CaMKII inhibition rectifies arrhythmic phenotype in a patient-specific model of catecholaminergic polymorphic ventricular tachycardia

Induced pluripotent stem cells (iPSC) offer a unique opportunity for developmental studies, disease modeling and regenerative medicine approaches in humans. The aim of our study was to create an in vitro ‘patient-specific cell-based system’ that could facilitate the screening of new therapeutic molecules for the treatment of catecholaminergic polymorphic ventricular tachycardia (CPVT), an inherited form of fatal arrhythmia. Here, we report the development of a cardiac model of CPVT through the generation of iPSC from a CPVT patient carrying a heterozygous mutation in the cardiac ryanodine receptor gene (RyR2) and their subsequent differentiation into cardiomyocytes (CMs). Whole-cell patch-clamp and intracellular electrical recordings of spontaneously beating cells revealed the presence of delayed afterdepolarizations (DADs) in CPVT-CMs, both in resting conditions and after β-adrenergic stimulation, resembling the cardiac phenotype of the patients. Furthermore, treatment with KN-93 (2-[N-(2-hydroxyethyl)]-N-(4methoxybenzenesulfonyl)]amino-N-(4-chlorocinnamyl)-N-methylbenzylamine), an antiarrhythmic drug that inhibits Ca2+/calmodulin-dependent serine–threonine protein kinase II (CaMKII), drastically reduced the presence of DADs in CVPT-CMs, rescuing the arrhythmic phenotype induced by catecholaminergic stress. In addition, intracellular calcium transient measurements on 3D beating clusters by fast resolution optical mapping showed that CPVT clusters developed multiple calcium transients, whereas in the wild-type clusters, only single initiations were detected. Such instability is aggravated in the presence of isoproterenol and is attenuated by KN-93. As seen in our RyR2 knock-in CPVT mice, the antiarrhythmic effect of KN-93 is confirmed in these human iPSC-derived cardiac cells, supporting the role of this in vitro system for drug screening and optimization of clinical treatment strategies.

[1]  Laura Iop,et al.  Dantrolene rescues arrhythmogenic RYR2 defect in a patient-specific stem cell model of catecholaminergic polymorphic ventricular tachycardia , 2012, EMBO molecular medicine.

[2]  S. Priori,et al.  Short Communication: Flecainide Exerts an Antiarrhythmic Effect in a Mouse Model of Catecholaminergic Polymorphic Ventricular Tachycardia by Increasing the Threshold for Triggered Activity , 2011, Circulation research.

[3]  S. Priori,et al.  Flecainide and antiarrhythmic effects in a mouse model of catecholaminergic polymorphic ventricular tachycardia. , 2012, Trends in cardiovascular medicine.

[4]  M. Latronico,et al.  A lentiviral vector with a short troponin-I promoter for tracking cardiomyocyte differentiation of human embryonic stem cells , 2008, Gene Therapy.

[5]  D. Roden,et al.  Flecainide prevents catecholaminergic polymorphic ventricular tachycardia in mice and humans , 2009, Nature Medicine.

[6]  Katriina Aalto-Setälä,et al.  Cell Model of Catecholaminergic Polymorphic Ventricular Tachycardia Reveals Early and Delayed Afterdepolarizations , 2012, PloS one.

[7]  M. Yano,et al.  Dantrolene, a therapeutic agent for malignant hyperthermia, markedly improves the function of failing cardiomyocytes by stabilizing interdomain interactions within the ryanodine receptor. , 2009, Journal of the American College of Cardiology.

[8]  Wenjun Guo,et al.  Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds , 2008, Nature Biotechnology.

[9]  G. Condorelli,et al.  Generation of human cardiomyocytes: a differentiation protocol from feeder-free human induced pluripotent stem cells. , 2013, Journal of visualized experiments : JoVE.

[10]  S. Priori,et al.  Inherited calcium channelopathies in the pathophysiology of arrhythmias , 2012, Nature Reviews Cardiology.

[11]  Silvia G Priori,et al.  Inherited dysfunction of sarcoplasmic reticulum Ca2+ handling and arrhythmogenesis. , 2011, Circulation research.

[12]  Carlo Napolitano,et al.  Induced pluripotent stem cell-derived cardiomyocytes in studies of inherited arrhythmias. , 2013, The Journal of clinical investigation.

[13]  Lior Gepstein,et al.  Modelling the long QT syndrome with induced pluripotent stem cells , 2011, Nature.

[14]  Euan A. Ashley,et al.  Patient-Specific Induced Pluripotent Stem Cells as a Model for Familial Dilated Cardiomyopathy , 2012, Science Translational Medicine.

[15]  Wei-Zhong Zhu,et al.  Neuregulin/ErbB Signaling Regulates Cardiac Subtype Specification in Differentiating Human Embryonic Stem Cells , 2010, Circulation research.

[16]  D. Melton,et al.  Derivation of Human Embryonic Stem Cells , 2007 .

[17]  Dmitry Terentyev,et al.  miR-1 Overexpression Enhances Ca2+ Release and Promotes Cardiac Arrhythmogenesis by Targeting PP2A Regulatory Subunit B56α and Causing CaMKII-Dependent Hyperphosphorylation of RyR2 , 2009, Circulation research.

[18]  Jonathan A. Bernstein,et al.  Using iPS cells to investigate cardiac phenotypes in patients with Timothy Syndrome , 2011, Nature.

[19]  Karl-Ludwig Laugwitz,et al.  Patient-specific induced pluripotent stem-cell models for long-QT syndrome. , 2010, New England Journal of Medicine.

[20]  Lei Yang,et al.  Patient-specific induced pluripotent stem cell derived models of LEOPARD syndrome , 2010, Nature.

[21]  S. Marx,et al.  Dysfunctional ryanodine receptors in the heart: new insights into complex cardiovascular diseases. , 2013, Journal of molecular and cellular cardiology.

[22]  S. Priori,et al.  Mutations in the Cardiac Ryanodine Receptor Gene (hRyR2) Underlie Catecholaminergic Polymorphic Ventricular Tachycardia , 2001, Circulation.

[23]  Azra Fatima,et al.  In vitro Modeling of Ryanodine Receptor 2 Dysfunction Using Human Induced Pluripotent Stem Cells , 2011, Cellular Physiology and Biochemistry.

[24]  G. Horgan,et al.  Relative expression software tool (REST©) for group-wise comparison and statistical analysis of relative expression results in real-time PCR , 2002 .

[25]  Y. Korchev,et al.  Functional interaction between charged nanoparticles and cardiac tissue: a new paradigm for cardiac arrhythmia? , 2013, Nanomedicine.

[26]  Johannes E. Schindelin,et al.  Fiji: an open-source platform for biological-image analysis , 2012, Nature Methods.

[27]  S. Priori,et al.  A Clinical Approach to Inherited Arrhythmias , 2012, Circulation. Cardiovascular genetics.

[28]  S. Priori,et al.  Calmodulin kinase II inhibition prevents arrhythmias in RyR2(R4496C+/-) mice with catecholaminergic polymorphic ventricular tachycardia. , 2011, Journal of molecular and cellular cardiology.

[29]  Bruce D Gelb,et al.  Induced pluripotent stem cell-derived cardiomyocytes as models for genetic cardiovascular disorders , 2011, Current opinion in cardiology.

[30]  José Jalife,et al.  Optical Imaging of Voltage and Calcium in Cardiac Cells & Tissues , 2012, Circulation research.

[31]  E. Zaklyazminskaya,et al.  Cardiac channelopathies: genetic and molecular mechanisms. , 2013, Gene.

[32]  A. J. Williams,et al.  Flecainide reduces Ca2+ spark and wave frequency via inhibition of the sarcolemmal sodium current , 2013, Cardiovascular research.

[33]  Michael Glikson,et al.  Modeling of catecholaminergic polymorphic ventricular tachycardia with patient-specific human-induced pluripotent stem cells. , 2012, Journal of the American College of Cardiology.

[34]  Erik Willems,et al.  Induced Pluripotent Stem Cells in Cardiovascular Drug Discovery , 2013, Circulation research.

[35]  Carlo Napolitano,et al.  Clinical and Molecular Characterization of Patients With Catecholaminergic Polymorphic Ventricular Tachycardia , 2002, Circulation.

[36]  T. Ichisaka,et al.  Induction of Pluripotent Stem Cells From Adult Human Fibroblasts by Defined Factors , 2008 .

[37]  T. Ichisaka,et al.  Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors , 2007, Cell.

[38]  George Q. Daley,et al.  Disease-Specific Induced Pluripotent Stem Cells , 2008, Cell.

[39]  Donald M Bers,et al.  Local &bgr;-Adrenergic Stimulation Overcomes Source-Sink Mismatch to Generate Focal Arrhythmia , 2012, Circulation research.

[40]  Ofer Binah,et al.  Cardiomyocytes generated from CPVTD307H patients are arrhythmogenic in response to β-adrenergic stimulation , 2012, Journal of cellular and molecular medicine.

[41]  M. Eldar,et al.  A missense mutation in CASQ2 is associated with autosomal recessive catecholamine‐induced polymorphic ventricular tachycardia in Bedouin families from Israel , 2004, Annals of medicine.

[42]  L. Blatter,et al.  Dantrolene prevents arrhythmogenic Ca2+ release in heart failure. , 2012, American journal of physiology. Heart and circulatory physiology.

[43]  D. Bers,et al.  Ca2+/Calmodulin–Dependent Protein Kinase Modulates Cardiac Ryanodine Receptor Phosphorylation and Sarcoplasmic Reticulum Ca2+ Leak in Heart Failure , 2005, Circulation research.

[44]  Shulan Tian,et al.  Induced Pluripotent Stem Cell Lines Derived from Human Somatic Cells , 2007, Science.