Hunt, Robert F

Lim, Byung Chan

English

eScholarship - University of California

UC IrvineUC Irvine Previously Published WorksTitleEpilepsy Therapy Goes ViralPermalinkhttps://escholarship.org/uc/item/1qr3p832JournalEpilepsy Currents, 19(5)ISSN1535-7597AuthorsLim, Byung ChanHunt, Robert FPublication Date2019-09-01DOI10.1177/1535759719870506Copyright InformationThis work is made available under the terms of a Creative Commons Attribution License, availalbe at https://creativecommons.org/licenses/by/4.0/ Peer reviewedeScholarship.org Powered by the California Digital LibraryUniversity of CaliforniaCurrent Literaturein Basic ScienceEpilepsy Therapy Goes ViralEpilepsy Gene Therapy Using an Engineered Potassium ChannelSnowball A, Chabrol E, Wykes RC, Shekh-Ahmad T, Cornford JH, Lieb A, Hughes MP, Massaro G, Rahim AA, Hashemi KS,Kullmann DM, Walker MC, Schorge S. J Neurosci. 2019;39(16):3159-3169.Refractory focal epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy representsa promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must becarefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vectoroptimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon opti-mized for human expression and mutated to accelerate the recovery of the channels from inactivation. For improved safety,this engineered potassium channel (EKC) gene was packaged into a nonintegrating lentiviral vector under the control of a celltype-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled preclinical trial, the EKC lentivector robustlyreduced seizure frequency in a male rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures.When packaged into an adeno-associated viral vector (AAV2/9), the EKC gene was also effective at suppressing seizures in amale rat model of temporal lobe epilepsy. This demonstration of efficacy in a clinically relevant setting, combined with theimproved safety conferred by cell type–specific expression and integration-deficient delivery, identifies EKC gene therapy asbeing ready for clinical translation in the treatment of refractory focal epilepsy.Keywordsepilepsy, virus, gene therapy, KCNA1, neocortical focal epilepsyCommentaryGene therapy has been envisioned since the 1970s but has faceda number of difficult obstacles, both in the laboratory and theclinic. Following early success of a clinical trial for adenosinedeaminase - severe combined immunodeficiency (ADA-SCID)in 1990, subsequent clinical failures quickly led to dismissal ofgene therapy as nothing more than an experimental technique.However, over the last 15 years, there have been dramaticimprovements in human genetics, genome editing strategies,methods for gene delivery, and disease modeling, all of whichhas made this innovative therapeutic strategy move at anincreasingly rapid pace toward every day clinical practice.1This has led to a renewed interest in the promise of gene ther-apy for a wide range of human diseases, highlighted by a recentFood and Drug Administration approval for the treatment ofspinal muscular atrophy.2 However, despite many convincingadvances in the field, antiepileptic gene therapy remains at apreclinical proof-of-principle stage. There are a number ofimportant challenges that must be overcome in order to movethis exciting technology to the clinic for epilepsy, such aschoice of gene or pathway to target, viral packaging,off-target effects, successful delivery into the human brain,safety, and even the inherent heterogeneous etiology of epilepsyitself. With potentially more than 100 genes reported to causeepilepsy,3 many of which are expressed throughout the brainand/or body, it is difficult to envision applying the traditionalconcept of gene therapy (ie, to fix a defective gene or pathway)for each individual monogenetic disorder. In contrast,approaches that target more downstream pathways to manipulatethe expression of genes that control neuronal or circuit excitabil-ity in affected brain areas may have broader application.4Snowball and colleagues built upon their prior work5,6developing a viral vector–mediated gene therapy to alter neu-ronal and circuit excitability in focal neocortical epilepsy. Thisapproach involves overexpression of the human KCNA1 gene,which encodes the voltage-gated potassium channel Kv1.1, inepileptogenic foci. Here, the authors engineered and optimizedthe viral construct used in earlier studies with the goal of gen-erating an effective gene therapy that might be ready for clin-ical translation. To do this, they first replicated and extendedEpilepsy Currents2019, Vol. 19(5) 328-329ª The Author(s) 2019Article reuse guidelines:sagepub.com/journals-permissionsDOI: 10.1177/1535759719870506journals.sagepub.com/home/epiCreative Commons Non Commercial No Derivs CC BY-NC-ND: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 License (http://www.creativecommons.org/licenses/by-nc-nd/4.0/) which permits non-commercial use, reproduction anddistribution of the work as published without adaptation or alteration, without further permission provided the original work is attributed as specified on theSAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).EPILEPSY CURRENTStheir prior work by demonstrating a dramatic suppression ofspontaneous seizures following the delivery of a CMV-drivenKCNA1 lentivirus directly into the seizure focus in visual cor-tex of a tetanus toxin model. This construct was then modifiedin 3 main ways: (1) the non-cell-type-specific CMV promotorwas exchanged for a CAMK2A promotor to drive Kv1.1 expres-sion in excitatory projection neurons, (2) the Kv1.1 gene wasmodified to reduce its inactivation, and (3) the resulting plas-mid was packaged into a nonintegrating lentiviral vector toreduce the potential risk of mutagenesis. This new, refinedconstruct had even more powerful therapeutic effects on sei-zures in the tetanus toxin model of focal epilepsy, presumablydue to increased specificity for targeting excitatory neurons,and it was also effective in a kainic acid model of temporallobe epilepsy, when packaged into an AAV vector and injectedbilaterally into hippocampus.Neocortical epilepsy is often drug-resistant, and surgery toremove the epileptogenic zone is not always feasible or effec-tive. As such, development of a spatially restricted, cell-type-specific gene therapy that selectively modifies the activity ofneurons at the seizure focus could be an important advance. Ifsuccessful, this strategy might also spare unaffected brainregions or minimize the types of side effects seen in currentpharmacological approaches. Although the results reported bySnowball and colleagues are promising, additional hurdlesremain before eventual translation to the clinic. For example,the authors only evaluated the effect of this gene therapy onseizures, and it is not yet known how overexpression of Kv1.1in CAMK2A neurons might alter visual cortex function. Inthinking about moving from rodent to human, large animalstudies may be necessary for determining how to preciselydeliver the vector to epileptic foci of a gyrated neocortex.Ultimately, for clinical trials to succeed, one must decidewho will be an appropriate candidate for therapy. A patientwith drug-resistant epilepsy and a well-defined focal lesion ineloquent cortex has been considered to be an ideal candidate.The most prevalent pathologic substrate of epilepsy surgery isfocal cortical dysplasia, which has been identified in both mag-netic resonance imaging positive and negative cases.7However, if a gene therapy is effective and safe, and can bebroadly applied to many forms of the disease as indicated in thecurrent study, it may be advantageous to develop this technol-ogy for a wider range of patients regardless of whether or not afocal epileptogenic zone is present. It is possible that such atherapy could be useful in acquired epilepsy, such as followinga traumatic brain injury or stroke, as well as in genetic epilep-sies. As such, this study by Snowball and colleagues providesan important step closer to achieving successful gene therapyfor epilepsy.By Byung Chan Lim and Robert F. HuntORCID iDRobert F. Hunt https://orcid.org/0000-0003-4490-8718References1. Dunbar CE, High KA, Joung JK, Kohn DB, Ozawa K, Sadelain M.Gene therapy comes of age. Science. 2018;359(6372):eaan4672.2. Mendell JR, Al-Zaidy S, Shell R, et al. Single-dose gene-replacement therapy for spinal muscular atrophy. N Engl J Med.2017;377(18):1713-1722.3. McTague A, Howell KB, Cross JH, Kurian MA, Scheffer IE. Thegenetic landscape of the epileptic encephalopathies of infancy andchildhood. Lancet Neurol. 2016;15(3):304-316.4. Kullmann DM, Schorge S, Walker MC, Wykes RC. Gene therapyin epilepsy-is it time for clinical trials? Nat Rev Neurol. 2014;10(5):300-304.5. Heeroma JH, Henneberger C, Rajakulendran S, Hanna MG,Schorge S, Kullmann DM. Episodic ataxia type 1 mutations differ-entially affect neuronal excitability and transmitter release. DisModel Mech. 2009;2(11-12):612-619.6. Wykes RC, Heeroma JH, Mantoan L, et al. Optogenetic and potas-sium channel gene therapy in a rodent model of focal neocorticalepilepsy. Sci Transl Med. 2012;4(161):161.7. Ryvlin P, Cross JH, Rheims S. Epilepsy surgery in children andadults. Lancet Neurol. 2014;13(11):1114-1126.Commentary 329

Epilepsy Therapy Goes Viral

UC IrvineUC Irvine Previously Published WorksTitleEpilepsy Therapy Goes Viral.Permalinkhttps://escholarship.org/uc/item/8n22m3w4JournalEpilepsy currents, 19(5)ISSN1535-7597AuthorsLim, Byung ChanHunt, Robert FPublication Date2019-09-01DOI10.1177/1535759719870506Copyright InformationThis work is made available under the terms of a Creative Commons Attribution License, availalbe at https://creativecommons.org/licenses/by/4.0/ Peer reviewedeScholarship.org Powered by the California Digital LibraryUniversity of CaliforniaCurrent Literaturein Basic ScienceEpilepsy Therapy Goes ViralEpilepsy Gene Therapy Using an Engineered Potassium ChannelSnowball A, Chabrol E, Wykes RC, Shekh-Ahmad T, Cornford JH, Lieb A, Hughes MP, Massaro G, Rahim AA, Hashemi KS,Kullmann DM, Walker MC, Schorge S. J Neurosci. 2019;39(16):3159-3169.Refractory focal epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy representsa promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must becarefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vectoroptimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon opti-mized for human expression and mutated to accelerate the recovery of the channels from inactivation. For improved safety,this engineered potassium channel (EKC) gene was packaged into a nonintegrating lentiviral vector under the control of a celltype-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled preclinical trial, the EKC lentivector robustlyreduced seizure frequency in a male rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures.When packaged into an adeno-associated viral vector (AAV2/9), the EKC gene was also effective at suppressing seizures in amale rat model of temporal lobe epilepsy. This demonstration of efficacy in a clinically relevant setting, combined with theimproved safety conferred by cell type–specific expression and integration-deficient delivery, identifies EKC gene therapy asbeing ready for clinical translation in the treatment of refractory focal epilepsy.Keywordsepilepsy, virus, gene therapy, KCNA1, neocortical focal epilepsyCommentaryGene therapy has been envisioned since the 1970s but has faceda number of difficult obstacles, both in the laboratory and theclinic. Following early success of a clinical trial for adenosinedeaminase - severe combined immunodeficiency (ADA-SCID)in 1990, subsequent clinical failures quickly led to dismissal ofgene therapy as nothing more than an experimental technique.However, over the last 15 years, there have been dramaticimprovements in human genetics, genome editing strategies,methods for gene delivery, and disease modeling, all of whichhas made this innovative therapeutic strategy move at anincreasingly rapid pace toward every day clinical practice.1This has led to a renewed interest in the promise of gene ther-apy for a wide range of human diseases, highlighted by a recentFood and Drug Administration approval for the treatment ofspinal muscular atrophy.2 However, despite many convincingadvances in the field, antiepileptic gene therapy remains at apreclinical proof-of-principle stage. There are a number ofimportant challenges that must be overcome in order to movethis exciting technology to the clinic for epilepsy, such aschoice of gene or pathway to target, viral packaging,off-target effects, successful delivery into the human brain,safety, and even the inherent heterogeneous etiology of epilepsyitself. With potentially more than 100 genes reported to causeepilepsy,3 many of which are expressed throughout the brainand/or body, it is difficult to envision applying the traditionalconcept of gene therapy (ie, to fix a defective gene or pathway)for each individual monogenetic disorder. In contrast,approaches that target more downstream pathways to manipulatethe expression of genes that control neuronal or circuit excitabil-ity in affected brain areas may have broader application.4Snowball and colleagues built upon their prior work5,6developing a viral vector–mediated gene therapy to alter neu-ronal and circuit excitability in focal neocortical epilepsy. Thisapproach involves overexpression of the human KCNA1 gene,which encodes the voltage-gated potassium channel Kv1.1, inepileptogenic foci. Here, the authors engineered and optimizedthe viral construct used in earlier studies with the goal of gen-erating an effective gene therapy that might be ready for clin-ical translation. To do this, they first replicated and extendedEpilepsy Currents2019, Vol. 19(5) 328-329ª The Author(s) 2019Article reuse guidelines:sagepub.com/journals-permissionsDOI: 10.1177/1535759719870506journals.sagepub.com/home/epiCreative Commons Non Commercial No Derivs CC BY-NC-ND: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 License (http://www.creativecommons.org/licenses/by-nc-nd/4.0/) which permits non-commercial use, reproduction anddistribution of the work as published without adaptation or alteration, without further permission provided the original work is attributed as specified on theSAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).EPILEPSY CURRENTStheir prior work by demonstrating a dramatic suppression ofspontaneous seizures following the delivery of a CMV-drivenKCNA1 lentivirus directly into the seizure focus in visual cor-tex of a tetanus toxin model. This construct was then modifiedin 3 main ways: (1) the non-cell-type-specific CMV promotorwas exchanged for a CAMK2A promotor to drive Kv1.1 expres-sion in excitatory projection neurons, (2) the Kv1.1 gene wasmodified to reduce its inactivation, and (3) the resulting plas-mid was packaged into a nonintegrating lentiviral vector toreduce the potential risk of mutagenesis. This new, refinedconstruct had even more powerful therapeutic effects on sei-zures in the tetanus toxin model of focal epilepsy, presumablydue to increased specificity for targeting excitatory neurons,and it was also effective in a kainic acid model of temporallobe epilepsy, when packaged into an AAV vector and injectedbilaterally into hippocampus.Neocortical epilepsy is often drug-resistant, and surgery toremove the epileptogenic zone is not always feasible or effec-tive. As such, development of a spatially restricted, cell-type-specific gene therapy that selectively modifies the activity ofneurons at the seizure focus could be an important advance. Ifsuccessful, this strategy might also spare unaffected brainregions or minimize the types of side effects seen in currentpharmacological approaches. Although the results reported bySnowball and colleagues are promising, additional hurdlesremain before eventual translation to the clinic. For example,the authors only evaluated the effect of this gene therapy onseizures, and it is not yet known how overexpression of Kv1.1in CAMK2A neurons might alter visual cortex function. Inthinking about moving from rodent to human, large animalstudies may be necessary for determining how to preciselydeliver the vector to epileptic foci of a gyrated neocortex.Ultimately, for clinical trials to succeed, one must decidewho will be an appropriate candidate for therapy. A patientwith drug-resistant epilepsy and a well-defined focal lesion ineloquent cortex has been considered to be an ideal candidate.The most prevalent pathologic substrate of epilepsy surgery isfocal cortical dysplasia, which has been identified in both mag-netic resonance imaging positive and negative cases.7However, if a gene therapy is effective and safe, and can bebroadly applied to many forms of the disease as indicated in thecurrent study, it may be advantageous to develop this technol-ogy for a wider range of patients regardless of whether or not afocal epileptogenic zone is present. It is possible that such atherapy could be useful in acquired epilepsy, such as followinga traumatic brain injury or stroke, as well as in genetic epilep-sies. As such, this study by Snowball and colleagues providesan important step closer to achieving successful gene therapyfor epilepsy.By Byung Chan Lim and Robert F. HuntORCID iDRobert F. Hunt https://orcid.org/0000-0003-4490-8718References1. Dunbar CE, High KA, Joung JK, Kohn DB, Ozawa K, Sadelain M.Gene therapy comes of age. Science. 2018;359(6372):eaan4672.2. Mendell JR, Al-Zaidy S, Shell R, et al. Single-dose gene-replacement therapy for spinal muscular atrophy. N Engl J Med.2017;377(18):1713-1722.3. McTague A, Howell KB, Cross JH, Kurian MA, Scheffer IE. Thegenetic landscape of the epileptic encephalopathies of infancy andchildhood. Lancet Neurol. 2016;15(3):304-316.4. Kullmann DM, Schorge S, Walker MC, Wykes RC. Gene therapyin epilepsy-is it time for clinical trials? Nat Rev Neurol. 2014;10(5):300-304.5. Heeroma JH, Henneberger C, Rajakulendran S, Hanna MG,Schorge S, Kullmann DM. Episodic ataxia type 1 mutations differ-entially affect neuronal excitability and transmitter release. DisModel Mech. 2009;2(11-12):612-619.6. Wykes RC, Heeroma JH, Mantoan L, et al. Optogenetic and potas-sium channel gene therapy in a rodent model of focal neocorticalepilepsy. Sci Transl Med. 2012;4(161):161.7. Ryvlin P, Cross JH, Rheims S. Epilepsy surgery in children andadults. Lancet Neurol. 2014;13(11):1114-1126.Commentary 329

Epilepsy Therapy Goes Viral.

UC IrvineUC Irvine Previously Published WorksTitleEpilepsy Therapy Goes Viral.Permalinkhttps://escholarship.org/uc/item/99b9g1vhJournalEpilepsy currents, 19(5)ISSN1535-7597AuthorsLim, Byung ChanHunt, Robert FPublication Date2019-09-01DOI10.1177/1535759719870506Copyright InformationThis work is made available under the terms of a Creative Commons Attribution License, availalbe at https://creativecommons.org/licenses/by/4.0/ Peer reviewedeScholarship.org Powered by the California Digital LibraryUniversity of CaliforniaCurrent Literaturein Basic ScienceEpilepsy Therapy Goes ViralEpilepsy Gene Therapy Using an Engineered Potassium ChannelSnowball A, Chabrol E, Wykes RC, Shekh-Ahmad T, Cornford JH, Lieb A, Hughes MP, Massaro G, Rahim AA, Hashemi KS,Kullmann DM, Walker MC, Schorge S. J Neurosci. 2019;39(16):3159-3169.Refractory focal epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy representsa promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must becarefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vectoroptimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon opti-mized for human expression and mutated to accelerate the recovery of the channels from inactivation. For improved safety,this engineered potassium channel (EKC) gene was packaged into a nonintegrating lentiviral vector under the control of a celltype-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled preclinical trial, the EKC lentivector robustlyreduced seizure frequency in a male rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures.When packaged into an adeno-associated viral vector (AAV2/9), the EKC gene was also effective at suppressing seizures in amale rat model of temporal lobe epilepsy. This demonstration of efficacy in a clinically relevant setting, combined with theimproved safety conferred by cell type–specific expression and integration-deficient delivery, identifies EKC gene therapy asbeing ready for clinical translation in the treatment of refractory focal epilepsy.Keywordsepilepsy, virus, gene therapy, KCNA1, neocortical focal epilepsyCommentaryGene therapy has been envisioned since the 1970s but has faceda number of difficult obstacles, both in the laboratory and theclinic. Following early success of a clinical trial for adenosinedeaminase - severe combined immunodeficiency (ADA-SCID)in 1990, subsequent clinical failures quickly led to dismissal ofgene therapy as nothing more than an experimental technique.However, over the last 15 years, there have been dramaticimprovements in human genetics, genome editing strategies,methods for gene delivery, and disease modeling, all of whichhas made this innovative therapeutic strategy move at anincreasingly rapid pace toward every day clinical practice.1This has led to a renewed interest in the promise of gene ther-apy for a wide range of human diseases, highlighted by a recentFood and Drug Administration approval for the treatment ofspinal muscular atrophy.2 However, despite many convincingadvances in the field, antiepileptic gene therapy remains at apreclinical proof-of-principle stage. There are a number ofimportant challenges that must be overcome in order to movethis exciting technology to the clinic for epilepsy, such aschoice of gene or pathway to target, viral packaging,off-target effects, successful delivery into the human brain,safety, and even the inherent heterogeneous etiology of epilepsyitself. With potentially more than 100 genes reported to causeepilepsy,3 many of which are expressed throughout the brainand/or body, it is difficult to envision applying the traditionalconcept of gene therapy (ie, to fix a defective gene or pathway)for each individual monogenetic disorder. In contrast,approaches that target more downstream pathways to manipulatethe expression of genes that control neuronal or circuit excitabil-ity in affected brain areas may have broader application.4Snowball and colleagues built upon their prior work5,6developing a viral vector–mediated gene therapy to alter neu-ronal and circuit excitability in focal neocortical epilepsy. Thisapproach involves overexpression of the human KCNA1 gene,which encodes the voltage-gated potassium channel Kv1.1, inepileptogenic foci. Here, the authors engineered and optimizedthe viral construct used in earlier studies with the goal of gen-erating an effective gene therapy that might be ready for clin-ical translation. To do this, they first replicated and extendedEpilepsy Currents2019, Vol. 19(5) 328-329ª The Author(s) 2019Article reuse guidelines:sagepub.com/journals-permissionsDOI: 10.1177/1535759719870506journals.sagepub.com/home/epiCreative Commons Non Commercial No Derivs CC BY-NC-ND: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 License (http://www.creativecommons.org/licenses/by-nc-nd/4.0/) which permits non-commercial use, reproduction anddistribution of the work as published without adaptation or alteration, without further permission provided the original work is attributed as specified on theSAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).EPILEPSY CURRENTStheir prior work by demonstrating a dramatic suppression ofspontaneous seizures following the delivery of a CMV-drivenKCNA1 lentivirus directly into the seizure focus in visual cor-tex of a tetanus toxin model. This construct was then modifiedin 3 main ways: (1) the non-cell-type-specific CMV promotorwas exchanged for a CAMK2A promotor to drive Kv1.1 expres-sion in excitatory projection neurons, (2) the Kv1.1 gene wasmodified to reduce its inactivation, and (3) the resulting plas-mid was packaged into a nonintegrating lentiviral vector toreduce the potential risk of mutagenesis. This new, refinedconstruct had even more powerful therapeutic effects on sei-zures in the tetanus toxin model of focal epilepsy, presumablydue to increased specificity for targeting excitatory neurons,and it was also effective in a kainic acid model of temporallobe epilepsy, when packaged into an AAV vector and injectedbilaterally into hippocampus.Neocortical epilepsy is often drug-resistant, and surgery toremove the epileptogenic zone is not always feasible or effec-tive. As such, development of a spatially restricted, cell-type-specific gene therapy that selectively modifies the activity ofneurons at the seizure focus could be an important advance. Ifsuccessful, this strategy might also spare unaffected brainregions or minimize the types of side effects seen in currentpharmacological approaches. Although the results reported bySnowball and colleagues are promising, additional hurdlesremain before eventual translation to the clinic. For example,the authors only evaluated the effect of this gene therapy onseizures, and it is not yet known how overexpression of Kv1.1in CAMK2A neurons might alter visual cortex function. Inthinking about moving from rodent to human, large animalstudies may be necessary for determining how to preciselydeliver the vector to epileptic foci of a gyrated neocortex.Ultimately, for clinical trials to succeed, one must decidewho will be an appropriate candidate for therapy. A patientwith drug-resistant epilepsy and a well-defined focal lesion ineloquent cortex has been considered to be an ideal candidate.The most prevalent pathologic substrate of epilepsy surgery isfocal cortical dysplasia, which has been identified in both mag-netic resonance imaging positive and negative cases.7However, if a gene therapy is effective and safe, and can bebroadly applied to many forms of the disease as indicated in thecurrent study, it may be advantageous to develop this technol-ogy for a wider range of patients regardless of whether or not afocal epileptogenic zone is present. It is possible that such atherapy could be useful in acquired epilepsy, such as followinga traumatic brain injury or stroke, as well as in genetic epilep-sies. As such, this study by Snowball and colleagues providesan important step closer to achieving successful gene therapyfor epilepsy.By Byung Chan Lim and Robert F. HuntORCID iDRobert F. Hunt https://orcid.org/0000-0003-4490-8718References1. Dunbar CE, High KA, Joung JK, Kohn DB, Ozawa K, Sadelain M.Gene therapy comes of age. Science. 2018;359(6372):eaan4672.2. Mendell JR, Al-Zaidy S, Shell R, et al. Single-dose gene-replacement therapy for spinal muscular atrophy. N Engl J Med.2017;377(18):1713-1722.3. McTague A, Howell KB, Cross JH, Kurian MA, Scheffer IE. Thegenetic landscape of the epileptic encephalopathies of infancy andchildhood. Lancet Neurol. 2016;15(3):304-316.4. Kullmann DM, Schorge S, Walker MC, Wykes RC. Gene therapyin epilepsy-is it time for clinical trials? Nat Rev Neurol. 2014;10(5):300-304.5. Heeroma JH, Henneberger C, Rajakulendran S, Hanna MG,Schorge S, Kullmann DM. Episodic ataxia type 1 mutations differ-entially affect neuronal excitability and transmitter release. DisModel Mech. 2009;2(11-12):612-619.6. Wykes RC, Heeroma JH, Mantoan L, et al. Optogenetic and potas-sium channel gene therapy in a rodent model of focal neocorticalepilepsy. Sci Transl Med. 2012;4(161):161.7. Ryvlin P, Cross JH, Rheims S. Epilepsy surgery in children andadults. Lancet Neurol. 2014;13(11):1114-1126.Commentary 329

UC IrvineUC Irvine Previously Published WorksTitleEpilepsy Therapy Goes Viral.Permalinkhttps://escholarship.org/uc/item/8sd8770jJournalEpilepsy currents, 19(5)ISSN1535-7597AuthorsLim, Byung ChanHunt, Robert FPublication Date2019-09-01DOI10.1177/1535759719870506Copyright InformationThis work is made available under the terms of a Creative Commons Attribution License, availalbe at https://creativecommons.org/licenses/by/4.0/ Peer reviewedeScholarship.org Powered by the California Digital LibraryUniversity of CaliforniaCurrent Literaturein Basic ScienceEpilepsy Therapy Goes ViralEpilepsy Gene Therapy Using an Engineered Potassium ChannelSnowball A, Chabrol E, Wykes RC, Shekh-Ahmad T, Cornford JH, Lieb A, Hughes MP, Massaro G, Rahim AA, Hashemi KS,Kullmann DM, Walker MC, Schorge S. J Neurosci. 2019;39(16):3159-3169.Refractory focal epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy representsa promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must becarefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vectoroptimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon opti-mized for human expression and mutated to accelerate the recovery of the channels from inactivation. For improved safety,this engineered potassium channel (EKC) gene was packaged into a nonintegrating lentiviral vector under the control of a celltype-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled preclinical trial, the EKC lentivector robustlyreduced seizure frequency in a male rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures.When packaged into an adeno-associated viral vector (AAV2/9), the EKC gene was also effective at suppressing seizures in amale rat model of temporal lobe epilepsy. This demonstration of efficacy in a clinically relevant setting, combined with theimproved safety conferred by cell type–specific expression and integration-deficient delivery, identifies EKC gene therapy asbeing ready for clinical translation in the treatment of refractory focal epilepsy.Keywordsepilepsy, virus, gene therapy, KCNA1, neocortical focal epilepsyCommentaryGene therapy has been envisioned since the 1970s but has faceda number of difficult obstacles, both in the laboratory and theclinic. Following early success of a clinical trial for adenosinedeaminase - severe combined immunodeficiency (ADA-SCID)in 1990, subsequent clinical failures quickly led to dismissal ofgene therapy as nothing more than an experimental technique.However, over the last 15 years, there have been dramaticimprovements in human genetics, genome editing strategies,methods for gene delivery, and disease modeling, all of whichhas made this innovative therapeutic strategy move at anincreasingly rapid pace toward every day clinical practice.1This has led to a renewed interest in the promise of gene ther-apy for a wide range of human diseases, highlighted by a recentFood and Drug Administration approval for the treatment ofspinal muscular atrophy.2 However, despite many convincingadvances in the field, antiepileptic gene therapy remains at apreclinical proof-of-principle stage. There are a number ofimportant challenges that must be overcome in order to movethis exciting technology to the clinic for epilepsy, such aschoice of gene or pathway to target, viral packaging,off-target effects, successful delivery into the human brain,safety, and even the inherent heterogeneous etiology of epilepsyitself. With potentially more than 100 genes reported to causeepilepsy,3 many of which are expressed throughout the brainand/or body, it is difficult to envision applying the traditionalconcept of gene therapy (ie, to fix a defective gene or pathway)for each individual monogenetic disorder. In contrast,approaches that target more downstream pathways to manipulatethe expression of genes that control neuronal or circuit excitabil-ity in affected brain areas may have broader application.4Snowball and colleagues built upon their prior work5,6developing a viral vector–mediated gene therapy to alter neu-ronal and circuit excitability in focal neocortical epilepsy. Thisapproach involves overexpression of the human KCNA1 gene,which encodes the voltage-gated potassium channel Kv1.1, inepileptogenic foci. Here, the authors engineered and optimizedthe viral construct used in earlier studies with the goal of gen-erating an effective gene therapy that might be ready for clin-ical translation. To do this, they first replicated and extendedEpilepsy Currents2019, Vol. 19(5) 328-329ª The Author(s) 2019Article reuse guidelines:sagepub.com/journals-permissionsDOI: 10.1177/1535759719870506journals.sagepub.com/home/epiCreative Commons Non Commercial No Derivs CC BY-NC-ND: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 License (http://www.creativecommons.org/licenses/by-nc-nd/4.0/) which permits non-commercial use, reproduction anddistribution of the work as published without adaptation or alteration, without further permission provided the original work is attributed as specified on theSAGE and Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).EPILEPSY CURRENTStheir prior work by demonstrating a dramatic suppression ofspontaneous seizures following the delivery of a CMV-drivenKCNA1 lentivirus directly into the seizure focus in visual cor-tex of a tetanus toxin model. This construct was then modifiedin 3 main ways: (1) the non-cell-type-specific CMV promotorwas exchanged for a CAMK2A promotor to drive Kv1.1 expres-sion in excitatory projection neurons, (2) the Kv1.1 gene wasmodified to reduce its inactivation, and (3) the resulting plas-mid was packaged into a nonintegrating lentiviral vector toreduce the potential risk of mutagenesis. This new, refinedconstruct had even more powerful therapeutic effects on sei-zures in the tetanus toxin model of focal epilepsy, presumablydue to increased specificity for targeting excitatory neurons,and it was also effective in a kainic acid model of temporallobe epilepsy, when packaged into an AAV vector and injectedbilaterally into hippocampus.Neocortical epilepsy is often drug-resistant, and surgery toremove the epileptogenic zone is not always feasible or effec-tive. As such, development of a spatially restricted, cell-type-specific gene therapy that selectively modifies the activity ofneurons at the seizure focus could be an important advance. Ifsuccessful, this strategy might also spare unaffected brainregions or minimize the types of side effects seen in currentpharmacological approaches. Although the results reported bySnowball and colleagues are promising, additional hurdlesremain before eventual translation to the clinic. For example,the authors only evaluated the effect of this gene therapy onseizures, and it is not yet known how overexpression of Kv1.1in CAMK2A neurons might alter visual cortex function. Inthinking about moving from rodent to human, large animalstudies may be necessary for determining how to preciselydeliver the vector to epileptic foci of a gyrated neocortex.Ultimately, for clinical trials to succeed, one must decidewho will be an appropriate candidate for therapy. A patientwith drug-resistant epilepsy and a well-defined focal lesion ineloquent cortex has been considered to be an ideal candidate.The most prevalent pathologic substrate of epilepsy surgery isfocal cortical dysplasia, which has been identified in both mag-netic resonance imaging positive and negative cases.7However, if a gene therapy is effective and safe, and can bebroadly applied to many forms of the disease as indicated in thecurrent study, it may be advantageous to develop this technol-ogy for a wider range of patients regardless of whether or not afocal epileptogenic zone is present. It is possible that such atherapy could be useful in acquired epilepsy, such as followinga traumatic brain injury or stroke, as well as in genetic epilep-sies. As such, this study by Snowball and colleagues providesan important step closer to achieving successful gene therapyfor epilepsy.By Byung Chan Lim and Robert F. HuntORCID iDRobert F. Hunt https://orcid.org/0000-0003-4490-8718References1. Dunbar CE, High KA, Joung JK, Kohn DB, Ozawa K, Sadelain M.Gene therapy comes of age. Science. 2018;359(6372):eaan4672.2. Mendell JR, Al-Zaidy S, Shell R, et al. Single-dose gene-replacement therapy for spinal muscular atrophy. N Engl J Med.2017;377(18):1713-1722.3. McTague A, Howell KB, Cross JH, Kurian MA, Scheffer IE. Thegenetic landscape of the epileptic encephalopathies of infancy andchildhood. Lancet Neurol. 2016;15(3):304-316.4. Kullmann DM, Schorge S, Walker MC, Wykes RC. Gene therapyin epilepsy-is it time for clinical trials? Nat Rev Neurol. 2014;10(5):300-304.5. Heeroma JH, Henneberger C, Rajakulendran S, Hanna MG,Schorge S, Kullmann DM. Episodic ataxia type 1 mutations differ-entially affect neuronal excitability and transmitter release. DisModel Mech. 2009;2(11-12):612-619.6. Wykes RC, Heeroma JH, Mantoan L, et al. Optogenetic and potas-sium channel gene therapy in a rodent model of focal neocorticalepilepsy. Sci Transl Med. 2012;4(161):161.7. Ryvlin P, Cross JH, Rheims S. Epilepsy surgery in children andadults. Lancet Neurol. 2014;13(11):1114-1126.Commentary 329

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Epilepsy Therapy Goes Viral.

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