Exon Skipping and Gene Transfer Restore Dystrophin Expression in Human Induced Pluripotent Stem Cells-cardiomyocytes Harboring DMD Mutations

Overview

Journal Stem Cells Dev

Date 2013 Jul 9

PMID 23829870

Citations 44

Authors

Emily Dick

Spandan Kalra

David Anderson

Vinoj George

Morten Ritso

Steven H Laval

Rita Barresi

Annemieke Aartsma-Rus

Hanns Lochmuller

Chris Denning

Affiliations

Soon will be listed here.

Abstract

With an incidence of ∼1:3,500 to 5,000 in male children, Duchenne muscular dystrophy (DMD) is an X-linked disorder in which progressive muscle degeneration occurs and affected boys usually die in their twenties or thirties. Cardiac involvement occurs in 90% of patients and heart failure accounts for up to 40% of deaths. To enable new therapeutics such as gene therapy and exon skipping to be tested in human cardiomyocytes, we produced human induced pluripotent stem cells (hiPSC) from seven patients harboring mutations across the DMD gene. Mutations were retained during differentiation and analysis indicated the cardiomyocytes showed a dystrophic gene expression profile. Antisense oligonucleotide-mediated skipping of exon 51 restored dystrophin expression to ∼30% of normal levels in hiPSC-cardiomyocytes carrying exon 47-50 or 48-50 deletions. Alternatively, delivery of a dystrophin minigene to cardiomyocytes with a deletion in exon 35 or a point mutation in exon 70 allowed expression levels similar to those seen in healthy cells. This demonstrates that DMD hiPSC-cardiomyocytes provide a novel tool to evaluate whether new therapeutics can restore dystrophin expression in the heart.

Citing Articles

From bench to bedside: cutting-edge applications of base editing and prime editing in precision medicine.

Xu W, Zhang S, Qin H, Yao K J Transl Med. 2024; 22(1):1133.

PMID: 39707395 PMC: 11662623. DOI: 10.1186/s12967-024-05957-3.

An Updated Analysis of Exon-Skipping Applicability for Duchenne Muscular Dystrophy Using the UMD-DMD Database.

Leckie J, Zia A, Yokota T Genes (Basel). 2024; 15(11).

PMID: 39596689 PMC: 11593839. DOI: 10.3390/genes15111489.

State of the art and perspectives of gene therapy in heart failure. A scientific statement of the Heart Failure Association of the ESC, the ESC Council on Cardiovascular Genomics and the ESC Working Group on Myocardial & Pericardial Diseases.

Van Linthout S, Stellos K, Giacca M, Bertero E, Cannata A, Carrier L Eur J Heart Fail. 2024; 27(1):5-25.

PMID: 39576264 PMC: 11798634. DOI: 10.1002/ejhf.3516.

Tracking single hiPSC-derived cardiomyocyte contractile function using CONTRAX an efficient pipeline for traction force measurement.

Pardon G, Roest A, Chirikian O, Birnbaum F, Lewis H, Castillo E Nat Commun. 2024; 15(1):5427.

PMID: 38926342 PMC: 11208611. DOI: 10.1038/s41467-024-49755-3.

Physiological stress improves stem cell modeling of dystrophic cardiomyopathy.

Fullenkamp D, Willis A, Curtin J, Amaral A, Dittloff K, Harris S Dis Model Mech. 2023; 17(6).

PMID: 38050701 PMC: 10820750. DOI: 10.1242/dmm.050487.

References

Matsa E, Rajamohan D, Dick E, Young L, Mellor I, Staniforth A . Drug evaluation in cardiomyocytes derived from human induced pluripotent stem cells carrying a long QT syndrome type 2 mutation. Eur Heart J. 2011; 32(8):952-62. PMC: 3076668. DOI: 10.1093/eurheartj/ehr073. View

Yin H, Saleh A, Betts C, Camelliti P, Seow Y, Ashraf S . Pip5 transduction peptides direct high efficiency oligonucleotide-mediated dystrophin exon skipping in heart and phenotypic correction in mdx mice. Mol Ther. 2011; 19(7):1295-303. PMC: 3128823. DOI: 10.1038/mt.2011.79. View

Ezzat K, Andaloussi S, Zaghloul E, Lehto T, Lindberg S, Moreno P . PepFect 14, a novel cell-penetrating peptide for oligonucleotide delivery in solution and as solid formulation. Nucleic Acids Res. 2011; 39(12):5284-98. PMC: 3130259. DOI: 10.1093/nar/gkr072. View

Dick E, Rajamohan D, Ronksley J, Denning C . Evaluating the utility of cardiomyocytes from human pluripotent stem cells for drug screening. Biochem Soc Trans. 2010; 38(4):1037-45. DOI: 10.1042/BST0381037. View

Lu Q, Yokota T, Takeda S, Garcia L, Muntoni F, Partridge T . The status of exon skipping as a therapeutic approach to duchenne muscular dystrophy. Mol Ther. 2010; 19(1):9-15. PMC: 3017449. DOI: 10.1038/mt.2010.219. View

Wood M, Gait M, Yin H . RNA-targeted splice-correction therapy for neuromuscular disease. Brain. 2010; 133(Pt 4):957-72. DOI: 10.1093/brain/awq002. View

Cirak S, Arechavala-Gomeza V, Guglieri M, Feng L, Torelli S, Anthony K . Exon skipping and dystrophin restoration in patients with Duchenne muscular dystrophy after systemic phosphorodiamidate morpholino oligomer treatment: an open-label, phase 2, dose-escalation study. Lancet. 2011; 378(9791):595-605. PMC: 3156980. DOI: 10.1016/S0140-6736(11)60756-3. View

Heemskerk H, de Winter C, van Kuik P, Heuvelmans N, Sabatelli P, Rimessi P . Preclinical PK and PD studies on 2'-O-methyl-phosphorothioate RNA antisense oligonucleotides in the mdx mouse model. Mol Ther. 2010; 18(6):1210-7. PMC: 2889733. DOI: 10.1038/mt.2010.72. View

Aartsma-Rus A, Fokkema I, Verschuuren J, Ginjaar I, van Deutekom J, van Ommen G . Theoretic applicability of antisense-mediated exon skipping for Duchenne muscular dystrophy mutations. Hum Mutat. 2009; 30(3):293-9. DOI: 10.1002/humu.20918. View

10.

Goemans N, Tulinius M, van den Akker J, Burm B, Ekhart P, Heuvelmans N . Systemic administration of PRO051 in Duchenne's muscular dystrophy. N Engl J Med. 2011; 364(16):1513-22. DOI: 10.1056/NEJMoa1011367. View

11.

Malerba A, Boldrin L, Dickson G . Long-term systemic administration of unconjugated morpholino oligomers for therapeutic expression of dystrophin by exon skipping in skeletal muscle: implications for cardiac muscle integrity. Nucleic Acid Ther. 2011; 21(4):293-8. DOI: 10.1089/nat.2011.0306. View

12.

Foldes G, Mioulane M, Wright J, Liu A, Novak P, Merkely B . Modulation of human embryonic stem cell-derived cardiomyocyte growth: a testbed for studying human cardiac hypertrophy?. J Mol Cell Cardiol. 2010; 50(2):367-76. PMC: 3034871. DOI: 10.1016/j.yjmcc.2010.10.029. View

13.

Sugita H, Takeda S . Progress in muscular dystrophy research with special emphasis on gene therapy. Proc Jpn Acad Ser B Phys Biol Sci. 2010; 86(7):748-56. PMC: 3066538. DOI: 10.2183/pjab.86.748. View

14.

Burridge P, Anderson D, Priddle H, Munoz M, Chamberlain S, Allegrucci C . Improved human embryonic stem cell embryoid body homogeneity and cardiomyocyte differentiation from a novel V-96 plate aggregation system highlights interline variability. Stem Cells. 2006; 25(4):929-38. DOI: 10.1634/stemcells.2006-0598. View

15.

Aartsma-Rus A, Kaman W, Bremmer-Bout M, Janson A, den Dunnen J, van Ommen G . Comparative analysis of antisense oligonucleotide analogs for targeted DMD exon 46 skipping in muscle cells. Gene Ther. 2004; 11(18):1391-8. DOI: 10.1038/sj.gt.3302313. View

16.

van Deutekom J, Bremmer-Bout M, Janson A, Ginjaar I, Baas F, den Dunnen J . Antisense-induced exon skipping restores dystrophin expression in DMD patient derived muscle cells. Hum Mol Genet. 2001; 10(15):1547-54. DOI: 10.1093/hmg/10.15.1547. View

17.

Rando T . The dystrophin-glycoprotein complex, cellular signaling, and the regulation of cell survival in the muscular dystrophies. Muscle Nerve. 2001; 24(12):1575-94. DOI: 10.1002/mus.1192. View

18.

Goyenvalle A, Vulin A, Fougerousse F, Leturcq F, Kaplan J, Garcia L . Rescue of dystrophic muscle through U7 snRNA-mediated exon skipping. Science. 2004; 306(5702):1796-9. DOI: 10.1126/science.1104297. View

19.

Aartsma-Rus A, van Deutekom J, Fokkema I, van Ommen G, den Dunnen J . Entries in the Leiden Duchenne muscular dystrophy mutation database: an overview of mutation types and paradoxical cases that confirm the reading-frame rule. Muscle Nerve. 2006; 34(2):135-44. DOI: 10.1002/mus.20586. View

20.

Dick E, Matsa E, Young L, Darling D, Denning C . Faster generation of hiPSCs by coupling high-titer lentivirus and column-based positive selection. Nat Protoc. 2011; 6(6):701-14. DOI: 10.1038/nprot.2011.320. View