Cardiac Myosin Activators: Up and Coming

This article refers to ‘Ribonucleotide reductase-mediated increase in dATP improves cardiac performance via myosin activation in a large animal model of heart failure’, by S. Kadota et al., published in this issue on pages XXX–XXX. Despite the central role of decreased contractile function in the pathogenesis of heart failure with reduced ejection fraction (HFrEF), many promising inotropic agents have failed to demonstrate clinical benefits outside of cardiogenic shock.1 The dissociation between improved inotropy and clinical outcomes with these agents may be due to their mechanisms of action and effect on cellular energetics and calcium handling rather than a requisite adverse impact of increased myocardial contractility. Improved contractile function thus remains a viable therapeutic target. Myocardial contractility is produced at the cellular level by the interaction of myosin and actin filaments, with their relationship dynamics modulated by ATP, calcium, and regulatory proteins.2 The details of the myosin cycle are informative. Overall, the chemical energy of ATP hydrolysis is coupled to mechanical movement of ∼10 nm by each activated myosin head. In resting myocardium, troponin and tropomyosin bind to actin and prevent myosin attachment. Following myocyte depolarization, calcium is released from the sarcoplasmic reticulum as well as imported from the extracellular space. The increased calcium binds troponin, allowing the myosin cycle to proceed. ATP binding releases myosin in its strongly bound post-contraction state from actin. Hydrolysis of ATP to ADP resets the myosin lever arm to a cocked position. Release of the hydrolysed phosphate occurs concomitant with actin binding and is coupled to the force producing movement of the myosin lever arm. The cycle then ends with the dissociation of ADP from the myosin and the myosin–actin complex awaiting the arrival of the next ATP. Currently available inotropic agents including beta-adrenergic agonists, phosphodiesterase inhibitors, and levosimendan, as well as many in the development pipeline including nitroxyl donors, modify this system either by increasing cellular calcium or by sensitizing the cardiomyocyte to endogenous calcium fluxes.1 Increased cytoplasmic calcium augments sarcomere activation and the velocity of contraction development (dP/dt). However, it also induces myocardial ischaemia and arrhythmias, clinically significant side effects of these agents. Some of the novel agents designed to improve cardiac contractility possess activities apart from their calcium-mediated myocardial effects, such as vasodilation, that may yet determine their utility. Enter the novel therapeutic class of cardiac myosin activators. These agents directly modulate the myosin and actin mechanism by accelerating the rate-limiting step of the myosin cycle, actin binding coupled to phosphate release. Although the force generated by each myosin head does not change, overall contractile function improves. Normally only a fraction of available myosin heads participate in each cardiac cycle; by speeding up the rate-limiting step, cardiac myosin activators allow a greater fraction of myosin to bind actin for each cycle, producing an augmented power stroke. The most advanced of the myosin activator class is omecamtiv mecarbil, a small molecule identified by optimization of compounds found by high-throughput screening of a biochemically reconstituted sarcomere.3 It increases the duration of cardiac contraction without increasing dP/dt, and functions independently of calcium homeostasis. Because it does not increase dP/dt, systole lengthens and diastole shortens. In canine models of both ischaemic and pacing-induced cardiac dysfunction, as well as a phase I healthy volunteer study and a phase II human HFrEF study, omecamtiv mecarbil produced concentration-dependent increases in stroke volume and LV ejection time.4-6 The dose-limiting effect is cardiac ischaemia at excessive concentrations, presumably due to diastolic shortening and consequent reductions in coronary perfusion. Initiation of a phase III programme awaits the final results of the Acute Treatment with Omecamtiv Mecarbil to Increase Contractility in Acute Heart Failure (ATOMIC-AHF) study, a completed phase II study of hospitalized patients with acute HFrEF (ClinicalTrials.gov NCT01300013), and the Chronic Oral Study of Myosin Activation to Increase Contractility in Heart Failure (COSMIC-HF; ClinicalTrials.gov NCT01786512), a phase II formulation and dose-finding study of oral omecamtiv mecarbil. In this setting, Kadota and colleagues have published here the use of a a distinct direct myosin activator in a large animal model.7 They tested an adeno-associated virus that delivers ribonucleotide reductase (RNR) gene therapy in an ischaemic porcine model of HFrEF. The foundation for this work was laid extensively by this same research group over the past two decades.8-10 The expressed RNR increases the intracellular fraction of 2-deoxy-ATP (dATP) which has been demonstrated to increase myosin activity in tissue culture and murine models. Cardiac-specific production of the enzyme complex is enabled with a troponin T promotor element, which ideally circumvents off-target effects, and it appears that <10% of the ATP pool needs to be replaced by dATP to facilitate these changes. Chronic RNR overexpression appears to lack adverse effects in a transgenic mouse model followed for 5 months.11 The dATP mechanism is similar to that of omecamtiv mecarbil in that both accelerate the rate-limiting step of myosin activation; however, some specific properties of dATP-mediated actuation are distinct. Biochemically, the change in substrate from ATP to dATP induces a conformational change in the actin-binding surface to allow increased myosin–actin attachment with stronger and quicker force generation. Inflated intracellular dATP levels increase myocardial contractile force and dP/dt without affecting lusitropy, thus preserving diastole. The study by Kadota and colleagues is the next step in the progression of this therapeutic. In a small unblinded pilot study of 17 Yucatan minipigs, ischaemic cardiomyopathy was generated by transient balloon occlusion of the left anterior descending coronary artery, a well-studied model system.7 However, this model is limited by a variable degree of systolic dysfunction (EF ranged from 30% to 55% following injury) and high mortality due to refractory arrhythmias (11 of the 28 pigs died prior to evaluation). Two weeks after the induction of HF, the surviving affected pigs were randomized to coronary infusion of adeno-associated virus (BB-R12 vector) or control solution. Echocardiographic and haemodynamic measurements were made until the pigs were euthanized 56 days following treatment. Although the variability of the model and the small number of animals used markedly limit the power of the study and its ability to recapitulate the findings of earlier work, there were modest but variable improvements in myocardial contractility and haemodynamics with high dose gene therapy, primarily apparent when analysing those animals that initially developed more severe systolic dysfunction. The authors should be applauded, as moving from bench to bedside is a colossal task. One major inflection point is the cumbersome transition to a large animal model to confirm advances characterized in cell culture or small animal settings. Although this study is small with significant weaknesses, the authors establish enough evidence to encourage expanded and more robust investigations.7 Nevertheless, hurdles and questions remain, and the enticing promise of this study needs to be balanced by the context of the findings. The intrinsic variability in this animal model and small sample sizes limit the robustness of these results, and the biological plausibility of the benefit could have been more adequately investigated. The effectiveness of the intervention on a biological level could have been reinforced by measuring RNR protein, addressing whether treatment actually increases dATP to the requisite levels in the myocardium, and demonstrating appropriate dose responsiveness. Furthermore, porcine–human differences may undermine the relevance of these results for predicting effects in humans. For instance, while dATP increases contractile force without calcium sensitization in porcine cultures, experiments with dATP in failing human heart tissue demonstrated a significant calcium sensitization effect.12, 13 In these experiments, the intracoronary injection was performed through the infarct-related artery; however, many patients with ischaemic cardiomyopathy have chronic total coronary occlusions and may require delivery method alteration. Finally, the durability of this intervention will need to be characterized, as will the potential adverse and possibly neoplastic effects of the presence of supratherapeutic concentrations of dATP on the fidelity of DNA polymerase.14 Apart from the above concerns, the hope that greeted the conception of gene therapy as a therapeutic agent should be tempered with the reality of its implementation. Although multiple pre-clinical experiments have suggested the potential of adeno-associated virus gene therapy in cardiovascular disease, much of this territory is uncharted. Only one agent in the arena of cardiovascular disease has made it to phase II trials, gene therapy to boost the calcium level in the sarcoplasmic reticulum to increase contractility. However, the EF did not increase significantly with treatment in the Calcium Upregulation by Percutaneous Administration of Gene Therapy in Cardiac Disease (CUPID) phase II HFrEF trial, and recently the CUPID-2 phase II trial did not meet either its primary or its secondary endpoints.15 Additional obstacles universal to gene therapy trials have been identified. Because adeno-associated viruses are endemic, there is antiviral immunity in the human population that limits their clinical utility. In the Kadota trial, pigs with evidence of viral immunity were excluded, and though the proportion of pigs in which this was the case was not reported, in the human trial of cardiac gene therapy >50% of the population were excluded for this reason.15 Nonetheless, given the tremendous and growing size of the HF population, the authors should be praised for their achievement and urged to continue their investigations. One day this innovative therapeutic approach may improve outcomes for these extremely ill patients. Conflict of interest: J.R.T. has received research funding and honoraria from Amgen, Cytokinetics, Johnson & Johnson, Novartis, Relypsa, Sorbent, Trevena, and ZS Pharma. M.A.P. has no conflicts to declare.