Pharmacological TRPC6 inhibition improves survival and muscle function in mice with Duchenne muscular dystrophy

Gene mutations causing loss of dystrophin result in the severe muscle disease known as Duchenne muscular dystrophy (DMD). Despite efforts at genetic repair, DMD therapy remains largely palliative. Loss of dystrophin destabilizes the sarcolemmal membrane impacting mechanosensitive cation channels to increase calcium entry, promoting cell damage, and eventually muscle dysfunction. One putative channel is transient receptor potential canonical 6 (TRPC6) that we showed contributes to abnormal force and calcium stress-responses in mouse cardiomyocytes lacking dystrophin and haplodeficient in utrophin mdx/utrn+/- (HET). Here, we show in both HET and the far more severe homozygous mdx/utrn-/- (DKO) mouse that TRPC6 gene deletion or its selective pharmacologic inhibition (BI 749327) prolongs survival 2-3-fold, improving skeletal and cardiac muscle and bone defects. Gene pathways reduced by BI 749327 treatment most prominently regulate fat metabolism and TGFβ1 signaling. These results support the testing of TRPC6 inhibitors in human trials for other diseases as a novel DMD therapy.

[1]  H. Wilton-Clark,et al.  Casimersen for Duchenne muscular dystrophy. , 2021, Drugs of today.

[2]  T. Yokota,et al.  Pharmacology and toxicology of eteplirsen and SRP-5051 for DMD exon 51 skipping: an update , 2021, Archives of Toxicology.

[3]  E. Mercuri,et al.  Long-Term Safety and Efficacy Data of Golodirsen in Ambulatory Patients with Duchenne Muscular Dystrophy Amenable to Exon 53 Skipping: A First-in-human, Multicenter, Two-Part, Open-Label, Phase 1/2 Trial , 2021, Nucleic acid therapeutics.

[4]  M. Guglieri,et al.  Life Expectancy in Duchenne Muscular Dystrophy , 2021, Neurology.

[5]  G. Trifirò,et al.  Global epidemiology of Duchenne muscular dystrophy: an updated systematic review and meta-analysis , 2020, Orphanet Journal of Rare Diseases.

[6]  E. Olson,et al.  Correction of muscular dystrophies by CRISPR gene editing. , 2020, The Journal of clinical investigation.

[7]  M. Bouxsein,et al.  Guidelines for the assessment of bone density and microarchitecture in vivo using high-resolution peripheral quantitative computed tomography , 2020, Osteoporosis International.

[8]  J. Ervasti,et al.  Social stress is lethal in the mdx model of Duchenne muscular dystrophy , 2020, EBioMedicine.

[9]  Zhenwei Ling,et al.  TRPC6-dependent Ca2+ signaling mediates airway inflammation in response to oxidative stress via ERK pathway , 2020, Cell Death & Disease.

[10]  Hiroshi Ueno,et al.  Effects of repetitive gentle handling of male C57BL/6NCrl mice on comparative behavioural test results , 2020, Scientific Reports.

[11]  Jose A. Adams,et al.  Contribution of TRPC Channels to Intracellular Ca2 + Dyshomeostasis in Smooth Muscle From mdx Mice , 2020, Frontiers in Physiology.

[12]  A. Zarbock,et al.  Intravascular adhesion and recruitment of neutrophils in response to CXCL1 depends on their TRPC6 channels , 2020, Journal of Molecular Medicine.

[13]  R. Spurney,et al.  TRPC Channels in Proteinuric Kidney Diseases , 2019, Cells.

[14]  I. Kalajzic,et al.  CTRP3 Regulates Endochondral Ossification and Bone Remodeling During Fracture Healing , 2019, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[15]  J. Molkentin,et al.  ERK1/2 signaling induces skeletal muscle slow fiber-type switching and reduces muscular dystrophy disease severity , 2019, JCI insight.

[16]  D. Kass,et al.  In vivo selective inhibition of TRPC6 by antagonist BI 749327 ameliorates fibrosis and dysfunction in cardiac and renal disease , 2019, Proceedings of the National Academy of Sciences.

[17]  C. Ward,et al.  GsMTx4-D provides protection to the D2.mdx mouse , 2018, Neuromuscular Disorders.

[18]  D. Duan.  Systemic AAV Micro-dystrophin Gene Therapy for Duchenne Muscular Dystrophy , 2018, Molecular therapy : the journal of the American Society of Gene Therapy.

[19]  D. Largaespada,et al.  Social stress shortens lifespan in mice , 2018, Aging cell.

[20]  P. Germain,et al.  Exploratory rearing: a context- and stress-sensitive behavior recorded in the open-field test , 2018, Stress.

[21]  D. Kass,et al.  Transient receptor potential channel 6 regulates abnormal cardiac S-nitrosylation in Duchenne muscular dystrophy , 2017, Proceedings of the National Academy of Sciences.

[22]  J. McArdle,et al.  Reducing sarcolipin expression mitigates Duchenne muscular dystrophy and associated cardiomyopathy in mice , 2017, Nature Communications.

[23]  M. Drazner,et al.  Predictors of Death in Adults With Duchenne Muscular Dystrophy–Associated Cardiomyopathy , 2017, Journal of the American Heart Association.

[24]  T. Gudermann,et al.  Classical transient receptor potential 6 (TRPC6) channels support myofibroblast differentiation and development of experimental pulmonary fibrosis. , 2017, Biochimica et biophysica acta. Molecular basis of disease.

[25]  R. Ma,et al.  Canonical Transient Receptor Potential 6 Channel: A New Target of Reactive Oxygen Species in Renal Physiology and Pathology. , 2016, Antioxidants & redox signaling.

[26]  David W. Hammers,et al.  Tadalafil Treatment Delays the Onset of Cardiomyopathy in Dystrophin‐Deficient Hearts , 2016, Journal of the American Heart Association.

[27]  Age-related changes in behavior in C57BL/6J mice from young adulthood to middle age , 2016, Molecular Brain.

[28]  Dongsheng Duan,et al.  In vivo genome editing improves muscle function in a mouse model of Duchenne muscular dystrophy , 2016, Science.

[29]  N. HellerKristin,et al.  Human α7 Integrin Gene (ITGA7) Delivered by Adeno-Associated Virus Extends Survival of Severely Affected Dystrophin/Utrophin-Deficient Mice. , 2015 .

[30]  Dong I. Lee,et al.  Prevention of PKG1α oxidation augments cardioprotection in the stressed heart. , 2015, The Journal of clinical investigation.

[31]  D. Kass,et al.  Combined TRPC3 and TRPC6 blockade by selective small-molecule or genetic deletion inhibits pathological cardiac hypertrophy , 2014, Proceedings of the National Academy of Sciences.

[32]  D. Kass,et al.  Hyperactive Adverse Mechanical Stress Responses in Dystrophic Heart Are Coupled to Transient Receptor Potential Canonical 6 and Blocked by cGMP–Protein Kinase G Modulation , 2014, Circulation research.

[33]  T. Gudermann,et al.  TRPC6: physiological function and pathophysiological relevance. , 2014, Handbook of experimental pharmacology.

[34]  T. Benzing,et al.  NOX2 interacts with podocyte TRPC6 channels and contributes to their activation by diacylglycerol: essential role of podocin in formation of this complex. , 2013, American journal of physiology. Cell physiology.

[35]  Y. Mori,et al.  Molecular determinants for cardiovascular TRPC6 channel regulation by Ca2+/calmodulin‐dependent kinase II , 2013, The Journal of physiology.

[36]  J. Schlöndorff,et al.  Gain-of-function Mutations in Transient Receptor Potential C6 (TRPC6) Activate Extracellular Signal-regulated Kinases 1/2 (ERK1/2)* , 2013, The Journal of Biological Chemistry.

[37]  C. Ward,et al.  X-ROS signaling in the heart and skeletal muscle: stretch-dependent local ROS regulates [Ca²⁺]i. , 2013, Journal of molecular and cellular cardiology.

[38]  L. Birnbaumer,et al.  A TRPC6-dependent pathway for myofibroblast transdifferentiation and wound healing in vivo. , 2012, Developmental cell.

[39]  F. Sbrana,et al.  Microtubules Underlie Dysfunction in Duchenne Muscular Dystrophy , 2012, Science Signaling.

[40]  G. V. Ommen,et al.  Comparison of skeletal muscle pathology and motor function of dystrophin and utrophin deficient mouse strains , 2012, Neuromuscular Disorders.

[41]  T. Gudermann,et al.  Activation of TRPC6 channels is essential for lung ischaemia–reperfusion induced oedema in mice , 2012, Nature Communications.

[42]  J. Tidball,et al.  Neuronal Nitric Oxide Synthase-Rescue of Dystrophin/Utrophin Double Knockout Mice does not Require nNOS Localization to the Cell Membrane , 2011, PloS one.

[43]  S. Muallem,et al.  Reactive Oxygen Species-mediated TRPC6 Protein Activation in Vascular Myocytes, a Mechanism for Vasoconstrictor-regulated Vascular Tone* , 2011, The Journal of Biological Chemistry.

[44]  E. Bertini,et al.  Mechanisms Inducing Low Bone Density in Duchenne Muscular Dystrophy in Mice and Humans , 2011, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[45]  Ching-On Wong,et al.  cAMP Activates TRPC6 Channels via the Phosphatidylinositol 3-Kinase (PI3K)-Protein Kinase B (PKB)-Mitogen-activated Protein Kinase Kinase (MEK)-ERK1/2 Signaling Pathway* , 2011, The Journal of Biological Chemistry.

[46]  D. Kass,et al.  Cyclic GMP/PKG-dependent inhibition of TRPC6 channel activity and expression negatively regulates cardiomyocyte NFAT activation Novel mechanism of cardiac stress modulation by PDE5 inhibition. , 2010, Journal of molecular and cellular cardiology.

[47]  D. Allen,et al.  Calcium and the damage pathways in muscular dystrophy. , 2010, Canadian journal of physiology and pharmacology.

[48]  C. Cognard,et al.  Regulation by scaffolding proteins of canonical transient receptor potential channels in striated muscle , 2009, Journal of Muscle Research and Cell Motility.

[49]  B. Aronow,et al.  Calcium influx is sufficient to induce muscular dystrophy through a TRPC-dependent mechanism , 2009, Proceedings of the National Academy of Sciences.

[50]  J. Chamberlain,et al.  Microutrophin delivery through rAAV6 increases lifespan and improves muscle function in dystrophic dystrophin/utrophin-deficient mice. , 2008, Molecular therapy : the journal of the American Society of Gene Therapy.

[51]  H. Brinkmeier,et al.  Transient receptor potential cation channels in normal and dystrophic mdx muscle , 2008, Neuromuscular Disorders.

[52]  D. Allen,et al.  Intracellular calcium handling in ventricular myocytes from mdx mice. , 2007, American journal of physiology. Heart and circulatory physiology.

[53]  John McAnally,et al.  TRPC6 fulfills a calcineurin signaling circuit during pathologic cardiac remodeling. , 2006, The Journal of clinical investigation.

[54]  J. Soboloff,et al.  A common mechanism underlies stretch activation and receptor activation of TRPC6 channels , 2006, Proceedings of the National Academy of Sciences.

[55]  M. Estacion,et al.  Human TRPC6 expressed in HEK 293 cells forms non‐selective cation channels with limited Ca2+ permeability , 2006, The Journal of physiology.

[56]  Robert A. H. White,et al.  Improvement in survival and muscle function in an mdx/utrn −/− double mutant mouse using a human retinal dystrophin transgene , 2006, Neuromuscular Disorders.

[57]  S. Day,et al.  Dystrophic heart failure blocked by membrane sealant poloxamer , 2005, Nature.

[58]  J. Tidball,et al.  Cardiomyopathy in dystrophin-deficient hearts is prevented by expression of a neuronal nitric oxide synthase transgene in the myocardium. , 2005, Human molecular genetics.

[59]  D. Allen,et al.  Effects of stretch‐activated channel blockers on [Ca2+]i and muscle damage in the mdx mouse , 2005, The Journal of physiology.

[60]  H. Debaix,et al.  Involvement of TRPC in the abnormal calcium influx observed in dystrophic (mdx) mouse skeletal muscle fibers , 2002, The Journal of cell biology.

[61]  Susan C. Brown,et al.  Utrophin-Dystrophin-Deficient Mice as a Model for Duchenne Muscular Dystrophy , 1997, Cell.