RANGRF

Sudden cardiac death is a sudden, unexpected death developed by one of the many different causes of cardiac arrest that occur within 1 hour of the onset of new symptoms. Sudden unexplained death (SUD) comprises a normal heart at postmortem examination and negative toxicological analysis. SUD often arises from cardiac genetic disease, particularly channelopathies. Channelopathies, or inherited arrhythmia syndromes, are a group of disorders characterized by an increased risk of sudden cardiac death, abnormal cardiac electrical function, and, typically, a structurally normal heart. They share an underlying genetic etiology where disease-causing genetic variants may lead to the absence or dysfunction of proteins involved in the generation and propagation of the cardiac action potential. Our study aimed to evaluate the importance of next-generation sequencing in the postmortem investigations of SUD cases. In this study, 5 forensic SUD cases were investigated for inherited cardiac disorders. We screened a total of 68 cardiac genes for the sibling of case 1, as well as case 2, and 51 genes for cases 3, 4, and 5. Of the 12 variants identified, 2 likely pathogenic variants (16.7%) were the TMEM43_ c.1000+2T>C splice site mutation and the SCN5A_ p.W703X nonsense mutation. The remaining 10 variants of uncertain significance were detected in the TRPM4, RANGRF, AKAP9, KCND3, KCNE1, DSG2, CASQ1, and SNTA1 genes. Irrespective of genetic testing, all SUD families require detailed clinical testing to identify relatives who may be at risk. Molecular autopsy and detailed premorbid clinical and family histories can survive family members of SUD cases.

The RAN Guanine Nucleotide Release Factor (RANGRF) gene encodes the protein MOG1, which binds to Nav1.5 and facilitates its transport to the cell membrane. Nav1.5 mutations have been linked to various cardiac arrhythmias and cardiomyopathy. To investigate the role of RANGRF in this process, we utilized the CRISPR/Cas9 gene editing system to generate a homozygous RANGRF knockout hiPSC line. The availability of the cell line will prove to be an invaluable asset in the study of disease mechanisms and the testing of gene therapies for cardiomyopathy.

Brugada syndrome (BrS) is a fatal arrhythmia that causes an estimated 4% of all sudden death in high-incidence areas. SCN5A encodes cardiac sodium channel Na_V1.5 and causes 25 to 30% of BrS cases. Here, we report generation of a knock-in (KI) mouse model of BrS (Scn5a^G1746R/+). Heterozygous KI mice recapitulated some of the clinical features of BrS, including an ST segment abnormality (a prominent J wave) on electrocardiograms and development of spontaneous ventricular tachyarrhythmias (VTs), seizures, and sudden death. VTs were caused by shortened cardiac action potential duration and late phase 3 early afterdepolarizations associated with reduced sodium current density (I_Na) and increased Kcnd3 and Cacna1c expression. We developed a gene therapy using adeno-associated virus serotype 9 (AAV9) vector-mediated MOG1 delivery for up-regulation of MOG1, a chaperone that binds to Na_V1.5 and traffics it to the cell surface. MOG1 was chosen for gene therapy because the large size of the SCN5A coding sequence (6048 base pairs) exceeds the packaging capacity of AAV vectors. AAV9-MOG1 gene therapy increased cell surface expression of Na_V1.5 and ventricular I_Na, reversed up-regulation of Kcnd3 and Cacna1c expression, normalized cardiac action potential abnormalities, abolished J waves, and blocked VT in Scn5a^G1746R/+ mice. Gene therapy also rescued the phenotypes of cardiac arrhythmias and contractile dysfunction in heterozygous humanized KI mice with SCN5A mutation p.D1275N. Using a small chaperone protein may have broad implications for targeting disease-causing genes exceeding the size capacity of AAV vectors.