Omecamtiv mercabil and blebbistatin modulate cardiac contractility by perturbing the regulatory state of the myosin filament

Kampourakis, Thomas; Zhang, Xuemeng; Sun, Yin-Biao; Irving, Malcolm

doi:10.1113/jp275050

Cited by 86 publications

(152 citation statements)

References 60 publications

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“…Subsequent studies confirmed that OM increases the rate of phosphate release 6,7 ; however, it was also shown that OM inhibits the velocity of actin gliding in the in vitro motility assay at all concentrations tested 6,[8][9][10] and reduces the rate of tension development and relaxation in myocytes at micromolar concentrations 1,9,[11][12][13] . These phenomena, along with observations of decreased isometric force in fully activated cardiomyocytes, 11,14 are inconsistent with the originally proposed model for OM-activation of myosin in muscle.…”

Section: Introductioncontrasting

confidence: 70%

“…We carried out simulations to determine whether the results we observed in the optical trapping assay could explain the effect of OM on the actin gliding rate in the in vitro motility assay 6,8-10 and on the calcium-dependence of contractile function of cardiomyocytes 11,14 . We used a simple, two-state model of the actomyosin cycle along with cooperative activation of the thin filament as simulated by Walcott et al 27,28 (See Online Methods).…”

Section: Calcium Sensitization and Motility Inhibition By Om Is Explamentioning

confidence: 99%

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Positive Cardiac Inotrope, Omecamtiv Mecarbil, Activates Muscle Despite Suppressing the Myosin Working Stroke

Woody

Greenberg

Barua

et al. 2018

Preprint

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Omecamtiv mecarbil (OM) is a positive cardiac inotrope in phase-3 clinical trials for treatment of heart failure. Although initially described as a direct myosin activator, subsequent studies are at odds with this description and do not explain OM-mediated increases in cardiac output. Single-molecule, biophysical experiments on cardiac myosin show that OM suppresses myosin's working stroke and prolongs actomyosin attachment 5-fold, which explain inhibitory actions of the drug observed in vitro. Surprisingly, the increased myocardial force output in the presence of OM can be explained by cooperative thin filament activation by OM-inhibited myosin molecules. Selective suppression of myosin is an unanticipated route to muscle activation that may guide future development of therapeutic drugs. RESULTS OM Reduces the Size of Myosin's Working StrokeWe kinetically and mechanically characterized the interaction of a recombinant, human -cardiac heavy-meromyosin (HMM) construct with actin in the presence and absence of OM using single-molecule, optical trapping. In this assay, a single actin filament suspended between two optically-trapped polystyrene beads (known as an actin "dumbbell") is brought into proximity with a single myosin molecule adsorbed to a pedestal bead which is anchored to the coverslip surface 15,16 . Single myosin molecules bind and displace the dumbbell when it is near the pedestal, and binding events are detected by analysis of the covariance of the bead position fluctuations 17,18 (see Online Methods). Binding events as short as 12 ms can be detected, and the displacement of the dumbbell by the myosin working stroke is determined with sub-nanometer resolution (Figure 1, inset).

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Section: Introductioncontrasting

confidence: 70%

Section: Calcium Sensitization and Motility Inhibition By Om Is Explamentioning

confidence: 99%

Positive Cardiac Inotrope, Omecamtiv Mecarbil, Activates Muscle Despite Suppressing the Myosin Working Stroke

Woody

Greenberg

Barua

et al. 2018

Preprint

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“…The mechanism of action of a cardiac myosin activator, Omecamtiv mecarbil (OM) has been elucidated: OM favors the PPS conformation and thus increases the number of heads able to participate in force production upon the systole on-set 11 . Interestingly, a recent study has concluded that OM is not compatible with the sequestered state of cardiac myosin while the myosin inhibitor blebbistatin (BS) favors the sequestered state 40 . Our results are consistent with this study since the site of blebbistatin is internal and is not affected by IHM contacts.…”

Section: Implications For Future Treatmentsmentioning

confidence: 99%

Hypertrophic cardiomyopathy disease results from disparate impairments of cardiac myosin function and auto-inhibition

Robert-Paganin

Auguin²,

Houdusse³

2018

Preprint

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Abstract:Hypertrophic cardiomyopathies (HCM) result from distinct single-point mutations in sarcomeric proteins that lead to muscle hypercontractility. While different models account for a pathological increase in the power output, clear understanding of the molecular basis of dysfunction in HCM is the mandatory next step to improve current treatments. An optimized quasi-atomic model of the sequestered state of cardiac myosin coupled to X-ray crystallography and in silico analysis of the mechanical compliance of the lever arm allowed us to perform a systematic study for a large set of HCM mutations. We define different classes of mutations depending on their effects on lever arm compliance, sequestered state stability and motor functions. This work reconciles previous models and explains how distinct HCM mutations can have disparate effects on the motor mechano-chemical parameters and yet lead to the same disease. The framework presented here can guide future investigations aiming at finding HCM treatments.

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“…7) will be important in contracting cardiac muscle. Mechanisms likely regulating this transition include RLC phosphorylation [35], myosin-binding protein-C [31], thick-filament mechano-sensing [36], and small molecules [37].…”

Section: Resultsmentioning

confidence: 99%

Mavacamten stabilizes the auto-inhibited state of two-headed cardiac myosin

Rohde

Thomas

Muretta

2018

Preprint

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We used transient biochemical and structural kinetics to elucidate the molecular mechanism of mavacamten, an allosteric cardiac myosin inhibitor and prospective treatment for hypertrophic cardiomyopathy. We find that mavacamten stabilizes an auto-inhibited state of two-headed cardiac myosin, not found in the isolated S1 myosin motor fragment. We determined this by measuring cardiac myosin actin-activated and actin-independent ATPase and single ATP turnover kinetics. A two-headed myosin fragment exhibits distinct auto-inhibited ATP turnover kinetics compared to a single-headed fragment. Mavacamten enhanced this auto-inhibition. It also enhanced auto-inhibition of ADP release. Furthermore, actin changes the structure of the auto-inhibited state by forcing myosin lever-arm rotation. Mavacamten slows this rotation in two-headed myosin but does not prevent it. We conclude that cardiac myosin is regulated in solution by an interaction between its two heads and propose that mavacamten stabilizes this state. Significance StatementSmall-molecule allosteric effectors designed to target and modulate striated and smooth myosin isoforms for the treatment of disease show promise in preclinical and clinical trials. Beta-cardiac myosin is an especially important target, as heart disease remains a primary cause of death in the U.S. One prevalent type of heart disease is hypertrophic cardiomyopathy (HCM), which is hypothesized to result from dysregulated force generation by cardiac myosin. Mavacamten is a potent cardiac myosin ATPase activity inhibitor that improves cardiac output in HCM animal models. Our results show that mavacamten selectively stabilizes a two-head dependent, auto-inhibited state of cardiac myosin in solution. The kinetics and energetics of this state are consistent with the auto-inhibited super-relaxed state, previously only observed in intact sarcomeres. Introduction.Familial hypertrophic cardiomyopathies, abbreviated HCM, represent one of the most common classes of genetic disease, affecting 1 in 500 people [1]. HCM is hypothesized to result from cardiac hyper-contractility [2]. Direct inhibition of force generation by cardiac myosin is thus a putative treatment [3]. Mavacamten (Mava), previously termed Myk461, is a small-molecule allosteric inhibitor of cardiac myosin that shows promise in preclinical and clinical trials for the treatment of HCM [3]. Mavacamten binds with sub-micromolar affinity to cardiac myosin in the presence of adenosine triphosphate (ATP) and inhibits steady-state actin-activated and actin-independent (basal) ATPase cycling and calcium regulated ATPase activity in permeablized cardiac myofibrils [3,4]. Mavacamten also inhibits the kinetics of rigor actin-binding as well as the kinetics of actin-activated phosphate release [4]. It decreases in vitro actin filament sliding velocity in the actomyosin motility assay [4], decreases force generation by skinned cardiac preparations from mouse HCM models [3], and decreases cardiac output in live feline hearts [5]. Mavacamten's effect on cardia...

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Omecamtiv mercabil and blebbistatin modulate cardiac contractility by perturbing the regulatory state of the myosin filament

Cited by 86 publications

References 60 publications

Positive Cardiac Inotrope, Omecamtiv Mecarbil, Activates Muscle Despite Suppressing the Myosin Working Stroke

Positive Cardiac Inotrope, Omecamtiv Mecarbil, Activates Muscle Despite Suppressing the Myosin Working Stroke

Hypertrophic cardiomyopathy disease results from disparate impairments of cardiac myosin function and auto-inhibition

Mavacamten stabilizes the auto-inhibited state of two-headed cardiac myosin

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