Rationally engineered Troponin C modulates in vivo cardiac function and performance in health and disease

Abstract

Treatment for heart disease, the leading cause of death in the world, has progressed little for several decades. Here we develop a protein engineering approach to directly tune in vivo cardiac contractility by tailoring the ability of the heart to respond to the Ca2+ signal. Promisingly, our smartly formulated Ca2+-sensitizing TnC (L48Q) enhances heart function without any adverse effects that are commonly observed with positive inotropes. In a myocardial infarction (MI) model of heart failure, expression of TnC L48Q before the MI preserves cardiac function and performance. Moreover, expression of TnC L48Q after the MI therapeutically enhances cardiac function and performance, without compromising survival. We demonstrate engineering TnC can specifically and precisely modulate cardiac contractility that when combined with gene therapy can be employed as a therapeutic strategy for heart disease.

A Tension-Based Model Distinguishes Hypertrophic versus Dilated Cardiomyopathy

“…There are other examples of TG and knock-in mouse models of HCM that fail to phenocopy the asymmetric hypertrophic thickening of the myocardium observed in HCM patients, even independently generated L48Q TG mice (Shettigar et al, 2016; Davis et al, 2012; Michele et al, 2002). Thus, we hypothesized that the effects of L48Q cTnC are masked by the known high adrenergic drive and heart rate of the mouse, which accelerates diastolic function through increasing Ca 2+ reuptake by SERCA2 and desensitizing the sarcomere to Ca 2+ by phosphorylating the amino terminal serine residues (23/24) on cTnI (Metzger and Westfall, 2004).…”

Overexpression of miR-223 Tips the Balance of Pro- and Anti-hypertrophic Signaling Cascades toward Physiologic Cardiac Hypertrophy

“…To compare the performance between miR-223 TG and WT mice, age-matched mice were acclimatized and subjected to treadmill exercise training. The same protocol was applied as above except the speed was increased by 2 m/min every minute to up to 30 m/min, and total duration was recorded when the mice reached exhaustion (unresponsive to stimuli for 10s) (47).…”

“…Mice were trained 5 days a week for 4 weeks, heart samples were collected at the end of weeks 1, 2, and 4. Heart weight to body weight ratio was calculated (47). To compare the performance between miR-223 TG and WT mice, age-matched mice were acclimatized and subjected to treadmill exercise training.…”

Gene Transfer of Engineered Calmodulin Alleviates Ventricular Arrhythmias in a Calsequestrin‐Associated Mouse Model of Catecholaminergic Polymorphic Ventricular Tachycardia

“…The construct also contained a sequence encoding mCherry for transfection verification 24. Figure 6A demonstrates that ≈40% of the ventricular myocytes could be transduced with our viral construct based on mCherry fluorescence.…”

“…Adeno‐ associated viral vector serotype 9 (AAV9) vectors were produced by co‐transfecting HEK293 cells with three plasmids as previously described,24 which were then expanded in 10% DMEM medium. Five days after transfection, the cells were harvested for downstream purification using PEG precipitation and an iodixanol gradient protocol 25.…”