P236Evaluation of safety and feasibility of Mybpc3 gene therapy in a mouse model of hypertrophic cardiomyopathy.

Cardiovascular Research

PubMedID: 25020650

Mearini G, Simpel D, Geertz B, Kraemer L, Schlossarek S, Weinberger F, Mueller O, Voit T, Eschenhagen T, Carrier L, ISHR. P236Evaluation of safety and feasibility of Mybpc3 gene therapy in a mouse model of hypertrophic cardiomyopathy. Cardiovasc Res. 2014;103 Suppl 1S42.
Hypertrophic cardiomyopathy (HCM) is characterized by left ventricular hypertrophy (LVH) and diastolic dysfunction, and is frequently caused by MYBPC3 mutations, which mainly result in haploinsufficiency of cardiac myosin-binding protein C (cMyBP-C). We recently provided proof-of-concept studies that adeno-associated virus (AAV)-mediated RNA-based therapies (exon skipping and trans-splicing) remove the mutation in a homozygous Mybpc3-targeted knock-in (KI) mouse model of HCM. However, the level of cMyBP-C remained low. In the present study, we evaluated the feasibility and safety of Mybpc3 gene therapy to correct cMyBP-C protein haploinsufficiency and prevent the cardiac phenotype in KI mice.

FLAG-tagged Mybpc3 cDNA under the control of the human TNNT2 promoter was packaged in AAV9. Different doses of AAV9 (10^11, 3x10^11, 10^12, 3x10^12 viral genome (Vg)/mouse) or PBS were administered in the temporal vein of 1-day-old KI mice. Cardiac function was analyzed at different postnatal times until 34 wks by echocardiography and compared to age-matched wild-type (WT) mice. Levels of cMyBP-C protein were determined by Western blot and localization of transgenic protein was evaluated by immunofluorescence with specific antibodies in cardiac sections.

KI mice did not show a cardiac phenotype at postnatal day 1 (P1), but exhibited reduced fractional area shortening (FAS) at P2 and LVH at P3. AAV9-mediated Mybpc3 gene transfer dose-dependently and partially prevented the reduction of FAS and increase in LVM/BW ratio over time. In particular, doses of 10^12 and of 3x10^12 Vg/mouse fully prevented the development of cardiac dysfunction and hypertrophy 24 wks post-administration, respectively. The amount of FLAG-cMyBP-C and therefore of total-cMyBP-C was dose-dependently increased in ventricular tissue of AAV9-FLAG-Mybpc3-treated mice. FLAG-cMyBP-C protein was restricted to the heart and was not found in liver and skeletal muscle. In addition, immunofluorescence of cardiac sections showed the classical sarcomere striation pattern of cMyBP-C as doublet in the A-band.

This study provides proof-of-principle of gene therapy as a feasible treatment for HCM associated with MYBPC3 mutation. This therapeutic strategy is particularly attractive for severe forms of HCM affecting children.