The hidden genetics of epilepsy a clinically important new paradigm

Transcription

1 FOCUS ON EPILEPSY The hidden genetics of epilepsy a clinically important new paradigm Rhys H. Thomas and Samuel F. Berkovic Abstract Understanding the aetiology of epilepsy is essential both for clinical management of patients and for conducting neurobiological research that will direct future therapies. The aetiology of epilepsy was formerly regarded as unknown in about three-quarters of patients; however, massively parallel gene-sequencing studies, conducted in a framework of international collaboration, have yielded a bounty of discoveries that highlight the importance of gene mutations in the aetiology of epilepsy. These data, coupled with clinical genetic studies, suggest a new paradigm for use in the clinic: many forms of epilepsy are likely to have a genetic basis. Enquiry about a genetic cause of epilepsy is readily overlooked in the clinic for a number of understandable but remediable reasons, not least an incomplete understanding of its genetic architecture. In addition, the importance of de novo mutagenesis is often underappreciated, particularly in the epileptic encephalopathies. Other genomic surprises are worth emphasizing, such as the emerging evidence of a genetic contribution to focal epilepsies long regarded as acquired conditions and the complex role of copy number variation. The importance of improved understanding of the genetics of the epilepsies is confirmed by the positive outcomes, in terms of treatment selection and counselling, of receiving a genetic diagnosis. Thomas, R. H. & Berkovic, S. F. Nat. Rev. Neurol. 10, (2014); published online 15 April 2014; doi: /nrneurol Epilepsy Research Centre, Melbourne Brain Centre, Department of Medicine, Austin and Northern Health, 245 Burgundy Street, Heidelberg, Vic 3084, Australia (R.H.T., S.F.B.). Correspondence to: S.F.B. s.berkovic@ unimelb.edu.au Introduction The current belief among ancient writers, and among many at the present day, is that epilepsy is pre-eminently an [sic] hereditary affection. John Russell Reynolds (Epilepsy: Its Symptoms, Treatment and Relation to Other Chronic Convulsive Diseases, 1861). 1 John Russell Reynolds was a distinguished British physician and member of the Royal Society. His textbook from 150 years ago 1 is enlightening in that much of what he relates remains pertinent to a contemporary understanding of epilepsy and its aetiology. He estimated that one-third of the patients then characterized as having idiopathic epilepsy had a family history of epilepsy, and that the ratio of familial to nonfamilial epilepsies was equal between the sexes. If this view was previously incontrovertible, the reader might well ask what changed in the intervening eras to necessitate its reassertion here. The rise of eugenics in the early 20 th century 2 4 was one factor that probably limited investigation into the heritability of epilepsy, Competing interests S.F.B. declares that he has received honoraria and/or payments for development of educational presentations from Janssen- Cilag, Novartis Pharmaceuticals, Sanofi-Aventis, and UCB Pharma, and that he is an inventor on a patent for SCN1A testing owned by Bionomics and licensed to various diagnostic companies (WO/2006/133508); he has also applied for a patent for PCDH19 testing (WO/2009/086591). R.H.T. declares no competing interests. and downplaying the importance of transmissible epilepsy might have been expedient for physicians of that era. That the great epileptologist William Lennox, who conducted landmark twin and family studies that confirmed the importance of genetic causes in some types of epilepsy, 4 also struggled with eugenic views demonstrates in microcosm the conflict that then gripped the scientific community. Today, Reynolds would probably still recognize many aspects discussed in this Review, particularly the ongoing challenges in epilepsy research. By contrast, he would not have been able to predict the exponential increase in genetic information, driven primarily by improvements in gene-sequencing technology. These advances have flooded the medical literature in such a way that even enthusiasts struggle to keep abreast of important discoveries. Although there has never been a more promising era for studying the genetic basis of epilepsy, some neurologists have argued that the discoveries made so far have failed to translate into meaningful improvements in clinical practice. In this Review, we will robustly refute the view that genomic data have little practical relevance to epileptologists by exploring the hidden genetics of epilepsy that is, the opportunities for detecting genetic causes that exist in every epilepsy clinic. Furthermore, we discuss the beneficial influence that receiving a genetic diagnosis can have on epilepsy management. Genetic architecture of the epilepsies The aetiology of the genetic epilepsies was first decoded in familial monogenic diseases, and in disorders in which NATURE REVIEWS NEUROLOGY VOLUME 10 MAY

2 REVIEWS Key points Many forms of epilepsy have long been suspected to have a genetic background, and improvements in sequencing technology have now enabled detailed dissection of their genetic basis The likelihood of a genetic cause of epilepsy is easily underestimated in routine clinical practice De novo mutagenesis is increasingly being recognized as an important mechanism in some epilepsies, particularly the epileptic encephalopathies A genetic aetiology is not synonymous with generalized epilepsy; many focal epilepsies have a known genetic cause Genetic diagnosis can facilitate appropriate counselling and, in some patients, can also have therapeutic implications Neoplasm Stroke Trauma Birth anoxia Congenital Other lesions Infectious Idiopathic 1975 (Hauser & Kurland 15 ) Autoimmune Single-gene epilepsies: familial, de novo Epilepsies with complex inheritance 2014 paradigm Focal epilepsy with MRIdetectable lesions Modifiers and susceptibility alleles Figure 1 Advances in understanding the causes of epilepsy. a In 1975, the majority of epilepsies were characterized as idiopathic. b Today, epilepsy of unknown cause comprises a much smaller proportion, owing to the discovery of autoimmune epilepsies, epilepsies with lesions that are only detectable by MRI and, most importantly, the reclassification of many epilepsies previously considered idiopathic as having a genetic cause. The exact proportions of monogenic and complex or polygenic epilepsies remain uncertain. Data for part a were obtained from Hauser & Kurland. 15 seizures are a recognized but nonobligatory (comorbid) feature, such as the triplet repeat expansion in fragile X syndrome. 5 The molecular background of many Mendelian disorders that involve seizures including disorders associated with metabolic defects (such as phenylketonuria and pyridoxine deficiency 6 ) or structural changes (tuberous sclerosis 7 and familial cavernous angiomas 8 ) is now well understood. Even though these disorders are individually rare, understanding their molecular background has had important implications: a number of them are potentially treatable, and identifying the underlying disorder can help direct pharmacotherapy or surgery. However, the existence of Mendelian epilepsies that lack metabolic or structural markers, such as benign familial neonatal epilepsy, 9 has long been recognized. Contemporary studies have also revealed an increasing number of new and perhaps less obviously Mendelian epilepsies. These disorders usually have modest penetrance and, consequently, casual enquiry might fail to elicit important details from the family history. The majority of familial epilepsies, such as the genetic generalized epilepsies and familial temporal lobe epilepsies, do not show Mendelian inheritance. 10,11 Rather, the inheritance pattern is complex, indicating that the phenotype is determined by more than one gene, perhaps in interaction with environmental factors. For the genetic generalized epilepsies and familial temporal lobe epilepsies, evidence for environmental contributions to aetiology (as opposed to precipitation of seizures) has not yet emerged, although it cannot be excluded; the weight of evidence suggests that these are essentially oligogenic or polygenic disorders. 10,11 The clinical evidence for gene environment interactions in human epilepsy is robust, although the role of obstetric factors was previously overemphasized. 12 Most of the environmental factors have not been fully elucidated, but considerable evidence supports a role for gene environment interactions in determining the likelihood of developing epilepsy 12 following major brain insults as diverse as traumatic brain injury 13 and cerebral malaria. 14 The 2014 paradigm The classic treatise on epidemiology of epilepsy in a firstworld environment was published in 1975, 15 and also included information on its aetiology (Figure 1a). In the intervening years, further welcome detail has emerged on specific categories and causes of epilepsy, 16 but little else has changed, with the exception of the new class of autoimmune epilepsies (most commonly caused by antibodies directed against the N methyl d-aspartate receptor [NMDA] receptor or voltage-gated potassium channel proteins), 17,18 and the identification of subtle lesions by MRI. Of note, however, even epilepsies categoriz ed as acquired, such as those resulting from trauma, 13 stroke, 19 neoplasm, 20 infection 14 or congenital malformations, 21 are now known to have genetic contributions. The proportions of the aetiological categories that are attributed to genetic versus acquired factors best corres pond to those observed in the developed world; in other settings, endemic infections, such as neurocysticercosis, are an important (and often treatable) cause of epilepsy. 22 Notable ethnic differences, such as the elevated risk of febrile convulsions in Japan (6 9%, compared with 2 5% in European infants), are also evident. 23 A key difference between the 1975 paradigm and today s understanding of epilepsy is that we now have the confidence to leave the term idiopathic behind. 24 The label idiopathic epilepsy was too frequently used to pro vide the incorrect reassurance that a diagnosis had been made, while obfuscating the fact that the real cause was unknown. Importantly, the advent of MRI revealed lesions that could not be detected either clinically or by CT scans in at least 12% of adult patients with new-onset epilepsy. 25 Furthermore, although we are far from fully understanding the majority of epilepsies that would have been classed as unexplained in 1975, 15 it is disingenuous to state that we have no clues to their origin. The large sector that comprises the idiopathic epilepsies (Figure 1b) has, therefore, been transformed. Epidemiological studies revealed that genetic factors have a major role in these epilepsies, 4,26 28 and the results of clinical phenotyping and family studies should give clinicians the confidence to diagnose and counsel patients regarding the cause of their condition, even when the precise mutations are not yet known. 10,29 Several molecular genetic tests are already 284 MAY 2014 VOLUME 10

3 FOCUS ON EPILEPSY Table 1 Genetic testing in patients with epilepsy Genetic test Detects Indication Consequence Current relevance CGH array 42,43 Copy number variation Epilepsies with (comorbid) intellectual disability, autism or dysmorphic features Karyotyping 112,113 Large-scale chromosomal abnormality Suspected ring chromosome 20 Diagnosis Gene panel 49,65 Known mutations in (typically) genes HLA typing 109 Risk allele associated with hypersensitivity to carbamazepine Epileptic encephalopathies Probable Han Chinese or South Asian ethnic origin Diagnosis AED choice SCN1A 54,55 Mutations in sodium channel type 1α Dravet syndrome Diagnosis AED choice SLC2A Mutations in GLUT1 GLUT1 encephalopathy Early-onset childhood absence epilepsy Paroxysmal exertional dyskinesia PCDH19 61,92 Mutations in protocadherin 19 Female patients with clustered focal seizures (onset under 3 years of age) and often with intellectual disability Probable future relevance GRIN2A Mutations in NMDA receptor subunit ε1 Landau Kleffner syndrome Epilepsy aphasia spectrum disorders DEPDC5 69,70 Mutations in DEP domain-containing 5 Familial focal epilepsy with variable foci Most common mutation in focal epilepsy KCNT1 76,77 Mutations in potassium channel, subfamily T, member 1 Autosomal dominant nocturnal frontal lobe epilepsy Epilepsy in infancy with migrating focal seizures Ketogenic diet Diagnosis Seven widely available genetic tests have important management consequences that every neurologist should know about, and another three are likely to become of clinical importance soon. Abbreviations: AED, antiepileptic drug; CGH, comparative genomic hybridization; GLUT1, glucose transporter protein type 1; HLA, human leukocyte antigen; NMDA, N methyl d-aspartate receptor. in clinical use for diagnosis of epilepsy, and their number is rapidly growing (Table 1). Concomitantly, the proportion of patients with epilepsy of truly unknown aetiology has significantly shrunk. The new paradigm highlights four overlapping categories that describe the genetic architecture of epilepsy. At one extreme of the spectrum of genetic epilepsies are the monogenic syndromes, in which a single gene mutation (either inherited or de novo) with a large effect size essentially accounts for the phenotypic features. These syndromes include the Mendelian disorders in which seizures are an occasional or frequent comorbid feature, as well as the monogenic epilepsies such as Dravet syndrome (described in detail below) and benign familial neonatal epilepsy, both of which are paradigmatic examples. This continuum evolves into the complex epilepsies, including those with putative oligogenic or polygenic architecture 30 and little or no environmental contribution. This group includes the common electroclinical syndromes of genetic generalized epilepsy (such as childhood absence epilepsy and juvenile myoclonic epilepsy) and familial temporal lobe epilepsy. 10,31 Exome sequencing of large cohorts of people with genetic generalized epilepsy revealed that mutations in a single major unidentified gene were unlikely to account for these disorders, 32 even when the researchers focused specifically on finding mutations in genes encoding ion channel proteins. 33 Limitation of genetic analysis to scrutiny of protein-coding DNA (the exome) is a current pragmatic attempt to manage the enormity of genomic data, but the ENCODE consortium has also begun to investigate the intronic regions. 34 To date, only a few important examples of mutations associated with epilepsy occur in noncoding parts of the genome, such as the dodecamer repeat upstream of the transcription start site of cystatin B, which is the main cause of Unverricht Lundborg disease. 35 The paucity of currently known mutations that contribute to an individual s risk of developing an electroclinical syndrome have led some researchers to overlook compelling evidence of their heritability. 4,26 28 In many common diseases (including several with heritability estimates lower than those of epilepsies), robust evidence from genomewide association studies (GWAS) shows that common polymorphisms increase the risk of disease, although the individual attributable risks seem to be small. 36,37 To date, however, GWAS in epilepsy have been relatively limited and a clear picture of the role of common polymorphisms has not yet emerged The family and beyond If genetic factors are so important, why do most patients with epilepsy seem to lack an obvious family history of the disorder? At least two major scientific reasons account for this apparent paradox. First, in complex disorders the proportion of affected relatives is much lower than is observed in Mendelian disorders. For example, in fully penetrant, autosomal dominant disorders (Figure 2a), 50% of first-degree relatives are affected; whereas in complex dis orders, typically less than 5 10% of first-degree relatives have the disease phenotype. Second, for epilepsies caused by de novo mutations, which are increasingly recognized to be important, no family history of epilepsy NATURE REVIEWS NEUROLOGY VOLUME 10 MAY

4 REVIEWS a KCNQ2 b SCN1A (+/+) (+/+) (+/+) (m/+) (+/+) (+/+) (+/+) (m/+) (+/+) (m/+) (m/+) (+/+) (m/+) (+/+) (+/+) (+/+) (m/+) (+/+) (m/+) (+/+) (m/+) (m/+) (+/+) (+/+) (+/+) (+/+) Male Female Either sex Deceased c GLUT1 (m/+) (m/+) (+/+) (m/+) (m/+) (m/+) Individual heterozygous for mutation (+/+) Individual with two x x normal alleles Numbers indicate multiple individuals (m/+) d 15q13.3 deletion (m/+) (m/+) (m/+) (m/+) 2 2 (+/+) (m/+) 7 2 (m/+) (m/+) (m/+) (+/+) (m/+) (+/+) (+/+) (m/+) (+/+) Figure 2 Inheritance patterns and phenotypic heterogeneity in familial epilepsies. a Autosomal dominant inheritance with phenotypic homogeneity, such as in KCNQ2-related benign familial neonatal epilepsy. Blue denotes an individual with epilepsy. b De novo SCN1A mutation leading to Dravet syndrome. Blue denotes the affected child. c GLUT1-associated familial epilepsy (red, myoclonic astatic epilepsy; yellow, generalized epilepsy; green, early-onset absence epilepsy; purple, focal epilepsy 105 ). d 15q13.3 microdeletion is a risk factor rather than a cause of epilepsy: not all affected individuals have the deletion, and some unaffected individuals are carriers of it. 111 Blue represents individuals with juvenile myoclonic epilepsy. can be present by definition. However, a number of additional barriers can prevent us from fully appreciating the importance of genetic factors in people with epilepsy. These obstacles, for the most part, are created by failing to adequately ascer tain the family history and by not appreciating the complexity of epilepsy genetics. The hidden family history Obviously, inadequate inquiry into the wider family history will result in incomplete understanding; however, several potential pitfalls can be avoided. Failure to routinely ask about seizures in all family members of patients with epilepsy is often driven by underappreciation of the role of genetic factors in certain epilepsies, such as adult-onset focal epilepsy. Similarly, an inexact causal attribution, such as mistakenly ascribing an epileptic encephalopathy to birth trauma, might prevent thorough re-examination. Questioning a witness to determine the semiology of a blackout is common practice; however, similar questioning of the oldest living female relative (who tends to be the most enthusiastic and accurate custodian of family history) to reveal the most accurate family history is less common in adult practice. Young men in particular rarely have adequate awareness of their family history. Epilepsy, because of its intermittent nature and psychosocial consequences, is often covert. Seizures in the patient s older relatives might not be disclosed, at least in part because of excessive social stigma. This situation can be circumvented by asking about the family history at repeated intervals, and specifically asking the individual to speak to aunts and grandparents (Box 1). Inheritance patterns are easiest to establish in exceptionally large pedigrees. However, over-reliance on these rare examples, 41 despite their being a valuable and necessary research tool, can discourage us from recognizing the existence of a familial history in a more modestly sized family. Genomic surprises Some of the most important revelations in epilepsy research have challenged us to reorganize our heuristic models. For example, researchers have abandoned the search for a single gene to explain juvenile absence epilepsy. The implications of four particularly important and surprising findings copy number variants; de novo mutagenesis; the resurgence of genetic focal epilepsies; and the phenotypic heterogeneity of epilepsy syndromes associated with specific genes are discussed below. Copy number variants Each genome is littered with chromosomal rearrangements, particularly deletions or duplications of genetic information. By convention, a deletion or duplication larger than a kilobase is known as a copy number variant (CNV), but they can be many megabases in size. CNVs can be conceptualized into two groups: private and 286 MAY 2014 VOLUME 10

5 FOCUS ON EPILEPSY Box 1 Tips for ascertaining a full epilepsy family history Identify all seizures Febrile seizures in family members are often not recalled unless specifically requested, and a history of febrile seizures in older relatives is often not known to younger family members Single seizures and seizures in the context of alcohol withdrawal may not be known about by the whole family Nonconvulsive seizures may not be considered to be of sufficient importance to be reported Establish who is affected, and what part of the family Do not miss half-siblings who have been raised outside of the family home Where appropriate, discreetly enquire about potential uncertainties regarding paternity Medical knowledge might not be shared across the whole family; you might need to seek permission to speak with other branches of the family yourself Understand the family background Could the parents be consanguineous? Do the parents both come from a culturally isolated population? Do they self-identify as having a heritage that has been associated with a specific epilepsy syndrome? Is there a prominent history of miscarriages? Return to enquire about the family again Epilepsy syndromes evolve Covert stories come forward Younger relatives might develop epilepsy only after the initial discussion For a checklist for routine pedigree construction, see Scheffer et al. 114 recurrent. Private CNVs are rare, can occur anywhere across the genome, affect many genes and, when large, are usually of direct clinical relevance. 42,43 In contrast with private CNVs, recurrent CNVs occur at genomic hot spots that increase the likelihood of a range of disorders, including epilepsy. Recognizing the significance of the second group has imparted new knowledge about many neurological and psychiatric disorders Predicting the effects of contiguous gene deletions or duplications, however, is an inexact science, and supposedly pathogenic CNVs are occasionally inherited from seemingly unaffected parents (Figure 2d). CNV microarray analysis has now entered routine clinical use and has largely replaced karyotyping as a key genetic diagnosis tool, although karyotyping is still required for diagnosis of rare forms of epilepsy, such as ring chromosome 20 syndrome (Table 1). De novo mutagenesis The importance of de novo mutagenesis meaning that a child is affected, but the parents do not have the mutation in epilepsy has been a great surprise. The epileptic encephalopathies are prime examples of mono genic (or primarily monogenic) epilepsies, the genetic aetiology of which has been unravelled with unpre cedented success Although these disorders are rare at the population level, correct explanation of the cause of epileptic encephalopathies is critical. Removing false attribution from the parents consciousness, and thereby absolving them from blame (that vaccination, minor birth trauma or other early life events could have produced the syndrome) is important. 52,53 Unlike the inherited mutations with variable penetrance seen in large families affected by epilepsy, Dravet syndrome a paradigmatic epileptic encephalopathy is primarily caused by de novo mutations in gametes or at a very early stage of embryonic development The majority (around 80%) of cases of Dravet syndrome are caused by mutations in SCN1A, which encodes the α subunit of the type 1 voltage-gated sodium channel. 54,55 In about three-quarters of patients, the causative variants are in the paternal allele. 57 However, Dravet syndrome seems to be something of an exception. In other eponymous epileptic encephalopathies (such as Ohtahara, West and Lennox Gastaut syndromes), no single gene accounts for the majority of cases. Dravet syndrome also illustrates another key feature of inherited epilepsy, in that novel mutations are also reported in other seemingly unconnected genes. Occasionally, de novo mutations in GABRG2 and PCDH19 cause a Dravet-like presentation. 58,59 The number of genes in which de novo mutations can produce an epi leptic encephalopathy currently exceeds 30, with new genes identified on an almost monthly basis. 48,49 Although any gene mutation that explains an individual s epi lepsy is important, some genes (such as STXBP1, 60 PCDH19, 59,61 SCN2A 62 and CDKL5 48,49,63 ) are recurrently mutated. Recognition of these more-common disorders permits the identification of their characteristic clinical features, such as movement disorders and head stereotypies with STXBP1 mutations, 64 or clustered focal seiz ures with affective symptoms in female patients with PCDH19 mutations. 61 The challenge for genetic screening in routine clinical practice is the number of genes that can produce a similar phenotype. In a study of 264 individuals, only 29 had de novo mutations in the same gene as at least one other patient in the group (nine genes in total). 49 Genetic screening techniques are already in transition to the clinical domain, which is causing great excitement; 65 however, the danger is that any de novo mutation present will be overinterpreted as the probable cause of epilepsy. Until genotype data from a greater number of patients are published, and online databases of genotype phenotype correlations become available, epileptologists are reliant on nascent bioinformatics strategies to stratify possible target genes by their gene families, putative function of their product, and tolerance of polymorphism. 48,66 The resurgence of monogenic focal epilepsies Focal epilepsies have long been regarded as largely acquired disorders. Surprisingly, however, the first crop of gene discoveries in Mendelian epilepsies included mutations in CHRNA4 (which encodes the nicotinic receptor subunit) causing an autosomal dominant nocturnal frontal lobe epilepsy, 43 and mutations in LGI1 causing autosomal dominant epilepsy with auditory features. 67 These discoveries did not, however, open the sluice gates for the elucidation of all focal epilepsies. The epilepsy literature is littered with examples of genes or genetic variants that were initially heralded as contributing to causation, largely on the basis of early association studies, but research in the intervening years failed to provide complementary evidence of a pathogenetic role. 68 Massively parallel sequencing has facilitated the discovery of novel disease-linked genes, and the focal epilepsies NATURE REVIEWS NEUROLOGY VOLUME 10 MAY

6 REVIEWS are once again at the vanguard. A prime example of a gene with a role in focal epilepsies that does not encode an ion channel protein is DEPDC5. Nonsense and missense mutations in DEPDC5 are responsible for familial focal epilepsy with variable foci in seven of the eight published large pedigrees, but DEPDC5 mutations also explain around 12% of focal epilepsies in 10 smaller families. 69 A further six families in which DEPDC5 mutations were causing focal epilepsy were subsequently described. 70 Surprisingly, the protein encoded by DEPDC5 is involved in the mammalian target of rapamycin (mtor) path way, which is also implicated in tuberous sclerosis. 71 Specifically, DEPDC5 is a component of the GATOR1 com plex, which is a negative regulator of mtor complex 1. Following the unexpected finding that DEPDC5 is involved in the pathway associated with tuberous sclerosis, some patients with DEPDC5 mutations and cortical malformations have been identified. 72 The NMDA-type glutamate receptor subunit ε1, encoded by GRIN2A, has major importance in the epilepsy aphasia spectrum, including patients with mild focal epilepsy but no intellectual disability. Familial and trio (both parents, plus the affected child) studies have confirmed the importance of both de novo and inherited mutations in this setting Mutations within GRIN2A were identified (by three independent groups) in a small proportion of patients who had focal epilepsy with centrotemporal spikes Importantly, however, altered ε1 subunit function has also been connected to severe presentations of epilepsy aphasia spectrum disorders, such as Landau Kleffner syndrome and epilepsy with continuous spike-and-wave in slow-wave sleep The proportion of patients with typical or atypical benign epilepsy with centrotemporal spikes who harbour a GRIN2A mutation (0 5%) is much lower than the corresponding proportion of patients with Landau Kleffner syndrome (12 20%). Genes linked to multiple epilepsy phenotypes The general rule in clinical genetics has been that a sin gle gene was associated with a single disorder (and vice versa). This rule has been repeatedly broken in the epilepsies below, we highlight two recent examples. KCNT1 KCNT1 is an interesting gene that contributes to focal epilepsy. The sodium-activated potassium channel sub unit encoded by KCNT1 is widely expressed in the brain, particularly in the neurons of the frontal cortex, in keeping with its association with autosomal dominant noc turnal fron tal lobe epilepsy. 76 In addition to refractory seizures, affected family members exhibit notable intellec tual disability, behavioural problems and/or psychiatric features. KCNT1 mutations can also cause a quite different epilepsy phenotype: 77 epilepsy in infancy with migrating focal seizures, which was previously known as malignant migrating partial seizures of infancy. This severe epileptic encephalopathy has a very poor developmental outcome. Seizure onset takes place before 6 months of age and, in contrast to the fixed seizure focus observed in autosomal dominant nocturnal frontal lobe epilepsy, seizures occur apparently randomly across both hemispheres; furthermore, the brain shows an intraictal activation pattern, in which a seizure starts in one region and seems to migrate to another. The functional consequences of mutations in potassium channel genes that are associated with migrating focal seizures are more severe than those associated with autosomal dominant nocturnal frontal lobe epilepsy and, remarkably, the changes might be reversible by quinidine (see below 78 ). KCNQ2 KCNQ2 is the most commonly mutated gene in patients with benign familial neonatal epilepsy, 79,80 a disorder that has a favourable outcome in terms of seizure remission and intellectual function. In stark clinical contrast, KCNQ2 mutations are also associated with a severe encephalopathy that, like benign familial neonatal epilepsy, has a neo natal onset, but results in cognitive decline and sometimes refractory epilepsy that can resemble the electroclinical phenotype of Ohtahara syndrome. 81 KCNQ2 encephalopathy is typically associated with mutations that have severe functional effects, 82 but rare examples of families affected by both benign and severe forms of epilepsy due to a familial KCNQ2 mutation are known. 83 The most devastating forms of KCNQ2 epilepsy usually result from de novo mutagenesis. 82 Massively parallel sequencing The cost of sequencing the majority of protein-coding genes in the human genome has decreased to approximate that of assessing and interpreting the role of only one or two genes using traditional methods. This change has provoked a strategic shift. We are already moving towards routine use of massively parallel sequencing to address specific research questions, and incorporation of this technique into routine clinical investigation is on the horizon. 84,85 Another way to overcome the genetic complexity and heterogeneity of epilepsy is to use targeted panels. 49 The mutation detection rate depends on the selection of suitable patients, as well as the gene content of the panel. In one study using this approach, a panel of 256 genes relevant to epilepsy phenotypes was assessed, and potentially causative mutations were detected in 16 of 33 patients. 65 A persuasive argument can be made in favour of the use of whole exome sequencing rather than epilepsy-related gene panels. The advantage of whole-exome sequenc ing is that this technique does not predefine the set of genes to be studied, enabling the identification of genes that have no currently known function or have been previously (erroneously) discounted, as well as those suggest ing a pathogenetic role that had not previously been described. 86 Whole-exome sequencing will, however, present novel ethical, bio statistical and procedural challenges that epilepsy researchers as a community need to understand, investigate and address. 84,87 More over, the technology is far from a diagnostic panacea and, in the majority of patients, a single whole-exome study is currently uninterpretable, partly due to the vast degree of genomic variation seen even in healthy individuals. Additionally, even 288 MAY 2014 VOLUME 10

7 FOCUS ON EPILEPSY whole-exome sequencing is never fully comprehensive, as some parts of the genome are resistant to this type of analysis; certain exons are more difficult to sequence adequately, and regulatory sequences are not interrogated. As a result, most published studies share similar biases. Cur rently, which of the massively parallel sequencing techniques is the most advantageous depends on the clinical question being posed, as well as the specific health-care setting. The evolving role of clinicians Clinicians could soon be in the position of having accurate or unequivocal genomic information despite an inconclusive clinical picture. Whether it is possible to work backwards from genetic findings is currently unclear. In future, clinicians will increasingly need to deal with the uncertainties thrown up by these pioneering days of genomic personalized medicine, and will be responsible for interpreting genetic variations of uncertain significance or in genes of unknown relevance to the clinical context. The definitive clinical questions will be whether the patient s phenotype and genetic findings are in keeping with those of a typical presentation of the disorder, perhaps with some atypical features, and whether a given genetic (or phenotypic) finding is diagnostically relevant. Furthermore, identifying the specific phenotypes associated with mutations in some genes, such as SCN1A (Dravet syndrome and generalized epilepsy with febrile seizures plus [GEFS+]) and SLC2A1 (early-onset absence epilepsy and GLUT1 encephalo pathy) and differentiating them from the am orphous class of epilepsies will be of great clinical utility. Targeted management The reader might well ask what all this genetic information has done for patients with epilepsy. 88 We acknowledge that this question is borne of frustration that the majority of epilepsies do not yet have a recognizable cure. Although gene therapy is not yet a reality, a roadmap towards clini cal trials of vector-borne gene therapy for epilepsy 89 is out lined by Kullmann et al. in a Perspectives article in this issue. 90 Nonetheless, we should not overlook the areas of clinical practice where we have made demonstrable progress. The direct consequences for the individual and family of receiving a genetic diagnosis must be considered, even if the gene function and effects of the mutation have not been unravelled. Parents and/or patients are often relieved at finding a genetic cause, in part because self-blame for incorrect attribution of the epilepsy to something that they did (or did not do) can be erased. Moreover, individuals from a large family affected by epilepsy can experience complex feelings of guilt, whether or not they have epilepsy themselves. 91 Formalized counselling can be arranged once the genetic architecture and mode of inheritance of the epilepsy is fully understood. Epilepsy-specific genetic counselling is best administered by a specialist service, which is underused and not available in all health-care settings. For example, the advice given to the family of a young girl with epilepsy and intellectual disability with a PCDH19 mutation and that given to the family of a child with Dravet syndrome resulting from an SCN1A mutation can be starkly different (Table 1). PCDH19 mutations can be inherited in an X linked dominant fashion, such that male carriers transmit the affected allele only to their daughters (Figure 2). 92 PCDH19-associated epilepsy is, therefore, limited to female patients, and the degree of intellectual disability is variable. 93 Dravet syndrome is usually caused by a de novo mutation, but when counselling the families of such patients, it is important to know that in addition to genuinely novel mutagenesis, Dravet syndrome can be secondary to a low-level parental mosaicism in a minority of patients The ethical and practical aspects of genetic testing and counselling in epilepsies have been discussed in detail by Poduri et al. in a Perspectives article of this issue. 98 Surgery and neurostimulation The promise of identifying a specific genetic defect whets the appetite of clinicians and patients alike, who are attracted by the potential for development of a magic bullet therapeutic agent or at least a rationale for personalized selection of antiepileptic drugs. Epilepsies with a genetic background can, however, also be treated nonpharmacologically. For example, surgical resection of the tubers is possible in patients with tuberous sclerosis, and the success of localized removals in alleviating seizures is well-established. 99 Furthermore, in addition to its role in the immediate treatment of haemorrhage, targeted surgery can alleviate seizures in patients with familial cavernous haemangioma. Surgery can be useful even in some patients with nonlesional genetic epilepsies in which mutant proteins are likely to be widely expressed, such as temporal lobe epilepsy with hippocampal sclerosis associated with SCN1B mutations. 100 Pathway-directed therapies The discovery that classic epileptic encephalopathies, such as West syndrome and Lennox Gastaut syndrome, are particularly genetically heterogeneous 48 suggests that the genetic defects might converge on common pathways. Besides providing the opportunity to investigate therapies that reverse the effects of a particular mutation, this finding also encourages us to understand the aberrant network at genetic, cellular and neurophysiological levels and to devise pharmacological and nonpharmacological intervention strategies. The ketogenic diet The prominent publication of favourable results for a ran domized controlled trial of the ketogenic diet in 145 children 101 legitimized this important therapy. Signi ficant seizure reduction was seen in children with the most severe epilepsies; 38% of children on the diet had a more than 50% reduction in seizure frequency (compared with 6% of controls on a normal diet). The efficacy of the ketogenic diet varies across epilepsy syndromes; how ever, it should be considered early in the treatment of children with Dravet NATURE REVIEWS NEUROLOGY VOLUME 10 MAY

8 REVIEWS syndrome. 102 The ketogenic diet can also be surprisingly efficacious in other gen etic and acquired epilepsies, such as lissencephaly and hypoxic ischaemic encephalopathy. 101 Although the mechanisms through which the ketogenic diet ameliorates seizures are currently still being eluci dated, and selection of the patients who are most likely to benefit from a ketogenic diet is still being optimized, certain individuals with SLC2A1 mutations and GLUT1-deficiency disorders (that is, GLUT1-related encephalopathy and GLUT1-associated refractory epilepsy) should be considered as candidates for early trials of the ketogenic diet In addition to classical GLUT1-related encephalopathy, 106 the chance of identifying a patient with GLUT1- deficiency is enhanced by focusing on early-onset absence epilepsy (seizure onset under 4 years of age), where it has a frequency of 10% GLUT1 also provides a number of further important lessons: mutations can cause either absence epilepsy, paroxysmal exertional dyskinesia, or both. The movement disorder in individuals with GLUT1 deficiency can be so subtle that it is only recognized when the whole family is re-examined following genetic diagnos is of a clinically affected member. 105 Rapamycin analogues The remarkable finding that DEPDC5, a gene associated with familial focal epilepsies (see above), is linked to the mtor pathway 71 raises the possibility that rapamycin an alogues of which several are currently in various stages of clinical trials could one day be used to treat patients with these types of epilepsy, in addition to their established role in tuberous sclerosis. Quinidine Gain-of-function KCNT1 mutations can cause autosomal dominant nocturnal frontal lobe epilepsy and epilepsy in infancy with migrating focal seizures. A new in vitro study suggests that abnormally large currents, associated with gain-of-function mutation in KCNT1 channels, can be reversibly attenuated by quinidine, which is a KCNT1 channel blocker and a recognized yet superseded treatment for cardiac arrhythmia. 78 While these observations require confirmation, this strategy could provide a targeted therapy for two forms of epilepsy that are currently a challenge to treat. Drug response and pharmacogenomics The goal of pharmacogenomics is to predict how an individual will respond to a drug before it is prescribed. Clearly, the ability to make such predictions would be an attractive proposition for epilepsy; however, currently available tools for predicting seizure recurrence rates, despite elegant refinement, are fairly blunt. 107 In a GWAS of 889 individuals with newly diagnosed epilepsy, the researchers concluded that at the population level, no single variant could account for more than 5% of the variation in treatment outcomes. 38 A polygenic model, or a model in which rare mutations confer a strong likelihood of achieving seizure remission, remains plausible but unproven. Substantial progress has, however, been made in predicting adverse drug reactions. Finding a disease-associated SCN1A variant in a child with an epileptic encephalopathy can aid the selection of antiepileptic drugs, since carba mazepine and lamotrigine can potentially exacerbate seiz ures in individuals with Dravet syn drome. Conversely, valproate would be a beneficial choice, and stiripentol has superior efficacy (above even that of valproate and clobazam) in randomized controlled trials. 108 A very important finding is that in Han Chinese, and certain other ethnic groups of Southeast Asian origin, the HLA B*1502 allele confers a high risk of severe hypersensitivity reactions to carbamazepine. In Taiwan, 7.7% of people with epilepsy have the HLA B*1502 allele. Screening for this allele before starting therapy could potentially be cost-effective. Indeed, in a study of over 4,000 patients screened for HLA B*1502, in which those with the risk allele were advised to avoid carbamazepine, none of the participants developed either Stevens Johnson syndrome or toxic epidermal necrolysis, whereas 10 patients might otherwise have been expected (from his torical control data) to have developed these complications. This finding strongly argues for the power of pharmaco genomics at the population level. 109 In white people of northern European origin, the HLA A*3101 allele seems to confer an elevated risk of skin reactions to carbamaz epine, but the clinical utility of screening for this marker has not been established. 110 Conclusions Although much remains to be clarified, great advances have been made in understanding of the causes of epilepsy, particularly the forms previously categorized as idiopathic. Indeed, rather than the majority of epilepsies being of unknown cause, clinicians can now determine the cause of epilepsy in an appreciable proportion of their patients. This change is driven by a new appreciation of the importance of genetics, including the identification of causative genes in a minority of patients and powerful epidemiological evidence of genetic causation in many epilepsies, complemented by the appropriate use of highquality imaging. These clinical and molecular data lead us to assert that genetic variants offer a new paradigm for the aetiology of many epilepsies. Further genomic surprises might yet await us. The rewards of a thorough understanding of the genetic causes that underpin both common and rare epilepsies have already started to cascade into the clinical domain. This trickle could become a torrent when massively parallel sequencing becomes a routine diagnostic tool. Review criteria A search for original articles published between 1950 and 2014 and focusing on epilepsy was performed in MEDLINE and PubMed. The search terms used were gene, mutation, linkage and epidemiology, alone and in combination. All articles identified were Englishlanguage, full-text papers. We also searched the reference lists of identified articles for further relevant papers. 290 MAY 2014 VOLUME 10

9 FOCUS ON EPILEPSY 1. Reynolds, J. R. Epilepsy: Its Symptoms, Treatment, and Relation to Other Chronic Convulsive Diseases (Churchill, 1861). 2. Lennox, W. G. Should they live? Certain economic aspects of medicine. American Scholar 7, (1938). 3. Offen, M. L. Dealing with defectives. Foster Kennedy and William Lennox on eugenics. Neurology 61, (2003). 4. Lennox, W. G. The heredity of epilepsy as told by relatives and twins. JAMA 146, (1951). 5. Yu, S. et al. Fragile X genotype characterized by an unstable region of DNA. Science 252, (1991). 6. Mastrangelo, M., Celato, A. & Leuzzi, V. A diagnostic algorithm for the evaluation of early onset genetic-metabolic epileptic encephalopathies. Eur. J. Paediatr. Neurol. 16, (2012). 7. Crino, P. B., Nathanson, K. L. & Henske, E. P. The tuberous sclerosis complex. N. Engl. J. Med. 355, (2006). 8. Fischer, A., Zalvide, J., Faurobert, E., Albiges-Rizo, C. & Tournier-Lasserve, E. Cerebral cavernous malformations: from CCM genes to endothelial cell homeostasis. Trends Mol. Med. 19, (2013). 9. Rett, A., Teubel, R. Neugeborenenkrämpfe im Rahmen einer epileptisch belasteten Familie [German]. Wien. Klin. Wochenschr. 74, (1964). 10. Crompton, D. E. et al. Familial mesial temporal lobe epilepsy: a benign epilepsy syndrome showing complex inheritance. Brain 133, (2010). 11. Helbig, I., Scheffer, I. E., Mulley, J. C. & Berkovic, S. F. Navigating the channels and beyond: unravelling the genetics of the epilepsies. Lancet Neurol. 7, (2008). 12. Berkovic, S. F., Mulley, J. C., Scheffer, I. E. & Petrou, S. Human epilepsies: interaction of genetic and acquired factors. Trends Neurosci. 29, (2006). 13. Christensen, J. et al. Long-term risk of epilepsy after traumatic brain injury in children and young adults: a population-based cohort study. Lancet 373, (2009). 14. Kariuki, S. M. et al. The genetic risk of acute seizures in African children with falciparum malaria. Epilepsia 54, (2013). 15. Hauser, W. A. & Kurland, L. T. The epidemiology of epilepsy in Rochester, Minnesota, 1935 through Epilepsia. 16, 1 66 (1975). 16. Shorvon, S. D., Andermann, F. & Guerrini, R. (Eds) The Causes of Epilepsy: Common and Uncommon Causes in Adults and Children (Cambridge University Press, 2011). 17. Brenner, T. et al. Prevalence of neurologic autoantibodies in cohorts of patients with new and established epilepsy. Epilepsia 54, (2013). 18. Quek, A. M. et al. Autoimmune epilepsy: clinical characteristics and response to immunotherapy. Arch. Neurol. 69, (2012). 19. Dichgans, M. Genetics of ischaemic stroke. Lancet Neurol. 6, (2007). 20. Frattini, V. et al. The integrated landscape of driver genomic alterations in glioblastoma. Nat. Genet. 45, (2013). 21. Barkovich, A. J., Guerrini, R., Kuzniecky, R. I., Jackson, G. D. & Dobyns, W. B. A developmental and genetic classification for malformations of cortical development: update Brain 135, (2012). 22. Otte, W. M., Singla, M., Sander, J. W. & Singh, G. Drug therapy for solitary cysticercus granuloma: a systematic review and meta-analysis. Neurology. 80, (2013). 23. Tsuobi, T. Epidemiology of febrile and afebrile convulsions in children in Japan. Neurology 34, (1984). 24. Berg, A. T. et al. Revised terminology and concepts for organization of seizures and epilepsies: report of the ILAE Commission on Classification and Terminology, Epilepsia 51, (2010). 25. King, M. A. et al. Epileptology of the first-seizure presentation: a clinical, electroencephalographic, and magnetic resonance imaging study of 300 consecutive patients. Lancet 352, (1998). 26. Corey, L. A., Pellock, J. M., Kjeldsen, M. J. & Nakken, K. O. Importance of genetic factors in the occurrence of epilepsy syndrome type: a twin study. Epilepsy Res. 97, (2011). 27. Annegers, J. F., Hauser, W. A., Anderson, V. E. & Kurland, L. T. The risks of seizure disorders among relatives of patients with childhood onset epilepsy. Neurology. 32, (1982). 28. Ottman, R. Genetic epidemiology of epilepsy. Epidemiol. Rev. 19, (1997). 29. Peljto, A. L. et al., Familial risk of epilepsy: a population-based study. Brain 137, (2014). 30. Dibbens, L. M., Heron, S. E. & Mulley, J. C. A polygenic heterogeneity model for common epilepsies with complex genetics. Genes Brain Behav. 6, (2007). 31. [No author listed] Concordance of clinical forms of epilepsy in families with several affected members. Italian League Against Epilepsy Genetic Collaborative Group. Epilepsia 34, (1993). 32. Heinzen, E. L. et al. Exome sequencing followed by large-scale genotyping fails to identify single rare variants of large effect in idiopathic generalized epilepsy. Am. J. Hum. Genet. 91, (2012). 33. Klassen, T. et al. Exome sequencing of ion channel genes reveals complex profiles confounding personal risk assessment in epilepsy. Cell 145, (2011). 34. The ENCODE Project Consortium. Identification and analysis of functional elements in 1% of the human genome by the ENCODE pilot project. Nature 447, (2007). 35. Pennacchio, L. A. et al. Mutations in the gene encoding cystatin B in progressive myoclonus epilepsy (EPM1). Science 271, (1996). 36. Cross-Disorder Group of the Psychiatric Genomics Consortium & Genetic Risk Outcome of Psychosis (GROUP) Consortium. Identification of risk loci with shared effects on five major psychiatric disorders: a genome-wide analysis. Lancet 381, (2013). 37. The International Multiple Sclerosis Genetics Consortium and The Wellcome Trust Case Control Consortium 2. Genetic risk and a primary role for cell-mediated immune mechanisms in multiple sclerosis. Nature 476, (2011). 38. Speed, D. et al. A genome-wide association study and biological pathway analysis of epilepsy prognosis in a prospective cohort of newly treated epilepsy. Hum. Mol. Genet. 23, (2014). 39. EPICURE Consortium & EMINet Consortium. Genome-wide association analysis of genetic generalized epilepsies implicates susceptibility loci at 1q43, 2p16.1, 2q22.3 and 17q Hum. Mol. Genet. 21, (2012). 40. Kasperaviciūte, D. et al. Common genetic variation and susceptibility to partial epilepsies: a genome-wide association study. Brain 133, (2010). 41. Steinlein, O. K. et al. A missense mutation in the neuronal nicotinic acetylcholine receptor α4 subunit is associated with autosomal dominant nocturnal frontal lobe epilepsy. Nat. Genet. 11, (1995). 42. Mefford, H. C. et al., Rare copy number variants are an important cause of epileptic encephalopathies. Ann. Neurol. 70, (2011). 43. Mulley, J. C. & Mefford, H. C. Epilepsy and the new cytogenetics. Epilepsia 52, (2011). 44. Sebat, J. et al. Strong association of de novo copy number mutations with autism. Science 316, (2007). 45. de Kovel, C. G. et al. Recurrent microdeletions at 15q11.2 and 16p13.11 predispose to idiopathic generalized epilepsies. Brain 133, (2010). 46. International Schizophrenia Consortium. Rare chromosomal deletions and duplications increase risk of schizophrenia. Nature 455, (2008). 47. Mefford, H. C. et al. Genome-wide copy number variation in epilepsy: novel susceptibility loci in idiopathic generalized and focal epilepsies. PLoS Genet. 6, e (2010). 48. Epi4K Consortium & Epilepsy Phenome/Genome Project. De novo mutations in epileptic encephalopathies. Nature 501, (2013). 49. Carvill, G. L. et al. Targeted resequencing in epileptic encephalopathies identifies de novo mutations in CHD2 and SYNGAP1. Nat. Genet. 45, (2013). 50. Kodera, H. Targeted capture and sequencing for detection of mutations causing early onset epileptic encephalopathy. Epilepsia 54, (2013). 51. Veeramah, K. R. et al. Exome sequencing reveals new causal mutations in children with epileptic encephalopathies. Epilepsia 54, (2013). 52. McIntosh, A. M. et al. Effects of vaccination on onset and outcome of Dravet syndrome: a retrospective study. Lancet Neurol. 9, (2010). 53. Berkovic, S. F. et al. De-novo mutations of the sodium channel gene SCN1A in alleged vaccine encephalopathy: a retrospective study. Lancet Neurol. 5, (2006). 54. Claes, L. et al. De novo SCN1A mutations are a major cause of severe myoclonic epilepsy of infancy. Hum. Mutat. 21, (2003). 55. Harkin, L. A. et al. The spectrum of SCN1Arelated infantile epileptic encephalopathies. Brain 130, (2007). 56. Vadlamudi, L. et al. Timing of de novo mutagenesis a twin study of sodium-channel mutations. N. Engl. J. Med. 363, (2010). 57. Heron, S. E. et al. De novo SCN1A mutations in Dravet syndrome and related epileptic encephalopathies are largely of paternal origin. J. Med. Genet. 47, (2010). 58. Harkin, L. A. et al. Truncation of the GABA A - receptor γ2 subunit in a family with generalized epilepsy with febrile seizures plus. Am. J. Hum. Genet. 70, (2002). 59. Depienne, C. et al. Sporadic infantile epileptic encephalopathy caused by mutations in PCDH19 resemble Dravet syndrome but mainly affects females. PLoS Genet. 5, e (2009). 60. Deprez, L. et al. Clinical spectrum of early-onset epileptic encephalopathies associated with STXBP1 mutations. Neurology 75, (2010). NATURE REVIEWS NEUROLOGY VOLUME 10 MAY