Kade Scoresby

Introduction. Hypertrophic cardiomyopathy (HCM) is the most common heritable cardiomyopathy, affecting about 1 in 500 people.¹ HCM is genetically inherited, with mutations in the MYH7 (β-myosin heavy chain protein) and MYBPC3 (myosin-binding protein C) genes accounting for approximately 70% of HCM cases.^2,3 The most common HCM treatments include beta-blockers, calcium channel blockers, and implantable cardioverter defibrillators (ICD) to prevent sudden cardiac death.⁴ More recently, mavacamten, a cardiac-specific myosin ATPase inhibitor, shows promise in preventing adverse events related to HCM.⁵ However, none of the aforementioned interventions address the genetic cause of HCM, which, if targeted, may be more successful in slowing the rapid and devastating progression of HCM. In the near future, gene silencing therapy using sequence-specific RNA molecules to silence the expression of mutant genes may substantially lower autosomal familial risk for HCM.⁶ The MYH7-R403Q mutation causes impaired actin-myosin interaction due to destabilization of the myosin super-relaxed state (SRX), thereby increasing the number of myosin heads available to bind actin.^3,7 This mutant protein causes increased contractility and impaired relaxation in HCM patients, making the MYH7-R403Q mutant transcript a potential study target for gene silencing therapy.⁸ Methods. A study used human induced pluripotent stem cell cardiomyocytes (hiPSC-CMs) derived and differentiated from two siblings with a heterozygous MYH7-R403Q mutation.⁹ Short hairpin RNA (shRNA) and antisense oligonucleotides (ASO) were investigated as potential agents for gene silencing.⁹ Specific shRNA for MYH7-R403Q was transduced into hiPSC-CMs via a viral vector.⁹ ASOs were custom-designed to correct the mutation and transfected to the hiPSC-CMs using a transfection reagent (TransIT- TKO, Mirus Bio).⁹ Gene expression for both methods was measured using qPCR. Additionally, cell contractility was measured using traction force microscopy and cell size was measured by staining for α-actinin, actin, and DAPI.⁹ Results. Strikingly, the shRNA reduced R403Q expression by 57% in hiPSC-CMs and ASO reduced expression by 49%, with shRNA showing more significant molecular and contractile benefits, returning contractile properties to wild-type values.⁹ However, a potential drawback observed was that the shRNA also reduced the expression of the wild-type allele (33%).⁹ Furthermore, ASO was not specific to R403Q, increasing the likelihood of off-target silencing.⁹ Conclusions. Allele-specific RNA silencing of MYH7 gene mutations may provide an alternative treatment option for HCM that conveys wild-type phenotypes and is less invasive. Future studies will investigate the long-term effects of gene silencing, comparing the effects of additional RNA molecules, and the delivery options for silencing agents to patients.

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