Development and Validation of a Transgenic Rabbit Model of Dravet Syndrome

Dravet 综合征转基因兔模型的开发和验证

• 批准号: 10574719
• 负责人: Lori L. Isom
• 金额: $ 37.21万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10574719
• 关键词: Action Potentials; Age; Animal Genetics; Animal Model; Animals; Antisense Oligonucleotides; Apnea; Arrhythmia; Autonomic Dysfunction; Behavioral; Biological Markers; Brain; CRISPR/Cas technology; Calcium; Cardiac; Cardiac Myocytes; Cardiovascular system; Cause of Death; Cells; Cessation of life; Clinical; Complex; Data; Development; Dissection; Drug Combinations; Epilepsy; Frequencies; Future; Generalized seizures; Genes; Genetic; Genetic Models; Goals; Heart; Human; Incidence; Knowledge; Link; Michigan; Modeling; Mus; Neurons; New Zealand; Organism; Oryctolagus cuniculus; Patients; Pharmaceutical Preparations; Phase; Phenotype; Physiology; Rattus; Risk; Rodent; Role; SCN1A protein; SCN8A gene; Seizures; Sodium; Sodium Channel; Testing; Tissues; Transgenic Mice; Transgenic Organisms; Translating; Translational Research; Universities; Validation; Variant; Video Recording; Work; clinical diagnosis; clobazam; cohort; density; dravet syndrome; effective therapy; epileptic encephalopathies; gain of function; gene therapy; heart innervation; human disease; human old age (65+); induced pluripotent stem cell; insight; loss of function; mouse model; nerve supply; novel marker; novel therapeutic intervention; premature; prevent; respiratory; risk variant; stem cell model; success; sudden unexpected death in epilepsy; translational model; translational therapeutics; two-dimensional; voltage

ABSTRACT Sudden Unexpected Death in Epilepsy (SUDEP) is a leading cause of death in patients with epilepsy. SUDEP mechanisms are not understood, although there is evidence to implicate apnea, autonomic dysfunction, and cardiac arrhythmias. Our work in mice led to the hypothesis that cardiac arrhythmias contribute to the mechanism of SUDEP in channelopathy-linked genetic epilepsies. We demonstrated altered cardiac myocyte (CM) ionic currents, calcium handling, and action potentials (APs), as well as cardiac arrhythmias in mouse models of SCN1A-, SCN8A-, and SCN1B-linked developmental and epileptic encephalopathy (DEE). We showed that induced pluripotent stem cell (iPSC)-derived CMs from Dravet syndrome (DS) patients have substrates for arrhythmias. Importantly, no animal or iPSC model can completely replicate the human DS phenotype. Because mouse and human cardiac APs are very different, we used human iPSC-CM models to investigate cell autonomous effects of SCN1A haploinsufficiency. However, cells in 2-dimensional culture cannot replicate complex cardiac tissues, cardiovascular changes, or cardiac innervation. Here, we will develop and validate a large animal model in which the role of cardiac arrhythmias, in addition to seizures, in SUDEP can be investigated. Rabbits closely replicate the human cardiac AP and provide a complete organism to translate to the clinical setting. Our objective is to develop and validate a transgenic rabbit model of SCN1A-linked DS. We generated a New Zealand White (NZW) rabbit Scn1a deletion model using CRISPR-Cas9 gene editing. However, NZW Scn1a+/- rabbits showed neither seizures nor cardiac arrhythmia. We later found that F1: NZW x Dutch Belted Scn1a+/- rabbits have seizures, cardiac arrhythmia, and premature death. These exciting results suggest that we may have generated the first transgenic large animal model of a DEE, although this model must now be rigorously validated. If validated, this work will be a significant advance over currently available DS models. R61 Phase Specific Aims: 1. To characterize seizure onset, seizure types, seizure frequency and duration, and determine the rate of SUDEP in DS rabbits. 2. To characterize arrhythmia types, frequency, and duration, whether arrhythmias occur independently of seizures, and whether cardiac arrhythmia is associated with SUDEP in DS rabbits. R33 Phase Specific Aim: To validate the model using the ASO drug STK-001 or the drug combination stiripentol + clobazam to reduce seizures and SUDEP. Establishing a genetic rabbit model of DS will better inform the translatability of neuro-cardiac mechanisms of SUDEP to human disease.

摘要