Dravet syndrome (DS), a rare and severe form of epilepsy, typically presents in the first year of life. It is characterized by recurrent seizures that are often difficult to control with traditional antiepileptic drugs (AEDs).
The pathophysiology of DS is largely attributed to mutations in the SCN1A gene, which encodes the α subunit of NaV1.1—a voltage-gated sodium channel. The channel is critical for the proper functioning of inhibitory interneurons in the brain, which play a crucial role in regulating the excitability of neuronal networks.1
The most common genetic mutation in DS is a heterozygous loss-of-function mutation in the SCN1A gene. This mutation leads to a reduction in the number of functional NaV1.1 channels, since they must all be transcribed from the single healthy allele. This is termed haploinsufficiency.
As a result, there is impaired functioning of inhibitory interneurons, which leads to overexcitability of neuronal networks. This imbalance between excitation and inhibition is thought to be a key mechanism underlying the development of seizures in DS.1
An Urgent Need for New Treatment Options
Recent advancements in gene therapy have shown promise in treating this devastating condition. Specifically, gene therapy approaches that target the SCN1A gene are being developed to correct the genetic defect underlying DS. These therapies aim to either replace the faulty gene with a healthy copy or repair the mutation in it. The goal is to correct the defect and restore the gene’s normal neuronal functioning.
Read more about the genetics of DS
Although the development of these therapies is still in the early stages, they offer a potential new avenue for treating DS and other genetic forms of epilepsy. Given the high morbidity associated with DS, the need for effective and specific gene therapies is urgent.
Preclinical trials for DS are usually conducted in an inducible DS mouse model that harbors an A1783V mutation in the SCN1A gene. Through the Epilepsy Therapy Screening Program and sponsored by the National Institute of Neurological Disorders and Stroke, these DS mouse models are available free of cost for researchers investigating novel therapies for DS.2
The advent of novel gene editing tools, such as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) technology, and advancements in the development of adeno-associated virus (AAV)-based gene delivery systems, have paved the way for more precise and efficient gene therapy strategies for DS. Gene therapy has the potential to provide long-lasting benefits for patients with DS by targeting the root cause of the disorder and could offer a cure for this disease.
AAV-Based Gene Delivery With ETX101
The employment of recombinant, nonreplicating AAV-based vectors for gene replacement therapy has brought about revolutionary clinical advantages in the management of severe monogenic central nervous system (CNS) pathologies. One challenge in using this vector in DS is the size of the SCN1A gene, which exceeds the packing capacity of AAV.
To combat this problem, a group of researchers has developed AAV9-REGABA-eTFSCN1A (ETX101), an experimental AAV vector that has been formulated to regulate SCN1A gene expression via an engineered transcription factor (eTFSCN1A) that elicits endogenous SCN1A upregulation. Furthermore, the incorporation of a cell-selective regulatory element (REGABA) enables transgene expression targeting with high specificity towards GABAergic inhibitory interneurons.3
Read more about the pathophysiology of DS
Following injection with ETX101, SCN1A transcript levels in GABAergic inhibitory interneurons significantly increased, compared to untreated and excitatory cells in Scn1a+/- (haploinsufficient) mice. This finding demonstrates that ETX101 is cell-specific to the GABAergic inhibitory interneurons being targeted.
Additionally, the therapy resulted in an approximately 30% increase in membrane-associated NaV1.1 protein expression in the CNS tissue compared to vehicle-treated mice. These data demonstrate that ETX101 therapy can selectively upregulate the SCN1A gene in GABAergic inhibitory interneurons, suggesting its potential as a treatment for disorders associated with SCN1A loss of function mutations like DS.3
These findings correlated well in reducing the number and severity of seizures in the DS mouse models. DS mice in the treatment group had a 68% reduction in the average number of daily seizures. Additionally, 67% of mice were seizure-free following treatment during the study period vs 20% in the vehicle treatment group.3
Treatment with ETX101 also reduced temperature-induced seizures, a very common seizure trigger in DS.3,4 Approximately 87% of DS mice experienced seizures at an internal body temperature of 43.0 °C meanwhile 88% of treated mice did not experience seizures at this body temperature.3
The research team is planning to conduct an in-human trial of ETX101 to study safety and effectiveness during the first few years of treatment in a child’s life.5
RNA Modulation With STK001
In early 2022, researchers funded by Stoke Therapeutics were able to increase the SCN1A mRNA levels in DS mouse models using an antisense oligonucleotide named STK001 by employing a special technique called targeted augmentation of nuclear gene output (TANGO). This technique is not considered gene therapy, as it does not involve the manipulation or insertion of genes to the host’s DNA. Rather, it is considered RNA modulation. In addition to achieving increased SCN1A mRNA levels, STK001 was also able to increase NaV1.1 protein expression, reduce seizures, and improve survival in the mouse model of DS.6
These compelling results have led to the approval of 2 ongoing, parallel, phase 1/2a clinical trials in humans in the US and the UK: MONARCH and ADMIRAL.
The primary outcome of the MONARCH trial is the evaluation of the safety and tolerability of single and multiple doses of STK-001 in patients in the US aged 2 to 18 years. Secondary outcomes include measurement of seizure frequency and quality of life, among others.7 An open-label, multicenter, extension study is inviting and enrolling patients who have already received STK-001 to evaluate the safety of multiple doses of the experimental drug.8
Read about other clinical trials in DS
The ADMIRAL trial is taking place in the UK and is evaluating the safety and tolerability of multiple, ascending doses of STK-001 in people aged 2 to 18 years with DS.9
Interim analysis of both trials was made available in 2022 and highlighted some positive findings. The published data indicate that, to date, single and multiple doses of up to 30 mg of STK-001 are well tolerated with no safety concerns related to the drug. In addition, STK-001 levels were detected in cerebrospinal fluid up to 6 months after a single dose.10 The most recent press release by Stoke Therapeutics states that there is “a trend toward a reduction in median percent change from baseline in convulsive seizure frequency among patients treated with single doses of STK-001,” but did not disclose numerical figures.11
Tethered mRNA Amplifier and Engineered Transfer RNAs
In the summer of 2022, researchers published data on a novel technique that increases SCN1A expression by enhancing mRNA translation in haploinsufficient DS mice. Haploinsufficiency is a genetic condition in which one copy of a gene is insufficient to produce enough protein to avoid disease. To address this issue, scientists have been looking into ways of targeting endogenous transcripts, which are involved in post-transcriptional regulation and increasing protein output.12
This was achieved by developing a new technology called the tethered mRNA amplifier, which enhances the expression of specific mRNAs. They found that this technology can control both reporter and endogenous transcripts in human cells and has the potential to stimulate the translation of transcripts linked to haploinsufficiency disorders. The researchers optimized the mRNA amplifier so that it can be integrated into current AAV gene therapy strategies. They propose this approach could be an effective way to treat human haploinsufficiencies. Using this strategy, the researchers were successful in augmenting the quantity of a two-test protein by 1.5 to 2 fold.12
This technology is still in the preclinical stage of development but the hope is that this method could one day be employed to treat a wide array of haploinsufficiencies, including DS.
Read about experimental therapies in DS
Interestingly, a separate group of researchers has developed a similar technique that allows for the translation of mRNA with premature stop codons resulting from genetic frameshift mutations. Essentially, this allows for the translation of proteins that would otherwise be truncated due to premature termination codons.13 This lends hope to patients with genetic disease due to homozygous mutations. While homozygous SCN1A mutations are a very rare cause of DS, it has been described in case reports.14
Dravet and Beyond
Personalized medicine is considered by many experts to be the future of health care. More than 80% of DS cases are caused by mutations in the SCN1A gene, usually due to missense mutations resulting in haploinsufficiency. Gene therapy with AAV-based delivery systems is attempting to incorporate a properly sequenced gene into the patient’s DNA and offer a true cure. This area of research has become fast-paced and has the potential to treat hundreds of previously incurable diseases. Likewise, techniques that employ RNA modification show promise not only for DS, but also for the hundreds of human haploinsufficiencies that exist.
1. Chilcott E, Díaz JA, Bertram C, Berti M, Karda R. Genetic therapeutic advancements for Dravet syndrome. Epilepsy Behav. 2022;132. doi:10.1016/j.yebeh.2022.108741
2. Novel open-access mouse model of Dravet syndrome. Dravet Syndrome European Federation. Accessed April 24, 2023.
3. Tanenhaus A, Stowe T, Young A, et al. Cell-selective adeno-associated virus-mediated SCN1A gene regulation therapy rescues mortality and seizure phenotypes in a Dravet syndrome mouse model and is well tolerated in nonhuman primates. Hum Gene Ther. 2022;33(11-12):579-597. doi:10.1089/hum.2022.037
4. Warner TA, Liu Z, Macdonald RL, Kang J. Heat induced temperature dysregulation and seizures in Dravet Syndrome/GEFS+ Gabrg2+/Q390X mice. PubMed Central. Accessed April 25, 2023.
5. Askham A. Gene therapy targets interneurons to tackle Dravet syndrome. Spectrum. Published June 13, 2022. Accessed April 25, 2023.
6. Wengert ER, Wagley PK, Strohm SM, et al. Targeted augmentation of nuclear gene output (TANGO) of Scn1a rescues parvalbumin interneuron excitability and reduces seizures in a mouse model of Dravet syndrome. Brain Res. 2022;1775:147743. doi:10.1016/j.brainres.2021.147743
7. An open-label study to investigate the safety and pharmacokinetics of single and multiple ascending doses of antisense oligonucleotide STK-001 in children and adolescents with Dravet syndrome. ClinicalTrials.gov. June 22, 2020. Updated March 28, 2023. Accessed April 24, 2023.
8. An open-label extension study for patients with Dravet syndrome who previously participated in studies of STK-001. ClinicalTrials.gov. February 5, 2021. Updated February 10, 2023. Accessed April 24, 2023.
9. Admiral: a study of the safety of multiple increasing doses of STK-001 in children and adolescents with Dravet syndrome. ISRCTN registry. Accessed April 24, 2023.
10. Laux L. Interim analyses: an open-label study to investigate the safety and pharmacokinetics of single and multiple ascending doses of antisense oligonucleotide STK- 001 in children and adolescents with Dravet syndrome. Stoke Therapeutics. Accessed April 24, 2023.
11. Stoke therapeutics announces positive interim safety, PK and CSF exposure data from the phase 1/2a MONARCH study of STK-001 in children and adolescents with Dravet syndrome. News release. Stoke Therapeutics; September 21, 2021.
12. Torkzaban B, Kawalerski R, Coller J. Development of a tethered mRNA amplifier to increase protein expression. Biotechnol J. 2022;17(10):2200214. doi:10.1002/biot.202200214
13. Lueck JD, Yoon JS, Perales-Puchalt A, et al. Engineered transfer RNAs for suppression of premature termination codons. Nat Commun. 2019;10(1):822. doi:10.1038/s41467-019-08329-4
14. Brunklaus A, Ellis R, Stewart H, et al. Homozygous mutations in the SCN1A gene associated with genetic epilepsy with febrile seizures plus and Dravet syndrome in 2 families. Eur J Paediatr Neurol. 2015;19(4):484-488. doi:10.1016/j.ejpn.2015.02.001