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Annals of Clinical & Laboratory Science, vol. 40, no. 4, 2010 375 Clinical and Genetic Analysis of a Korean Family with Hereditary Spastic Paraplegia Type 3

Min-Jung Kwon,1 Seung-Tae Lee,1 Jong-Won Kim,1 Duk Hyun Sung,2 and Chang-Seok Ki1 Departments of 1Laboratory Medicine & Genetics, and 2Physical Medicine & Rehabilitation, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

Abstract. Hereditary spastic paraplegia (HSP) is a neurodegenerative disease characterized by progressive spasticity in the lower extremities. Mutations in the atlastin GTPase 1 (ATL1) cause approximately 10% of autosomal dominantly inherited HSP. For many subjects with an ATL1 mutation, spastic gait begins in early childhood and does not significantly worsen, even over many years; such cases resemble spastic diplegic cerebral palsy. Herein we report a heterozygous R239C mutation in the ATL1 gene in a Korean family. The family members exhibited early onset pure spastic paraplegia and had been previously diagnosed with the diplegic form of cerebral palsy. We suggest that spastic paraplegia type 3 (SPG3A) be included in the differential diagnosis of early onset spastic paraplegia. To the best of our knowledge, this is the first report of a genetically confirmed family affected with SPG3A in Korea.

Keywords: hereditary spastic paraplegia, atlastin GTPase 1, ATL1 mutation

Introduction Spastic paraplegia 3 (SPG3A; OMIM 182600) is caused by mutations in the atlastin GTPase 1 Hereditary spastic paraplegia (HSP) comprises a (ATL1) gene, which is also known as the SPG3A group of inherited neurodegenerative disorders that gene, located on band 14q22.1. The cause progressive weakness and spasticity, SPG3A form of HSP typically presents with a pure predominantly affecting the lower extremities. phenotype, early onset, and a benign prognosis. HSP is due to a length-dependent retrograde For many subjects with an ATL1 mutation, spastic degeneration of the corticospinal tracts and gait begins in early childhood and does not fasciculus gracilis [1]. Conventionally, patients with significantly worsen, even over many years. Such HSP are classified into pure and complicated types cases resemble spastic diplegic cerebral palsy, which based on the absence or presence of accompanying is the most common cause of spastic paraplegia extra-neurological features, such as retinopathy, affecting infants. However, spasticity characterized dementia, deafness, ataxia, and mental retardation by progressive gait impairment may also be the first [2], and on the mode of inheritance (autosomal sign of infantile onset HSP [4,5]. dominant [AD], autosomal recessive [AR], or In Korea, there is a report demonstrating that X-linked recessive). At least 41 spastic paraplegia SPAST gene mutations are the most common cause gene loci have been mapped and 17 affected of HSP, as in other ethnic groups [6]. However, have been identified to date [3]. although the ATL1 gene has been known to be the

Address correspondence to Duk Hyun Sung, M.D., Ph.D., Department of Physical Medicine and Rehabilitation, Samsung Medical Center, 50 Irwon-dong, Gangnam-gu, Seoul, Korea, 135-710; tel 82 2 3410 2818; fax 82 2 3410 0052; e-mail yays@ skku.edu; or to Chang-Seok Ki, M.D., Ph.D., Department of Laboratory Medicine and Genetics, Samsung Medical Center, 50 Irwon-dong, Gangnam-gu, Seoul, Korea, 135-710; tel 82 2 3410 2709; fax 82 2 3410 2719; e-mail [email protected].

0091-7370/10/0400-0375. $1.75. © 2010 by the Association of Clinical Scientists, Inc. 376 Annals of Clinical & Laboratory Science, vol. 40, no. 4, 2010 second most common cause of HSP in other dorsiflexion to plantarflexion in the sagittal plane during the populations, there have been no reports of a Korean mid-stance phase. These findings are characteristics of a spastic paretic gait. However, the patient could walk indoors HSP patient with an ATL1 gene mutation. In the and outdoors and climb stairs while holding a railing. She had present study, we report a point mutation in the no cerebellar ataxia, sensory symptoms, or urinary bladder ATL1 gene in a Korean family with infantile onset urgency, and she had normal intelligence and language spastic paraplegia whose members were originally function. She underwent brain and spine magnetic resonance diagnosed with the diplegic form of cerebral palsy. imaging (MRI) studies, with normal results. The family history revealed that the patient’s maternal To the best of our knowledge, this is the first report grandmother (age 80 yr, I-2), mother (age 51 yr, II-3), of a genetically confirmed, SPG3A affected family deceased maternal aunt (II-5), deceased maternal cousin (III- in Korea. 3), and maternal uncle (age 38 yr, II-6) also had gait disturbances beginning in childhood (Fig. 1). All affected Materials and Methods family members could walk without support, except the grandmother who had been wheelchair-bound since 75 yr of age. The patient’s mother (II-3) was affected in a similar Patients. The proband (Fig. 1, III-1) was a 28-yr-old woman manner and showed minimal gait abnormalities, such as with slowly progressive leg spasticity and weakness, which intermittent dragging of her feet for the past 7 yr and began in infancy. She was born to nonconsanguineous parents progressive lower extremity weakness, which had only been after an uncomplicated pregnancy and delivery. At one mo of recognized recently. Her neurological examination at age 50 life, she developed respiratory distress and a fever, which led yr revealed a stiff knee gait, positive Babinski sign, and to an initial diagnosis of cerebral palsy at a university hospital. increased deep tendon reflexes in the lower extremities. She had normal development except for her lower extremities However, her gait disturbance was unremarkable during slow and an abnormal tiptoeing gait, which was identified at two walking and became apparent only when she was running or yr of age. At seven yr of age, at a different regional hospital, moving quickly. the patient underwent tendon release surgery in both Achilles Detailed medical information of the patient’s maternal tendons to improve her walking ability and to reduce her aunt (II-5) and cousin (III-3) could not be obtained because spasticity under a presumptive diagnosis of spastic cerebral they had died in a car accident 15 yr ago. The patient’s palsy. At the same hospital, the patient again underwent maternal uncle (II-6) was diagnosed as having a diplegic form tendon release surgery in both Achilles tendons at age 21. of cerebral palsy at two yr of age. A mild regression in his Upon admission to our hospital, the patient’s motor ability had been noticed since early adulthood. At the neurological examination showed increased knee jerk and age of 28 yr, he underwent Achilles tendon release surgery at ankle jerk reflexes, positive Babinski sign, and ankle clonus. a university hospital, which did not improve his walking Her lower extremity motor strength was poor to good, and capacity. Although he exhibited some characteristics of a sensory examination revealed no abnormal findings. Her spastic paretic gait, such as bilateral stiff knee gait, he did not lower extremities were spastic and gait analysis revealed require supportive devices. Other family members and their excessive pelvic anterior tilt, stiff knees during the swing offspring did not have any histories of gait abnormalities. phase, and abrupt reversal of ankle movement direction from Genetic analysis. To identify the genetic defect, we performed molecular genetic testing for the SPA3A and SPG4 forms of HSP. After we obtained informed consent, blood samples were collected from the patient, her mother, and her maternal uncle. The maternal grandmother was unavailable for study. Genomic DNA was isolated from peripheral whole blood leukocytes using the Wizard Genomic DNA Purification Kit (Promega, Madison, WI) following the manufacturer’s directions. All coding exons and their flanking intronic sequences of the ATL1 and SPAST genes were amplified using the polymerase chain reaction (PCR) in a thermal cycler (Model 9700; Applied Biosystems, Foster City, CA) with primers designed by the authors (available upon request). Direct sequencing was performed using the same primer sets and the BigDye Terminator Cycle Sequencing Ready Reaction kit (Applied Biosystems) on the ABI Prism 3130 genetic analyzer (Applied Biosystems). To identify sequence variations, the subject’s sequences were compared with reference sequences for the ATL1 and SPAST genes (GenBank NM_015915.4 and NM_014946.3, respectively) using Fig 1. Pedigree of proband and her family. Sequencher software (Gene Codes Corp., Ann Arbor, MI). ATL1 mutation in hereditary spastic paraplegia 377

Fig 2. Direct sequencing of the ATL1 gene in the patient and her family. A heterozygous missense mutation (c.715C>T; p.Arg239Cys) was identified in the patient, her mother, and her maternal uncle (arrow).

Results was homozygous for major alleles in Asian samples according to HapMap data (http://www.hapmap. Direct sequencing analyses of the ATL1 and SPAST org; http://www.ncbi.nlm.nih.gov/projects/SNP), genes were performed and revealed a missense no other sequence variations were observed in the mutation in the ATL1 gene. This mutation caused ATL1 and SPAST genes. a C-to-T substitution at nucleotide position 715 in Family analysis revealed that the two other exon 7, replacing arginine with cysteine in codon affected individuals who participated in the 239 (c.715C>T; p.Arg239Cys) (Fig. 2). Except for molecular genetics study, the patient’s mother (II- an intronic SNP (rs2934684) where the patient 3) and maternal uncle (II-6), were both heterozygous 378 Annals of Clinical & Laboratory Science, vol. 40, no. 4, 2010 for the same R239C mutation as was found in the not been selected for early age of onset, ATL1 proband (Fig. 2). mutations were not found. In this kindred, the members were unaware Discussion they had a genetically transmissible disease and had been provided no information regarding Spastic paraplegia 3 (SPG3A; OMIM 182600) is hereditary paraplegia. The patient’s mother caused by mutations in the ATL1 gene. The ATL1 exhibited mild lower extremity symptoms and an gene includes 14 exons spanning 69 kb and encodes unrecognized spastic gait. The patient’s maternal atlastin-1, which is a member of the dynamin uncle had lower extremity symptoms that were superfamily of large GTPases. Atlastin-1 is diagnosed as a diplegic form of cerebral palsy when predominantly localized in the central nervous he was two yr old, and the patient was incorrectly system, where it is enriched in pyramidal neurons diagnosed by her original treating physicians. It has in the cerebral cortex and hippocampus [7,8]. This been reported that cases of infantile onset HSP can has been implicated in vesicle formation, be misdiagnosed as spastic diplegic cerebral palsy transport from the endoplasmic reticulum (ER) to [17-19]. Progressive spasticity of lower extremities is the Golgi interface, maintenance of the Golgi a symptom commonly used to distinguish these apparatus, and ER morphogenesis [9,10]. two diseases and the other distinctions include no The diagnosis of SPG3A is usually made based apparent underlying causes, minimal upper on genetic analysis. There are 27 disease-causing extremity symptoms, normal intelligence, and mutations reported in ATL1 ( Gene previous family history. However, some patients Mutation Database at http://www.hgmd.cf.ac.uk/ with SPG3A can have minimal worsening of their ac/gene.php?gene=ATL1, last updated Jan 2010), symptoms and a negative family history may be including 25 missense/nonsense mutations and 2 caused by mildly affected members who did not small insertions. Furthermore, for professional report any problems at the time of evaluation. subscribers, 39 mutations have been reported, and SPG3A should be included in the differential the Genetics Home Reference (http://ghr.nlm.nih. diagnosis of early onset spastic paraplegia and gov/gene/ATL1) currently states that there are more genetic testing for the causative gene, ATL1, may than 30 mutations in the ATL1 gene that have been be necessary to distinguish these two disease identified in SPG3A. In fact, ATL1 mutations entities, which cause gait disturbances early in life. cause approximately 10% of autosomal dominantly In summary, we present a Korean family with inherited uncomplicated HSP. However, the SPG3A and a missense mutation in the ATL1 gene. mutation detection rate varies for each report and Their clinical phenotypes corresponded to the pure geographical origin, including Tessa et al [11] (1/8, form of HSP with early onset of spastic paraplegia 12%), Smith et al [12] (6/70, 9%), Abel et al [13] and the family members had previously been (1/12, 8%), Meijer et al [14] (1/70, 1.4%), and diagnosed with a diplegic form of cerebral palsy. To Souter et al [15] (5/13, 38%). Namekawa et al [16] our knowledge, this is the first report of a genetically reported that SPG3A is the most frequent HSP in confirmed family affected with SPG3A in Korea. patients with onset before ten yr of age and that it This report highlights the necessity of considering is twice as frequent as SPG4 in this age group. SPG3A in the differential diagnosis of early onset However, there have been no previous reports of pure lower extremity motor impairment, particu- genetically confirmed SPG3A in Korean HSP larly in patients without significantly worsening patients, and there has been only one previous symptoms, even over many years. investigation of SPG3A in AD-HSP. Park et al [6] reported a mutation analysis of the ATL1 and Acknowledgements SPAST genes in 18 Korean patients with dominantly inherited and uncomplicated HSP (SPG3A and This study was supported by Samsung Biomedical SPG4). Although the samples analyzed included a Research Institute grant # C-B0-206-1 and by a small number of patients with AD-HSP who had Grant No. A080588 from the Korea Healthcare ATL1 mutation in hereditary spastic paraplegia 379

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