Gene Correction Approaches for Treating Long QT Syndrome in Children

PI’s: Kevin Maher, Gang Bao, TJ Cradick, Chunhui Xu & Mary Wagner

Long QT syndrome is an inherited disorder of the ion channels of the cardiomyocytes. It occurs in approximately in 1:2,000 children and is associated with ventricular tachycardia, fibrillation and sudden cardiac death. The genetic defects are typically missense mutations that produce abnormal ion channels in the cell membrane. Palliative treatment in these children involves medications and intracardiac defibrillators. There is no cure for this disease that carries a  lifelong risk of sudden death. Advances in biotechnology have allowed the development of induced pluripotent stem (iPS) cells and cardiomyocytes from these patients. The proposed research for this project will involve correcting of the genetic defect in these cells, with removal of the long QT phenotype from the cardiomyocytes, curing the disease at the cellular level

Aim 1. Develop iPS cell lines from children with long QT syndrome: Fibroblasts from skin biopsies obtained from children with long QT syndrome will be reprogrammed into iPS cells which will be then sub-cultured and stimulated to become cardiomyocytes. These cell lines will undergo genetic and physiologic testing to confirm the disease genotype and phenotype, as seen in the patients.

Aim 2. Genetically correct the DNA defect in the long QT cardiomyocytes: TAL Effector Nucleases (TALEN) technology will be employed to correct the mutation in genomic DNA. TALENs will be used to specifically cleave the DNA near the mutation site. The gene will be edited using a supplied donor DNA and cellular homologous recombination. These “corrected” cell lines will then undergo physiologic testing to demonstrate that the pathologic genotype and phenotype has been removed, resulting in normally functioning cardiomyocytes.

Publications Preininger MK, Jha R, Maxwell JT, Wu Q, Singh M, Wang B, Dalal A, Mceachin ZT, Rossoll W, Hales CM, Fischbach P, Wagner MB, Xu C. A stem cell model of catecholaminergic polymorphic ventricular tachycardia recapitulates patient-specific drug responses. Dis Model Mech 9:927-939 (2016). PMID: 27491078; PMCID: PMC5047684. Jha R, Wu Q, Singh M, Preininger MK, Han P, Ding G, Cho H, Jo H, Maher KO, Wagner MB, Xu C. Simulated microgravity and 3D culture enhance induction, viability, proliferation and differentiation of cardiac progenitors from human pluripotent stem cells. Sci Rep 6:30956 (2016). PMID: 27492371; PMCID: PMC4974658. Han J, Qian X, Wu Q, Jha R, Duan J, Yang Z, Maher KO, Nie S, Xu C. Novel surface-enhanced Raman scattering-based assays for ultra-sensitive detection of human pluripotent stem cells. Biomaterials 105:66-76 (2016). PMID: 27509304. Han J, Wu Q, Xia Y, Wagner MB and Xu C. Cell alignment induced by anisotropic electrospun fibrous alone has limited effect on cardiomyocyte maturation. Stem Cell Res 16, 740-750 (2016). PMID: 27131761; PMCID: PMC4903921. Rajneesh Jha, Monalisa Singh, Qingling Wu, Cinsley Gentillon, Marcela K. Preininger, Chunhui Xu, Downregulation of LGR5 Expression Inhibits Cardiomyocyte Differentiation and Potentiates Endothelial Differentiation from Human Pluripotent Stem Cells, Stem Cell Reports, Volume 9, Issue 2, 2017,Pages 513-527, ISSN 2213-6711, Jha R, Xu RH, Xu C. Efficient differentiation of cardiomyocytes from human pluripotent stem cells with growth factors. In Methods Mol Biol. Skuse GR, Ferran MC eds. Humana Press.1299:115-31 (2015) Preininger MK, Singh M, Xu C. Cryopreservation of human stem cell-derived cardiomyocytes: strategies, challenges, and future directions. Karimi-Busheri F and Weinfeld M eds. In Biobanking and Cryopreservation of Stem Cells. Springer Press. Adv Exp Med Biol. 951:123-135 (2016)

Related Research