www.neurologyindia.com Indian Perspective Progressive myoclonic

P. Satishchandra, S. Sinha

Department of , National Institute of Mental Health & Neurosciences, Bangalore, India

Abstract

Progressive (PME) is a complex and is characterized by the development of relentlessly progressive , cognitive impairment, , and other neurologic deficits. It encompasses different diagnostic entities and the common causes include Lafora body disease, neuronal ceroid lipofuscinoses, Unverricht–Lundborg disease, myoclonic epilepsy with ragged-red fiber (MERRF) syndrome, sialidoses, dentato-rubro-pallidal , storage , and some of the inborn errors of , among others. Recent advances in this area have clarified molecular genetic basis, biological basis, and natural history, and also provided Address for correspondence: a rational approach to the diagnosis. Most of the large studies related to PME are from Dr. P. Satishchandra, south India from a single center, National Institute of Mental Health and Neurological National Institute of Mental Health Sciences (NIMHANS), Bangalore. However, there are a few case reports and small series & Neurosciences (NIMHANS), Hosur Road, Bangalore - 560 029, about Lafora body disease, neuronal ceroid lipofuscinoses and MERRF from India. We India. review the clinical and research experience of a cohort of PME patients evaluated at E-mail: drpsatishchandra@yahoo. NIMHANS over the last two decades, especially the phenotypic, electrophysiologic, com pathologic, and genetic aspects.

Key words: Lafora body disease, myoclonic epilepsy with ragged-red fiber, PMID: *** neuronal ceroid lipofuscinoses, progressive myoclonic epilepsy, Unverricht–Lundborg DOI: 10.4103/0028-3886.68660 disease

Introduction There are a few case reports and case series on various PMEs from India.[10-15] However, most of the larger Progressive myoclonic epilepsy (PME) is a syndrome studies are from a single center, National Institute of complex characterized by progressive myoclonus, Mental Health and Neurological Sciences (NIMHANS), [5-7,16] cognitive impairment, ataxia, and other neurologic Bangalore, in south India. The research interest in deficits.[1] It encompasses several diagnostic entities and one of the PMEs, LBD has started in early 1990s and the often causes diagnostic problems leading to nosological genetic analyses of patients with LBD have been carried [9,17] confusion. Over the last two decades, considerable out in collaboration with IIT, Kanpur. developments have occurred in the field of PMEs and A total of 147 patients with PME have been evaluated at they have been recognized as a group of syndromes NIMHANS, Bangalore, India, till date, they include: LBD: with specific etiologies.[2-4] Genetic tests had further 54; NCL: 65; ULD: 08; MERRF: 10; and Tay–Sachs disease enhanced the understanding of the disease process. (TSD): 10 [Table 1]. We have earlier described a cohort The most important causes of PME include:Unverricht– 97 histopathologically confirmed patients with PME Lundborg disease (ULD), myoclonic epilepsy with [Table 2]. This cohort included 63 males and 34 females; ragged-red fiber (MERRF) syndrome, Lafora body mean age at onset of illness 10.7 ± 8.2 years (median: disease (LBD), neuronal ceroid lipofuscinoses (NCL), 11 years, range: 6 months to 48 years); and duration and sialidoses.[2] Further, recent advances have clarified of illness at presentation 2.5 ± 2.2 years. History of the clinical features and facilitated a rational diagnostic consanguineous parentage was evident in 60 patients approach. [1- 3,5-9] Understanding of the molecular genetics (61.8%). Most of the patients presented with the classical may help in determining the biological basis and also the triad: myoclonus, cognitive decline, and neurologic natural history of these disorders. deficits.

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Lafora Body Disease and retinal degeneration has been documented but normal retina is usually noted. Ataxia is often missed The characteristics of LBD include: generalized tonic– because of severe myoclonus. The characteristic EEG clonic (GTCS), resting and action myoclonus, pattern is slowing of background activity with recurrent ataxia, , polyspike and wave discharges in the epileptiform discharges: spikes/polyspikes, with or [20] electroencephalogram (EEG) basophilic cytoplasmic without slow waves. LBD is caused by in the inclusion bodies in portions of , liver, and , as PME 2 (EPM2A) on chromosome 6q and EPM2B [5] well as the duct cells of the sweat glands. The disease gene. has autosomal recessive inheritance with the age of onset between 5 and 20 years. Most often patients with LBD NIMHANS cohort Clinical characteristics present at 13 or 14 years of age with few exceptions.[18] Death usually occurs within 10 years of onset. Seizures, In our cohort, there was a slight male (M:F:: myoclonus, or learning disability may be the first 24:14). History of consanguineous parentage was note in 73.7% of patients. The mean age at onset of illness was symptom in the majority.[5,7,19] Myoclonus is said to be 14.4 ± 3.9 years (range: 10–35 years, median: 14 years) and more often fragmentary, asymmetric, arrhythmic, and the mean duration of illness was 2.8 ± 2.1 years. GTCS was progressively disabling.[5-7,20] Presence of optic atrophy the presenting symptom in 71% of patients. Myoclonus with or without generalized seizures and progressive Table 1: PME cohort from NIMHANS, Bangalore cognitive decline were universally present in all the Type of PME No. of cases Mode of Number patients. Seizures are often refractory to antiepileptic n (%) diagnosis n=147 medications. Occipital seizures with visualization of LBD 54 (36.7) Brain 02 Muscle 02 flashes of light (ictal phenomena) were reported in a Skin 50 third of the patients, while behavioral changes were NCL 65 (44.2) Brain 11 evident in almost one-fourth of the patients. In our Skin and 05 cohort, progression from a presymptomatic stage to muscle 49 only electrophysiologic abnormalities (EEG changes or Skin giant somatosensory-evoked potential [SSEP] potential), ULD 08 (5.4) Exclusion 08 diagnosis and finally to a clinically obvious stage have been [5] Normal skin documented. and muscle biopsy Electrophysiology MERRF 10 (6.8) Muscle 10 Scalp EEG was done in 37 patients and the findings TSD 10 (6.8) Biochemical 10 included varying degrees of slowing of background assay activity in 97.4% of patients [Figure 1a]. Generalized PME - Progressive myoclonic epilepsy; MERRF - Myoclonic epilepsy with ragged-red fiber; NCL - Neuronal ceroid lipofuscinosis; ULD - Unverricht– epileptiform discharges in 84.2% of patients, while Lundborg disease; LBD - Lafora body disease; TSD - Tay–Sachs disease focal discharges were present in ten patients. One

Table 2: Clinical and electrophysiologic profile of 97 patients of PME, cohort from NIMHANS, Bangalore[7] Parameters NCL (n=40) LBD (n=38) MERRF (n=10) ULD (n=9) Age at onset (years) 5.9 ± 9.1 14.4 ± 3.9 14.6 ± 5.8 13.8 ± 9.5 Duration of illness (years) 2.5 ± 1.4 14.4 ± 3.9 2.1 ± 5.2 4.1 ± 4.05 M:F 28:12 24:14 6:4 5:4 Positive family history (%) 12.5 7.9 10 Nil Consanguineous parentage (%) 62.5 73.7 30 44.4 Common manifestations Regression of milestones, Myoclonus, generalized Myoclonus, generalized Myoclonus and ataxia , myoclonus, chorea, seizure, occipital seizures, seizures, cognitive visual loss, and ataxia and cognitive decline decline, and ataxia EEG: n=37: n=37: n=7: n=9: Slowing of BGA 94.6% 97.4% 71.4% 55.5% Epileptiform discharges 81.1% 84.2% 71.4% 88.8% SSEP: n=25: n=31: n=6: n=7: giant potentials (>10 μV) 28% 77.4% 16.6% 57.1% VEP n=2: absent - 2, n=31: absent- 8, n=6: absent- 1 n=7: Prolonged P100- 7 Prolonged P100- 4 Normal Nerve conduction studies n=19: axonal neuropathy- 3 n=21: axonal n=10: - 1; n=9: neuropathy- 1 neurogenic- 1 Normal EEG - Electroencephalogram; BGA - Background activity; LBD - Lafora body disease; MERRF - Myoclonic epilepsy with ragged-red fiber; NCL - Neuronal ceroid lipofuscinosis; PME - Progressive myoclonic epilepsy; ULD - Unverricht–Lundborg disease; SSEP - Somatosensory-evoked potential; VEP - Visual-evoked potential

Neurology India | Jul-Aug 2010 | Vol 58 | Issue 4 515 Satishchandra and Sinha: PME in India of the patient had multifocal epileptiform activity. b Photosensitivity with fast frequency stimulus was observed in 25% of patients, significantly less as compared to the western series..[20] Presymptomatic EEG abnormalities were detected in 3 families.[5] Progressive worsening in the background activity was observed in 4 patients. Interestingly, even presymptomatic individuals were found to have EEG changes, first time documented b from our center.[6] a

Giant SSEP (14–175 μV) was demonstrated in 24 of the 31 patients studied [Figure 1b], while visual-evoked potential (VEP) studies revealed a prolonged P100 in four of the 31 patients studied and absent waveform in eight. Giant VEP potentials have also been documented for the first time from our center[5,6] [Figure 1b]. Brainstem c d auditory-evoked potential studies did not reveal any Figure 1: Laboratory parameters showing characteristic features of LBD: abnormality. Electrophysiologic features of neuropathy (a) scalp EEG with diffuse slowing of BGA with frequent spike/polyspike were present in only one patient of LBD.[7] and waves from both the hemispheres; (b) SSEP: giant potential (35.6 uV), while recording SSEP from contralateral cortex and stimulating at median nerve; (b) giant VEP P100 potential (13.2 uV) during pattern Imaging reversal stimulation; (c) intraneuronal PAS-positive, diastase-resistant Lafora body in the cytoplasm of neurons in the cerebral cortex (×160); Computed tomography (CT) (n=32) and magnetic (d) PAS-positive diastase-resistant Lafora bodies along the base of resonance imaging (MRI) (n=4) of brain revealed apocrine sweat glands within the myoepithelial cells (×160); Inset: inclusions are positive for ubiquitin immunostaining.[7] Abbreviations: diffuse cortical atrophy with no obvious parenchymal EEG - Electroencephalogram; BGA - Background activity; changes similar to the observations in the earlier studies. SSEP - Somatosensory-evoked potential; VEP - Visual-evoked potential; However, MR spectroscopic abnormalities have been PAS - Periodic acid–Schiff noted in patients with LBD with no structural MRI abnormalities: reduction in the N-acetylaspartate disease (laforin).” EPM2A is the gene's official symbol. (NAA):creatine ratio and altered NAA:choline, and The EPM2A gene located at 6q24 provides instructions choline:creatine ratios in frontal cortex, cerebellum, and for making a protein called laforin and the initial paper [25] basal ganglia.[21,22] related to it by Minassian et al., included cases from our center as well. Subsequently, EPM2B (malin) has Histopathologic features been discovered. Although laforin protein is active in In two patients brain biopsy established the diagnosis cells throughout the body, it appears to play a critical by the presence of neuronal intracytoplasmic basophilic, role in the survival of nerve cells (neurons) in the brain. round to oval bodies, which were periodic acid–Schiff Studies suggest that laforin has multiple functions within (PAS)-positive and diastase-resistant [Figure 1c]. Brain the cells. To carry out these functions, laforin interacts biopsy is rarely performed now a days as the diagnosis with several other proteins, including malin (which can easily be established by axillary skin biopsy. The is produced from the NHLRC1 gene). These proteins typical inclusions were described initially by Busard are part of complex networks that transmit chemical et al.,[23,24] and later by Berkovic et al..[1] Similar was the signals and break down unneeded or abnormal proteins. experience at our center. The axillary skin biopsies (n=35) Additionally, laforin may act as a tumor suppressor revealed characteristic oval to round PAS-positive, protein, which means that it keeps the cells from growing diastase-resistant Lafora body inclusions in the sweat and dividing in an uncontrolled way. Laforin and malin glands [Figure 1d]. The Lafora bodies were positive for probably play a critical role in regulating the production Lugol’s iodine and ubiquitin immunostaining [Figure 1d, of a complex sugar called glycogen. Glycogen is a major inset]. Histochemically, Lafora bodies are polyglucosan source of stored energy in the body. The body stores this and its accumulation could be an error of carbohydrate sugar in the liver and muscles, breaking it down when metabolism. Busard et al. had demonstrated normal it is needed for fuel. Researchers believe that laforin pyruvate metabolism in the body fluids and brain.[23,24] and malin may prevent a potentially damaging buildup of glycogen in tissues that do not normally store this Genetics molecule, such as those of the . Recently, High degree of consanguineous parentage in south Rao et al.[26] from National Brain Research Center (NBRC), India might be responsible for high clustering of the New Delhi, had suggested that malin is unstable, and disease in this region. The official name of this gene the aggregate-prone protein and co-chaperone CHIP is “epilepsy, progressive myoclonus type 2A, Lafora can modulate its stability and therefore cause death.

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Nearly 100 distinct have been discovered in NIMHANS cohort the 2 in over 200 independent LD families. Nearly Clinical characteristics half of them are missense mutations, and the deletion Our cohort included 28 males and 12 females; mean age at mutations account for one-quarter. Defects in at least 3 onset of symptom 5.9 ± 9.1years (median: 4 years); mean genes underlie LBD, of which 2 have been isolated and age at presentation 6.6 ± 5.5 years (median 5 years, range 6 their mutations characterized: The EPM2A gene (MIM# months to 48 years), and mean duration of illness 2.5 ± 1.4 [9,16,25,27] 607566) encoding laforin and the NHLRC1 gene years (range 45 days to 7 years). Five patients (12.5%) had [28] (MIM# 608072) encoding malin. Laforin is a protein positive family history with autosomal recessive pattern phosphatase, which is ubiquitinated by malin before of Mendelian inheritance. Consanguineous parentage was [9,16,29-31] degradation. Aberrant functions of laforin and/ noted in 25 (62.5%) patients. Based on the age of onset of or malin, which eventually affect the posttranslational illness, patients could be categorized into four clinical modification of target proteins, are likely to underlie the forms: infantile NCL - 8 (mean age: 1.5 ± 0.5 years), late onset and progression of LBD.[9,31] Almost all the work infantile NCL - 19 (mean age: 3.5 ± 0.5 years), juvenile NCL related to the genetics of LBD in India has been carried - 11 (mean age: 6.5 ± 1.8 years), and adult NCL - 2 (mean out by Ganesh’s team at IIT, Kanpur, on patients from . our center [Table 3]. age: 44 0 ± 5.6 years). Juvenile NCL is the commonest in most reported series.[32-34] The two Kuf’s adult variant Neuronal Ceroid Lipofuscinosis cases in our series could be categorized into type “B” with predominant behavioral changes, dementia, ataxia, and rigidity.[35] The oldest patient of NCL reported in the NCL represents one of the common progressive [36] neurodegenerative disorders during childhood. The literature was aged 63 years. Generally the infantile and term “NCL” was coined by Zeman et al.32 to distinguish late infantile NCL have a rapid course and children usually the familial cerebromacular degeneration clinically and die within 5–10 and 12 years, respectively. In juvenile NCL pathologically distinctive from gangliosidoses. The disease death occurs by 15–25 years, whereas the adult group has [1,8,32,37] group has an autosomal recessive pattern of inheritance a variable course. The presenting clinical features and is commonly characterized by progressive myoclonus in infantile NCL, late infantile NCL, and juvenile NCL with visual failure and accumulation of an autofluorescent group were regression of milestones, seizures, myoclonus, lipopigment in the neurons and glial elements.[33,34] The chorea, visual loss, and ataxia. Two patients with adult first report of NCL from India was a patient with juvenile variant of NCL presented with abnormal behavior and onset NCL by Gulati et al.,[13] subsequently we reported extrapyramidal features. One of them in addition had 11 patients initially[16] and later 40 patients.[7] pyramidal signs, ataxia, and dementia.[7,16]

Table 3: Summary of the identified EPM2A and NHLRC1 mutations in patients with LBD from our cohort Nucleotide change Predicted effect Domain Age at Symptoms affect onset (years) EPM2A Homozygous c.442A>T p.N148Y (missense) 11 GTCS, myoclonic jerks, cognitive decline, ataxia, visual disturbances, behavioral problem Homozygous deletion of exon 3 R159fsX65, (476nu-718nu Ex3) Exon 3 5 Myoclonic jerks Homozygous deletion of exon R159fsX65, (476nu-718nu Ex3) Exon 3 9 Cognitive decline, myoclonic jerks Homozygous deletion of exon R159fsX65, (476nu_718nuEx3) Exon 3 NA NA Homozygous deletion of exon R159fsX65, (476un-718n Ex3) Exon 3 12 GTCS, myoclonus Heterozygous c.28G>A p.E210K (missense) 17 GTCS, myoclonic jerks, congnitive decline, visual disturbances, behavioral problems Homozygous c.412G>T p.E138X (nonsense) NA NA NHLRC1 C64A>C p.S22R (missense) Close to 13 Myoclonic jerks, cognitive decline (inferred from mother) RING Homozygous 836T>C p.L279P (missense) NHL (4th) 13 GTCS, myoclonic jerks Homozygous c.836T>C P.l279p (missense) NHL (4th) 4 GTCS, myoclonic jerks Homozygous c.612ins T p.F205fs232 NHL (3-6) NA NA Homozygous c.676C>T p.0226X NHL (3-6) 16 GTCS, myoclonic jerks, cognitive decline, ataxia, and behavioral problem Homozygous c.377T>C L126P NHL (1st) 9 GTCS, myoclonic jerks, cognitive decline, visual disturbances, and behavioral problems GTCS - Generalized tonic–clonic seizures; LBD - Lafora body disease

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Visual impairment at onset was noted in ten patients of brain (n=5) and skin (n=28) showed characteristic (25%) and subsequently 26 (65%) of them developed curvilinear inclusions [Figure 2e] in several cell types: visual abnormalities, which gives diagnostic clue in neurons, occasional astrocytes, sweat glands, vascular patients with early onset PME syndrome. Two patients of endothelial and smooth muscle cells, fibroblasts, and INCL had retinal degeneration and one had optic atrophy. occasional Schwann cells. In a single muscle biopsy, Optic atrophy and retinal degeneration was evident in 11 the inclusions were seen in the myofibers. Lamellar and three patients of LINCL, respectively. The patients in and electron dense bodies were infrequent. Clearly, the juvenile group had ophthalmic abnormalities in the form presence of curvilinear inclusions in multiple cell types of primary optic atrophy (n=5), macular degeneration makes the latter a feasible choice for diagnostic biopsy at (n=3) and retinitis pigmentosa (n=1). The adult forms our center. Inclusions are also present in the peripheral did not have any ophthalmic abnormality. Zeman et al. lymphocytes and their morphology has been (1970) have found pigmentary changes in 25, macular found to correlate with the clinical course and genetic degeneration in 12, and optic atrophy in 5 patients giving analysis based on which NCL is further categorized clue to the diagnosis when associated with regression of as (1) infantile NCL—granular bodies/GRODs, (2) milestones and myoclonus in pediatric patients.[32] late infantile NCL—curvilinear bodies, (3) juvenile NCL—finger print bodies, and (4) adult onset NCL Electrophysiology and imaging with varied forms and combination of inclusions.[7,16,43]. Scalp EEG (n=37) showed varying degrees of diffuse Overlapping of the morphologic features of inclusions in slowing of background activity in 94.6% and generalized different subtypes can pose a problem in the diagnosis epileptiform discharges in 81.1% of patients [Figure 1a]. by ultrastructural studies, although they may reflect Slow frequency photic stimulation-evoked response in evolution of cytologic pathology.[1,16,42,43] five (22.7%) of the 22 patients studied [Figure 2a]. These observations were similar to the observations reported in Genetics [16,32,38] the literature. Giant SSEP was record in 7 of the 25 The details of genetics in NCL are mentioned in the patients studied [Figure 2b] and evaluation of VEP study Table 4.[44] All the mutations known to cause NCL are revealed a prolonged P100 in two, absent waveform in listed in the NCL mutation database. More than 150 seven. Nerve conduction showed axonal neuropathy in mutations are known. By 2008, at least 8 genes that cause three of the ten patients studied. CT (n=35)/MRI (n=3) NCL in children had been identified. They are theCLN10/ scans revealed diffuse atrophy of variable degrees, CTSD, CLN1, CLN2, CLN3, CLN5, CLN6, CLN7, CLN8 [39,40] similar to findings in the literature. genes and possibly also the CLCN6 gene. It is known as to what protein is encoded by each gene and for some it Histopathology is known as to what function the protein is involved in, The cornerstone in the diagnosis of PMEs is histopathologic and ultrastructural examination of the various tissues, such as brain, skin, muscle, and liver. [41] During the initial periods, diagnosis was based on brain biopsy but it has been conclusively observed that extracerebral tissues, such as lymphocytes, axillary skin, muscle, rectal mucosa, and peripheral nerves may also reveal abnormalities on electron microscopy studies to establish the diagnosis.[41,42] Ultrastructural studies of skin biopsy have yielded consistent results. a b Histopathologic examination of brain biopsy (n=12) revealed that the brunt of the disease is in the gray matter. The cortical mantle was atrophic and showed neuronal depletion and reactive astrocytosis. The d preserved neurons were normal in size and on H & E stains no identifiable inclusions could be seen. PAS stain for polysaccharide and Luxal fast blue stain for myelin c e revealed intense granular staining of the entire neuronal Figure 2: Laboratory parameters showing characteristic features of NCL: (a) scalp EEG: Generalized epileptiform discharges with slowing cytoplasm indicating the carbohydrate and lipid nature of BGA; (b) SSEP: giant potential (24.1 uV), while recording SSEP from of the stored material, similar to lipofuscin pigment contralateral cortex and stimulating at median nerve; (c) photograph revealing the presence of Luxol fast blue–positive ceroid material [Figure 2c]. Examination of unstained sections under within the neurons (×250); (d) photograph showing granules within fluorescent showed yellow autofluorescence neurons exhibiting autofluorescent ceroid material (×160); (e) electron micrograph revealing curvilinear bodies by electron microscopy (EM) in of the substance [Figure 2d]. Skin and muscle were normal skin biopsy (×35,000).[7] NCL - Neuronal ceroid lipofuscinoses; on light microscopy. However, ultrastructural studies BGA - Background activity; SSEP - Somatosensory-evoked potential

518 Neurology India | Jul-Aug 2010 | Vol 58 | Issue 4 Satishchandra and Sinha: PME in India but for others this is not yet clear. The CLN10/CTSD gene Table 4: Genetic classification of NCL causes the earliest onset of NCL, which affects babies at Clinical types Genetic Chromosomal Gene product birth or even before they are born as well as milder cases types loci that start in late infancy or even in the teenage years. The EO-NCL CLN10/ not known Not known CLN1 gene causes infantile NCL, and also milder cases CTSD that start in late infancy, at a juvenile age, and even in I-NCL CLN1 Ip32 Lysosomal palmitoy1 adulthood. CLN2, CLN5, CLN6, CLN7, and CLN8 all Protein thioesterase LI-NCL CLN2 11p15 Lysosomal pepstatin cause NCL that almost always starts in late infancy. Insensitive peptidase CLN2 is referred to as the classic late infantile gene and LI-NCL CLN5 13q31-32 Not known CLN5, CLN6, CLN7, and CLN8 are referred to as variant J-NCL CLN3 16p12 Transmembranous (?) late infantile genes because the disease course is slightly A-NCL CLN4 not known Not known different to that caused by CLN2. These 4 types of variant EJ-NCL CLN6 15q21-23 Not known LINCL are virtually indistinguishable clinically so are tLI-NCL CLN7 not known Not known sometimes called Finnish, Czech, or Turkish variant vLI-NCL CLN8 not known Not known LINCL, respectively, because they were first described I-NCL - Infantile NCL; LI-NCL - Late infantile NCL; J-NCL - Juvenile NCL; in families from these countries. CLN3 causes NCL that A-NCL - Adult NCL; EJ-NCL = Early juvenile NCL; EO-NCL: Early onset NCL begins at a juvenile age. Recently 2 patients with late onset (teenage and adulthood) were described with CT/MRI scan showed diffuse cerebral atrophy. One single mutations in CLCN6. There are no genetic studies patient had giant SSEP (27 μV). Histochemical stains of of NCL from India.[9,16,43,45-48] muscle biopsy showed subsarcolemmal accumulation of reaction product for nicotinamide adenine dinucleotide/ Myoclonic Epilepsy with Ragged-red Fibers succinate dehydrogenase (NADH/SDH) enzymes and classical ragged-red fibers by modified gomori Myoclonic epilepsy with ragged-red fibers (MERRF) is a trichrome (MGT) stain [Figure 3a]. Electron microscopic disorder that affects many parts of the body, particularly studies confirmed the aggregation of abnormal forms of muscles and the nervous system. In most cases, the signs mitochondria subsarcolemmally [Figure 3b]. and symptoms of this disorder appear during childhood or . The features of MERRF vary widely Genetics among affected individuals, even among members of MERRF is inherited in a mitochondrial pattern, the same family. MERRF is characterized by myoclonus, also known as maternal inheritance. This pattern of myopathy, and . The characteristic histological inheritance applies to genes contained in mtDNA. feature is feature abnormal muscle cells are called Mutations in the MT-TK, MT-TL1, MT-TH, and MT-TS1 ragged-red fibers. Other features of MERRF include genes cause MERRF. Mutations in the MT-TK gene are seizures, ataxia, and dementia. the most common cause of MERRF, occurring in more These patients may also develop deafness and optic than 80% percent of the cases. Less frequently, mutations atrophy. Affected individuals sometimes have short in the MT-TL1, MT-TH, and MT-TS1 genes have been stature and abnormalities, . Less reported to cause the of MERRF. commonly, people with MERRF develop . From Individuals with mutations in these genes often have India cases with classical have been described features of other mitochondrial disorders as well. These by Mehndiratta et al.[14] and Sundaram et al.[15] genes are contained in mitochondrial DNA (mtDNA). Mitochondria are structures within cells that use oxygen NIHMANS cohort to convert the energy from food into a form that cells can Clinical characteristics use. Although most DNA is packaged in chromosomes The clinical features of MERRF in our cohort were similar within the nucleus, mitochondria also have a small to those reported in the literature: six males and four amount of their own DNA. The genes associated with females and mean age at onset 14.6 ± 5.8 years (range: MERRF provide instructions for making molecules called 8–26 years). One patient had a family history of similar transfer RNAs, which are chemical cousins of DNA. illness and three patients had a history of consanguineous These molecules help assemble protein building blocks parentage. Eight patients had myoclonus and GTCS. called amino acids into full-length, functioning proteins Cognitive decline and ataxia were evident in seven within mitochondria. Mutations that cause MERRF patients. One patient had in addition, abnormal impair the ability of mitochondria to make proteins, behavior. EEG was abnormal in six of the nine patients use oxygen, and produce energy. These mutations and demonstrated slowing of background activity particularly affect organs and tissues with high energy with/without generalized epileptiform discharges. requirements, such as the brain and muscles. Researchers Electrodiagnostic studies showed neuropathy in one have not determined how changes in mtDNA lead to the patient and myopathic and neurogenic in one each. specific signs and symptoms of MERRF. They continue to

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NIMHANS cohort Clinical characteristics Nine patients of probable ULD, five females and four females, were studied. The diagnosis of ULD was after detailed evaluation including skin and/or muscle biopsy. A history of consanguineous parentage was reported in four patients. Their mean age at onset was 13.8 ± 9.5 years (range: 5–32 years) and the mean duration of illness 4.1 ± 4.05 years. The presenting features were myoclonus and ataxia with no evidence of cognitive dysfunction. Neuroimaging revealed diffuse atrophy of cerebrum, brainstem, and cerebellum. EEG (n=9) showed generalized epileptiform discharges. Somatosensory a evoked potential (SSEP) done in seven patients revealed giant potentials (32–60 μV) in four.

Genetics Mutations in the CSTB gene cause ULD. The CSTB gene provides instructions for making a protein called cystatin B. This protein reduces the activity of enzymes called cathepsins. Cathepsins help break down certain proteins in the lysosomes (compartments in the cell that digest and recycle materials). While the specific function of cystatin B is unclear, it may help protect the cells' proteins from cathepsins that leak out of the lysosomes. One region of the CSTB gene has a particular repeating sequence of 12 DNA building blocks (nucleotides). This sequence is normally repeated 2 or 3 times within the b gene and is called a dodecamer repeat. Most people with Figure 3: Muscle biopsy in patients with MERRF: (a) modified Gomori trichrome stain: classical ragged-red fibers; inset: SDH stain: this disorder have more than 30 repeats of the dodecamer subsarcolemmal oxidation reaction products (×250);(b) electron sequence in both copies of the CSTB gene. The increased micrograph of showing subsarcolemmal aggregation of mitochondria with paracrystalline inclusions (×9600).[7] number of dodecamer repeats in the CSTB gene seems MERRF - Myoclonic epilepsy with ragged-red fiber; SDH - Succinate to interfere with the production of the cystatin B protein. dehydrogenase Levels of cystatin B in affected individuals are only 5%–10% of normal, and cathepsin levels are significantly investigate the effects of mitochondrial gene mutations increased. These changes are believed to cause the signs [49-51] in various tissues. and symptoms of ULD, but it is unclear how a reduction in the amount of cystatin B leads to the features of this Unverricht–Lundborg Disease disorder.[53,54]

ULD is a rare inherited form of epilepsy, more commonly PME – Diagnostic Approach reported from Scandinavian countries. Affected individuals usually begin showing signs and symptoms Most of the patients with PME have onset of the disease of the disorder between the ages of 6 and 15 years. The within the first two decades of life and have an autosomal characteristic feature is myoclonus with increase in recessive pattern of inheritance. A general approach to frequency and severity over time and stimulus sensitive. patients with PME syndrome is mentioned in Figure 4. Within 5–10 years of onset of the disease the myoclonic Recent advances in the field of genetics of PME would jerk may become severe enough to interfere with walking differentiate these groups. This might lead to the and other activities daily living. GTCS is the other seizure development of drugs for the management of these fatal type. After several years of progression, the frequency forms of disease in future. of seizures may stabilize or decrease. Eventually these patients develop ataxia, depression, and mild decline in Acknowledgments intellectual functioning. Patients with ULD typically live [52] into adulthood and the life expectancy may be normal. We are grateful to Drs. A. Mahadevan, T, Yasha, and S.K. From India there no reports of genetically proven cases Shankar, Department of Neuropathology, NIMHANS, of ILD. However, we have reported the probable cases. Bangalore, India, and Dr. S. Ganesh, IIT, Kanpur, for their

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Patients with PME

Step 1: Clinical

LBD NCL ULD MERRF TSD

Early dementia Severe dementia Ataxia Myoclonus Startle Myoclonus

Visual failure Retinal degeneration Myoclonus Deafness Cherry red spot

Occipital seizures Optic atrophy Normal cognition Neuropathy 1st two years

Early 2nd decade 1st decade 1st/ 2nd decade Retinitis pigmentosa

Step 2: Electrophysiology

Giant SSEP Giant SSEP Giant SSEP Neuropathy Absent VEP

Photosensitive Photosensitive

Seiz at high freq PS Seiz. at low freq PS

Step 3: Final diagnosis

Pathological Biochemical Genetics

Skin Liver Muscle Brain TSD NCL: CLN 1-8

LBD LBD MERRF LBD LBD: EPM2A/B

NCL (EM) TSD LBD NCL MERRF: MT-TK

NCL ULD: CSTB LBD - Lafora body disease; MERRF - Myoclonic epilepsy with ragged-red fiber; NCL - Neuronal ceroid lipofuscinosis; PME - Progressive myoclonic epilepsy; TSD: Tay sach’s disease; ULD - Unverricht–Lundborg disease Figure 4: Approach to patients with progressive myoclonic epilepsy association with the research work on PME at various stages. pathological study from south India. J Neurol Sci 2007;252:16-23. Not the least, we are very thankful to our patients who 8. Shahwan A, Farrell M, Delanty N. Progressive myoclonic : A consented for the research work. review of genetic and therapeutic aspects. Lancet Neurol 2005;4:239-48. 9. Singh S, Ganesh S. Lafora progressive myoclonus epilepsy: A meta- analysis of reported mutations in the first decade following the discovery References of the EPM2A and NHLRC1 genes. Hum Mutat 2009;30:715-23. 10. Dastur DK, Singhal BS, Gootz M, Seitelberger M. Atypical inclusion 1. Berkovic SF, Andermann F, Carpenter S, Wolfe LS. Progressive bodies with myoclonic epilepsy. Acta Neuropathologica 1966;7:16-25. 11. Chowdhury C, Jain S, Roy S, Maheshwari MC. Lafora’s disease: A case myoclonus epilepsies: Specific causes and diagnosis. N Engl J Med report. Neurol India 1984;32:59-64. 1986;315:296-305. 12. Jain S, Maheshwari MC. Progressive myoclonic epilepsies without EEG 2. Berkovic SF, Cochius J, Andermann E, Andermann F. Progressive abnormality. Neurol India 1987;35:159-62. myoclonic epilepsies: Clinical and genetic aspects. Epilepsia 1993;34:19-30. 13. Gulati S, Maheshwari R, Kabra M, Verma IC, Kalra V. Juvenile neuronal 3. Lake BD. Lysosomal and peroxisomal disorders. In: Adams JH, Duchen ceroid lipofuscinosis. Indian J Pediatr 2000;67:689-91. th LW, editors. Greenfield’s Neuropathology. 6 ed. London: Edward 14. Mehndiratta MM, Agarwal P, Tatke M, Krishnamurthy M. Neurological Arnold; 1997. p. 668-76. mitochondrial cytopathies. Neurol India 2002;50:162-7. 4. Classification of progressive myoclonus epilepsies and related disorders. 15. Sundaram C, Meena AK, Murthy JM, Venkateswar RB, Mohandas Marseille Consensus Group. Ann Neurol 1990;28:113-6. S. Diagnosis of mitochondrial diseases: Clinical and histological study 5. Acharya JN, Satishchandra P, Asha T, Shankar SK. Lafora's disease of sixty patients with ragged red fibers. Neurol India 2004;52:353-8. in south India: A clinical, electrophysiological and pathological study. 16. Sinha S, Satishchandra P, Vani S, Shankar SK. Neuronal ceroid Epilepsia 1993;34:476-87. lipofuscinosis: A clinicopathological study. Seizure 2004;13:235-40. 6. Acharya JN, Satishchandra P, Shankar SK. Familial progressive 17. Ganesh S, Puri R, Singh S, Mittal S, Dubey D. Recent advances in the myoclonic epilepsy: Clinical and electrophysiologic observations. molecular basis of Lafora’s progressive myoclonus epilepsy. J Hum Epilepsia 1995;36:429-34. Genet 2006;51:1-8. 7. Sinha S, Satishchandra P, Yasha T, Gayathri N, Shankar SK. 18. Genton P, Borg M, Vigliano P, Pellisier J, Roger J. Semi-late and Progressive myoclonic epilepsy: A clinical, electrophysiological and rapidly progressive case of Lafora’s disease with predominant cognitive

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