Discussion
This study contributes new biophysical observations about the effects of
mavacamten on human cardiac muscle function at physiological
temperature. With recent therapeutic treatments for heart failure
showing promise at the sarcomere level (Teerlink et al., 2016; Heitner
et al., 2019), there is a need to better define and identify molecular
mechanisms of contraction that may ultimately be targeted to benefit
patient care. Sarcomere length-dependent increases in
Ca2+-sensitivity of contraction represent a key
physiological relationship at the cellular and tissue level that
underlies the Frank-Starling response in healthy hearts. Pharmaceuticals
that compromised length-dependent activation may ultimately prove to be
less effective therapies for heart disease (Gollapudi et al., 2017;
Henze et al., 2019). Herein we show that mavacamten preserves sarcomere
length-dependent increases in the Ca2+-sensitivity of
force, while reducing maximum levels of force production.
These data have implications for mavacamten affecting systolic and
diastolic aspects of cardiac muscle function. On the systolic side, HCM
has typically been viewed as a hypercontractile phenotype due to a ‘gain
of function’ mutation (Moore et al., 2012; Green et al., 2016). Thus,
reducing the Ca2+-sensitivity of the myocardium with
mavacamten treatment could reduce the hypercontractile state during
systole, normalizing function and potentially leading to beneficial
remodeling of the heart. Administering mavacamten (early in life) to
transgenic mice carrying the myosin R403Q mutation, which typically
displays a potent HCM phenotype, helped reduce ventricular hypertrophy,
cardiomyocyte disarray, and myocardial fibrosis (Green et al., 2016). On
the other side of a heartbeat, diastolic dysfunction leads to impaired
relaxation and ventricular filling, which underlies roughly 50% of
heart failure diagnoses (Aljaroudi et al., 2012; Lekavich et al., 2015;
Mozaffarian et al., 2016). The findings that mavacamten reduced passive
tension at both sarcomere lengths (Fig. 3B) and increased cross-bridge
detachment rates under maximally activated conditions (Fig. 6B) suggests
that mavacamten may improve relaxation.
Prior biophysical assays have shown that mavacamten slows cross-bridge
recruitment, Pi release, and actin-associated ADP
release, all of which underlie strong cross-bridge binding and
force-generation (Green et al., 2016; Kawas et al., 2017; Rohde et al.,
2018). In skinned myocardial mechanics and intact myocyte assays
mavacamten reduced contractility in a dose-dependent manner, slowing the
rate of force development (Mamidi et al., 2018) and increasing the
relaxation rate (Toepfer et al., 2019b, 2019a). Our frequency-dependent
viscoelastic stiffness data support these findings in human myocardium
at physiological temperature (Fig. 5). Mavacamten shifted the negative
viscous moduli-frequency relationships towards lower frequencies and
reduced the amount of work generated by the myocardiaum (Fig. 5C-D),
consistent with impaired force generation (Fig. 2) and slowed
cross-bridge recruitment (Fig 6A). The frequency range where negative
viscous moduli occur reflects the physiological range of heart rates
within a species, and the diminished range for mavacamten-treated strips
could indicate that mavacamten plays a limiting role at the upper end of
this range. Our findings that mavacamten speeds cross-bridge detachment
(Fig. 6B) agrees with data showing that mavacamten increases relaxation
rates in human myofibrils (Scellini et al., 2020). Similar measurements
by Scellini et al. show that mavacamten had no effect on relaxation rate
in rabbit skeletal myofibrils, which suggests that the effects of
mavacamten may vary with species and muscle type (Scellini et al.,
2020). Additional studies will be required to more fully define the
potential impacts of mavacamten on different myosin isoforms.
Previously, we used mathematical models to test how mechanosensitive
contributions of the myosin OFF-ON equilibrium influence
length-dependent activation of contraction (Campbell et al. 2018). These
data showed that elevated force levels, from passive and/or active
mechanisms, amplify cross-bridge recruitment and force production, more
fully illustrating the fascinating (yet complicated) impacts of dynamic
filament regulation (Fig. 1) that underly force production and
relaxation throughout a heartbeat. Knowing that mavacamten stabilizes
the OFF state (Anderson et al., 2018; Rohde et al., 2018), we wondered
if sarcomere length-dependent increases in
Ca2+-sensitivity would be amplified in mavacamten
treated strips. Our data do not show this response in a statistically
significant manner (Fig. 4A), although the pCa50 change
between 1.9 µm and 2.3 µm was 3-fold greater for mavacamten treated
strips (on average ΔpCa50=0.03 for control vs. 0.09 for
mavacamten-treated strips). These findings differ from a preliminary
report using permeabilized myocardial strips from pigs, where mavacamten
suppressed length-dependent increases from OFF-to-ON and abolished
length-dependent increases in Ca2+-sensitivity of
contraction (Henze et al., 2019). The functional effects of the OFF-ON
equilibrium are only starting to come to light for the muscle physiology
research field, yet this equilibrium represents a critical mechanism
underlying regulatory coupling between the thick- and thin-filaments and
the Frank-Starling law of the heart. One interesting aspect of this
OFF-ON relationship is the force-dependent recruitment of myosin from
the OFF-to-ON state, which encompasses a force-feedback effect to
amplify contractility as force develops during isometric contractions
and when muscles are stretched (Ait-Mou et al., 2016; Campbell, 2017;
Fusi et al., 2017; Reconditi et al., 2017; Zhang et al., 2017; Piazzesi
et al., 2018; Brunello et al., 2020). Thus, any means of modulating
length-dependent changes Ca2+-sensitivity leads to
significant functional effects, because myofilaments are highly
cooperative and subtle shifts in pCa50 can produce large
change in force-production as muscle length changes. Prior to the
discovery of force-dependent OFF-ON transitions, length-dependent
changes in Ca2+-sensitivity were typically thought to
reflect changes in thin filament function and/or the binding kinetics of
individual myosin heads (Gordon et al., 2000). Therefore, ongoing and
future research testing the effects of mavacamten to preserve or even
amplify length-dependent activation in transgenic models of heart
failure and human tissue samples from heart failure patients will be
helpful for informing the potential scope of mavacamten as a therapeutic
treatment for cardiac dysfunction. More generally, these findings also
imply that the OFF-ON equilibrium of myosin may be a particularly
important therapeutic target for modulating myocardial function.