Abstract 15204: Inhibition of Shp2’s Phosphatase Activity Ameliorates Cardiac Hypertrophy in LEOPARD Syndrome Models
Mutations in the protein tyrosine phosphatase Shp2 cause congenital heart disease, in particular Noonan Syndrome (NS) and LEOPARD Syndrome. Due to the substantial clinical overlap between the two syndromes, the latter is also known as NS with multiple lentigines (NSML). Hypertrophic cardiomyopathy (HCM) is very common in NSML patients. In most of these cases, the underlying Shp2 mutations are localized close to the protein’s catalytic pocket. Thorough structure-function analyses have not yet been conducted, but recent evidence indicates that Shp2 has phosphatase-dependent and -independent functions. We therefore tested the role of Shp2’s catalytic activity for the pro-hypertrophic effects of mutant Shp2.
We focused on the Q510E mutation, which causes a particularly aggressive form of HCM in children and therefore is ideally suited for proof-of-principle studies. Using adenoviral delivery, we expressed Q510E-Shp2 in cultured neonatal rat cardiomyocytes (NRCM). This resulted in NRCM hypertrophy with increased cell surface areas, protein content, and protein synthesis rate, and enhanced sarcomeric organization. To test the necessity of an intact catalytic center for Q510E-Shp2’s effects, we then introduced a second mutation into the construct, C459S. The C459S mutation completely abolishes Shp2 phosphatase activity by replacing the cysteine essential for nucleophilic attack of the substrate. The Q510E-C459S double mutant failed to induce NRCM hypertrophy.
Next, we used the pharmacological Shp2 inhibitor PHPS1. PHPS1 was designed to bind at the catalytic site and thereby disrupt interactions between Shp2 and its substrates. 10microM PHPS1 prevented Q510E-Shp2-induced NRCM hypertrophy.
For in vivo validation, we used transgenic mice. Cardiomycoyte-specific expression of Q510E-Shp2 in these mice leads to neonatal-onset HCM. Importantly, PHPS1 (3 mg/kg i.p. 3 times a week for 6 weeks) normalized LV posterior wall thickness and significantly improved contractile function.
In conclusion, the pro-hypertrophic effects of Q510E-Shp2 depend on the protein’s phosphatase activity and can be ameliorated by a pharmacological inhibitor. Translated into the clinic, this would provide a novel custom-tailored approach for NS / NSML patients.
- © 2013 by American Heart Association, Inc.