.4 mg/dL [0.6 mmol/L]) levels Cytochrome-c oxidase (complex IV) 1.53 (63) 2.43 ± 0.70 normalized, with sustained clinical improvement. Succinate cytochrome-c reductase 0.46 (45)† 1.03 ± 0.31 (complex II + III) Succinate cytochrome-c reductase 3.74 (363) 2.56 ± 0.72 COMMENT and coenzyme Q1 Citrate synthase‡ 36.54 (232) 15.74 ± 4.44

Coenzyme Q10 9.11 (46) 19.81 ± 2.61 Our findings in this case study suggest that CoQ10 deficiency and the concomitant significant reductions in com- Abbreviation: NADH, reduced nicotinamide adenine dinucleotide. plex I, I + III, and II + III enzymatic activities in the RC *Data represent the mean of 2 independent analyses on different muscle homogenates. Figures in parentheses represent percentage of normal reference were responsible for the mitochondrial disorder ob- mean. served in our patient. The observed muscle carnitine de- †Residual enzyme activity greater than 2 SDs below the normal reference ficiency is most likely related to an increased reduced nico- mean. ‡Citrate synthase activity was not influenced by the addition of coenzyme Q1 tinamide adenine dinucleotide–nicotinamide adenine to the assay (data not shown). dinucleotide ratio associated with respiratory chain defects.15 The increased ratio could impair ␤-oxidation at 19 the level of 3-hydroxyacyl-coenzyme A dehydroge- complex III. Coenzyme Q10 allows the extrusion of pro- nases, with a subsequent accumulation of acyl- tons from the matrix to the intermembrane space along coenzyme A ␤-oxidation intermediates. These interme- with the electron flow through the RC.20 Deficiency of diates, released as carnitine esters, are transported into CoQ10 impairs the proton transfer across the inner mi- plasma and eliminated in urine, leading to secondary car- tochondrial membrane, thus affecting generation of aden- nitine deficiency.16,17 osine triphosphate and all adenosine triphosphate– A recent study of 13 patients with childhood-onset cer- dependent metabolic processes. Although the antioxidant ebellar ataxia and marked CoQ10 deficiency suggested a treatment for RC defects has no proven efficacy, treat- cutoff for primary CoQ10 deficiency in muscle at 55% of ment of ubiquinone deficiency might represent an ex- 18 3 the normal reference mean. Our patient’s muscle CoQ10 ception. A defective incorporation of tritium ( H)- activity was 46% of the normal reference mean. This re- mevalonate into CoQ10 in fibroblasts initially suggested sult, in conjunction with the in vitro augmentation of re- a specific site of impairment of endogenous CoQ10 syn- sidual muscle complex II + III activity with the addition thesis.7 Rötig et al7 reported very low concentrations of of exogenous coenzyme Q1 to the assay and the success- labeled decaprenyl-diphosphate in patients’ fibroblast ful clinical outcome with CoQ10 therapy, suggests pri- extracts, consistent with a deficiency of trans-prenyl- mary CoQ10 deficiency in our patient. However, the mo- transferase; however, no mutations in the gene encod- lecular elucidation of this disorder will be required to ing trans-prenyltransferase were identified, suggesting that confirm a primary defect in the ubiquinone biosyn- another gene involved in this pathway may be affected. thetic pathway in all of these cases. We could hypoth- Recently, mutations in the trans-prenyltransferase gene esize that the partial deficiency of CoQ10 observed in our have been identified in 2 siblings with mild intellectual proband perhaps accounts for the late clinical manifes- retardation, profound deafness, optic atrophy, valvu- tation and isolated muscle involvement. However, de- lopathy, and obesity who had CoQ10 deficiency in fibro- tailed review of the reported cases indicates no clear cor- blasts but not in skeletal muscle.21 relation between the observed in vitro muscle or fibroblast At least 4 different clinical manifestations of CoQ10 de- CoQ10 levels and the severity in phenotype and/or age of ficiency have been described: the encephalomyopathic onset in the affected individuals. This is illustrated by the form with myoglobinuria, ataxia, and seizures1-4; a pre- presence of undetectable CoQ10 levels in fibroblasts in 2 dominantly cerebellar disease with ataxia and cerebellar siblings, one with widespread multisystem involvement atrophy5,18; a widespread multisystem involvement with and the other with a milder form of the disease.7 In an- hypertrophic cardiomyopathy, ataxia, optic nerve atro- other report of childhood-onset cerebellar ataxia and phy, deafness, generalized amyotrophy, and nephrotic syn- 18 7 marked CoQ10 deficiency, patients who exhibited muscle drome ; and Leigh encephalopathy with growth retar- 6 CoQ10 concentrations of 2.9 µg/g and 14.8 µg/g, respec- dation, ataxia, deafness, and lactic acidosis. The clinical tively, had a similar phenotype of ataxia and cerebellar heterogeneity found among patients with CoQ10 defi- atrophy by 9 years of age with no developmental delay ciency suggests that a number of biochemical and mo- or seizures. lecular defects may be involved in causing different clini- Coenzyme Q10 plays an important role in the mito- cal phenotypes. chondrial RC by acting as a redox carrier, transferring The isolated myopathy with absence of central ner- reducing equivalents from complex I and complex II to vous system involvement and recurrent myoglobinuria

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©2005 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/29/2021 3. Boitier E, Degoul F, Desguerre I, et al. A case of mitochondrial encephalomy- in our patient expands the clinical phenotype of CoQ10 deficiency. The complete recovery of myopathy with ex- opathy associated with a muscle coenzyme Q10 deficiency. J Neurol Sci. 1998; 156:41-46. ogenous CoQ10 supplementation observed in this pa- 4. Di Giovanni S, Mirabella M, Spinazzola A, et al. Coenzyme Q10 reverses patho- tient highlights the importance of early identification and logical phenotype and reduces apoptosis in familial CoQ10 deficiency. Neurology. treatment of this genetic disorder, perhaps offering a simi- 2001;57:515-518. lar prognosis to patients affected with the myopathic form 5. Musumeci O, Naini A, Slonim AE, et al. Familial cerebellar ataxia with muscle of this condition. Functional studies to identify the pos- coenzyme Q10 deficiency. Neurology. 2001;56:849-855. 6. Van Maldergem L, Trijbels F, DiMauro S, et al. Coenzyme Q-responsive Leigh’s sible defect of ubiquinone synthesis in our patient are cur- encephalopathy in two sisters. Ann Neurol. 2002;52:750-754. rently underway. This case demonstrates the need for de- 7. Rötig A, Appelkvist EL, Geromel V, et al. Quinone-responsive multiple respiratory- tailed biochemical assessment of mitochondrial function chain dysfunction due to widespread coenzyme Q10 deficiency. Lancet. 2000; in the diagnostic evaluation of isolated myopathies. 356:391-395. 8. King TE, Howard RL. Preparation and properties of NADH dehydrogenase from cardiac muscle. In: Estabrook R, Pullman M, eds. Methods in Enzymology: Oxi- Accepted for Publication: March 11, 2004. dation and Phosphorylation. New York, NY: Academic Press; 1967:275-294. Correspondence: Seema Lalani, MD, Department of Mo- 9. King TE. Preparation of succinate dehydrogenase and reconstitution of succi- lecular and Human Genetics, Baylor College of Medi- nate oxidase. In: Estabrook R, Pullman M, eds. Methods in Enzymology: Oxida- cine, One Baylor Plaza, Room T828, Houston, TX 77030 tion and Phosphorylation. New York, NY: Academic Press; 1967:322-331. 10. Yonetan T. Cytochrome oxidase: beef heart. In: Estabrook R, Pullman M, eds. ([email protected]). Methods in Enzymology: Oxidation and Phosphorylation. New York, NY: Aca- Author Contributions: Study concept and design: Lalani, demic Press; 1967:332-335. Vladutiu, and Scaglia. Acquisition of data: Lalani, Vladu- 11. Srere PA. Citrate synthase. In: Lowenstein J, ed. Methods in Enzymology: Oxi- tiu, Plunkett, Lotze, Adesina, and Scaglia. Analysis and dation and Phosphorylation. New York, NY: Academic Press; 1969:3-11. interpretation of data: Lalani, Vladutiu, Adesina, and Sca- 12. Norum K. PalmitoylCoA: carnitine palmityltransferase. Biochim Biophys Acta. 1964; 89:95-108. glia. Drafting of the manuscript: Vladutiu and Lalani. Criti- 13. Laaksonen R, Riihimaki A, Laitila J, Martensson K, Tikkanen MJ, Himberg JJ. cal revision of the manuscript for important intellectual con- Serum and muscle tissue ubiquinone levels in healthy subjects. J Lab Clin Med. tent: Vladutiu, Lalani, Plunkett, Lotze, Adesina, and 1995;125:517-521. Scaglia. Obtained funding: Lalani and Vladutiu. Study su- 14. Rousseau G, Varin F. Determination of ubiquinone 9 and 10 levels in rat tissues pervision: Lalani, Vladutiu, and Scaglia. and blood by high-performance liquid chromatography with ultraviolet detection. J Chromatogr Sci. 1998;36:247-252. Funding/Support: This study was supported by the Doris 15. Wijburg FA, Feller N, Scholte HR, Przyrembel H, Wanders RJ. 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