CRISPR/Cas9 gene editing in induced pluripotent stem cells to investigate the feline hypertrophic cardiomyopathy causing MYBPC3/R820W mutation

PLoS One. 2024 Oct 10;19(10):e0311761. doi: 10.1371/journal.pone.0311761. eCollection 2024.

Abstract

Hypertrophic cardiomyopathy (HCM) is the most common heart disease in domestic cats, often leading to congestive heart failure and death, with current treatment strategies unable to reverse or prevent progression of the disease. The underlying pathological processes driving HCM remain unclear, which hinders novel drug discovery. The aim of this study was to generate a cellular model of the feline HCM-causing MYBPC3 mutation R820W. Using CRISPR/Cas9 gene editing we introduced the R820W mutation into a human induced pluripotent stem cell (iPSC) line. We differentiated both homozygous mutant clones and isogenic control clones to cardiomyocytes (iPSC-CMs). Protein quantification indicated that haploinsufficiency is not the disease mechanism of the mutation. Homozygous mutant iPSC-CMs had a larger cell area than isogenic controls, with the sarcomere structure and incorporation of cMyBP-C appearing similar between mutant and control iPSC-CMs. Contraction kinetic analysis indicated that homozygous iPSC-CMs have impaired relaxation and are hypocontractile compared to isogenic control iPSC-CMs. In summary, we demonstrate successful generation of an iPSC model of a feline MYBPC3 mutation, with the cellular model recapitulating aspects of HCM including cellular hypertrophy and impaired relaxation kinetics. We anticipate that further study of this model will lead to improved understanding of the disease-causing molecular mechanism, ultimately leading to novel drug discovery.

MeSH terms

  • Animals
  • CRISPR-Cas Systems*
  • Cardiomyopathy, Hypertrophic* / genetics
  • Cardiomyopathy, Hypertrophic* / pathology
  • Cardiomyopathy, Hypertrophic* / veterinary
  • Carrier Proteins* / genetics
  • Cat Diseases / genetics
  • Cat Diseases / pathology
  • Cats
  • Cell Differentiation
  • Gene Editing*
  • Humans
  • Induced Pluripotent Stem Cells* / metabolism
  • Mutation*
  • Myocytes, Cardiac* / metabolism
  • Myocytes, Cardiac* / pathology

Substances

  • myosin-binding protein C
  • Carrier Proteins

Grants and funding

Royal Veterinary College grant funded by Boehringer Ingelheim Vetmedica and The Beryl Evetts and Robert Luff Welfare Trust (4985 awarded to DC) and The Winn Feline Foundation (W17-008, awarded to DC and JD). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.