Effect of the Novel Myotrope Danicamtiv on Cross‐Bridge Behavior in Human Myocardium

Figures

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  Figure 1. Representative cross‐bridge dynamic stretch activation. A, The dynamic stretch activation and force redevelopment are analyzed by holding myocardial preparations to a new muscle length for 8 seconds and returning to the original muscle length. The dynamic stretch activation is recorded during the initial 8 seconds and then the force redevelopment (k tr) is analyzed.B, An expanded view of dynamic stretch activation is shown. The sudden 2% stretch in muscle length produces an instantaneous spike in the force response (P1). This is due to a sudden strain of elastic elements within the strongly bound cross‐bridges (Phase 1). Then, the force rapidly decays (phase 2) to a minimum with a dynamic rate constant krel, an index of the rate of cross‐bridge detachment from actin. The magnitude of strained cross‐bridge detachment into a non–force‐bearing state is shown by P2, which is the lowest point of phase 2. A gradual ascent of force (phase 3) occurs after P2 with a dynamic rate constant kdf, which is an index of the rate of crossbridge recruitment as cross bridges transition into the force‐bearing state. The new steady‐state force (P3) is achieved from the prestretch steady‐state force…

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  Figure 2. Force relative percentage increase with omecamtiv mecarbil (OM) or danicamtiv (DN). The relative percentage increase with drugs was calculated by the net increase in parameters from the untreated baseline (pre‐OM or pre‐danicamtiv) after OM (0.5 μmol/L) or danicamtiv (2 μmol/L) incubation. Values are represented as mean±SEM. Eight hearts were used for the study, and 3 to 5 myocardial fibers from 1 heart were measured. Comparisons between the pre‐ group and the post‐ (OM or danicamtiv) group were analyzed with a linear mixed model with nested 2‐way ANOVA (ns=0.4). DN indicates danicamtiv; and OM, omecamtiv mecarbil.

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  Figure 3. Effect of omecamtiv mecarbil (OM) and danicamtiv (DN) on Ca2+ sensitivity in donor myocardium. The effect of OM and danicamtiv on the force‐pCa relationships was analyzed with the nonlinear fit (sigmoidal dose–response). Skinned myocardial preparations were incubated with OM (0.5 μmol/L) or danicamtiv (2 μmol/L) before analyzing in each pCa. Values are represented as mean±SEM. Five hearts were used for the study. DN indicates danicamtiv; and OM, omecamtiv mecarbil.

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  Figure 4. Representative cross‐bridge detachment kinetics in donor myocardium prior to and after omecamtiv mecarbil (OM) or danicamtiv (DN) incubations. Representative cross‐bridge detachment responses after a sudden 2% stretch in muscle length. The graph was analyzed by averaging the force response of 2 to 3 myocardial fibers from 6 hearts. Expanded views of gray solid lines are shown below A (pre‐ and posttreatment of OM [0.5 μmol/L]) and B (pre‐ and posttreatment of danicamtiv [2 μmol/L]) to demonstrate biphasic kinetics of k rel. The graph shows k rel (FAST) at the beginning of the detachment kinetic following k rel (SLOW) until the end of the detachment kinetic (ie, P2, T d, which is represented as blue arrows). DN indicates danicamtiv; OM, omecamtiv mecarbil; and T d, time required to reach the total cross‐bridge detachment

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  Figure 5. Relative percentage change of cross‐bridge detachment parameters with omecamtiv mecarbil (OM) or danicamtiv (DN). The parameters were measured at baseline or after a 5‐min incubation with OM (0.5 μmol/L) or danicamtiv (2 μmol/L). The net increase in parameters from the untreated baseline (pre‐OM or pre‐danicamtiv) after OM or danicamtiv incubation was calculated and is expressed as a relative percentage change in parameters from baseline. A, P1/P relative percentage change with drugs (ns=0.2). B, a (SLOW) relative percentage change with drugs (ns=0.4). C, b (FAST) relative percentage change with drugs (ns=0.4). D, k rel, (SLOW) (s−1) relative % decrease with drugs (P=0.02). E, k rel (FAST) (s−1) relative percentage decrease with drugs (ns=0.09). F, P2/P relative percentage change with drugs (ns=0.5). G, T d relative percentage change with drugs (P=0.002). Values are represented as mean±SEM. Eight hearts were used for the study, and 3 to 5 myocardial fibers from 1 heart were measured. Comparisons between the pre‐ group and the post‐ (OM or danicamtiv) group were analyzed with a linear mixed model with nested 2‐way ANOVA. DN indicates danicamtiv; k rel, rate constant of force…

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  Figure 6. Relative percentage change of cross‐bridge recruitment parameters with omecamtiv mecarbil (OM) or danicamtiv (DN). The parameters were measured at baseline or after a 5‐min incubation with OM (0.5 μmol/L) or danicamtiv (2 μmol/L). The net increase in parameters from the untreated baseline (pre‐OM or pre‐danicamtiv) after OM or danicamtiv incubation was calculated and is expressed as relative % change in parameters from baseline. A, k df (s−1) relative % decrease with drugs (P<0.0001). B, P3/P relative % change with drugs (ns=0.5). C, P df relative % change with drugs (ns=0.6). D, T r relative % change with drugs (P=0.002). E, T pk relative % change with drugs (P=0.0004). Values are represented as mean±SEM. Eight hearts were used for the study, and 3 to 5 myocardial fibers from 1 heart were measured. Comparisons between the pre‐ group and post‐ (OM or danicamtiv) group were analyzed with a linear mixed model with nested 2‐way ANOVA. DN indicates danicamtiv; k df, rate constant of delayed force development; OM, omecamtiv mecarbil; P3/P, the new steady state force; P df, amplitude of cross‐bridge recruitment; and T pk, time required to achieve a steady‐state force following a…

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  Figure 7. Representative stretch activation responses in donor myocardium prior to and after omecamtiv mecarbil (OM) or danicamtiv (DN) incubations. Representative cross‐bridge recruitment responses are shown after a sudden 2% stretch in muscle length. The graph was analyzed by averaging the force response of 2 to 3 myocardial fibers from 6 hearts. Orange arrows represent T pk, which is the time required to reach steady‐state force. A, Cross‐bridge recruitment response of pre‐ and posttreatment of OM. B, Cross‐bridge recruitment response of pre‐ and posttreatment of danicamtiv. Expanded views of gray solid lines are shown below A and B to demonstrate T r (green arrows), which is the time required to start force development. DN indicates danicamtiv; OM, omecamtiv mecarbil; T pk, time required to reach steady‐state force; and T r, time required to start force development.

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  Figure 8. Effect of omecamtiv mecarbil (OM) and danicamtiv (DN) on force redevelopment (k tr) in donor myocardium. A, Representative force redevelopment of pre‐ and post‐OM. B, Representative force redevelopment of pre‐ and post‐danicamtiv. The red arrows represent the time to achieve peak force development. Representative force redevelopment graphs are shown by normalizing the force responses to prestretch activated force with 2 to 3 myocardial fibers from 6 hearts. C, The net decrease in k tr (s−1) from the untreated baseline (pre‐OM or pre‐danicamtiv) after OM or danicamtiv incubation was calculated and is expressed as a relative percentage decrease from baseline. Values of relative percentage decrease are represented as mean±SEM. Eight hearts were used for the study, and 3 to 5 myocardial fibers from 1 heart were measured. Comparisons between the pre‐ group and the post‐ (OM or danicamtiv) group were analyzed with a linear mixed model with nested 2‐way ANOVA (P<0.0001). DN indicates danicamtiv; k tr, the rate of force redevelopment; and OM, omecamtiv mecarbil.