Abstract
KEY POINTS: We report that the small molecule CK-2066260 selectively slows the off-rate of Ca(2)(+) from fast skeletal muscle troponin, leading to increased myofibrillar Ca(2)(+) sensitivity in fast skeletal muscle. Rodents dosed with CK-2066260 show increased hindlimb muscle force and power in response to submaximal rates of nerve stimulation in situ. CK-2066260 has no effect on free cytosolic [Ca(2)(+) ] during contractions of isolated muscle fibres. We conclude that fast skeletal muscle troponin sensitizers constitute a potential therapy to address an unmet need of improving muscle function in conditions of weakness and premature muscle fatigue. ABSTRACT: Skeletal muscle dysfunction occurs in many diseases and can lead to muscle weakness and premature muscle fatigue. Here we show that the fast skeletal troponin activator, CK-2066260, counteracts muscle weakness by increasing troponin Ca(2+) affinity, thereby increasing myofibrillar Ca(2+) sensitivity. Exposure to CK-2066260 resulted in a concentration-dependent increase in the Ca(2+) sensitivity of ATPase activity in isolated myofibrils and reconstituted hybrid sarcomeres containing fast skeletal muscle troponin C. Stopped-flow experiments revealed a ∼2.7-fold decrease in the Ca(2+) off-rate of isolated troponin complexes in the presence of CK-2066260 (6 vs. 17 s(-1) under control conditions). Isolated mouse flexor digitorum brevis fibres showed a rapidly developing, reversible and concentration-dependent force increase at submaximal stimulation frequencies. This force increase was not accompanied by any changes in the free cytosolic [Ca(2+) ] or its kinetics. CK-2066260 induced a slowing of relaxation, which was markedly larger at 26°C than at 31°C and could be linked to the decreased Ca(2+) off-rate of troponin C. Rats dosed with CK-2066260 showed increased hindlimb isometric and isokinetic force in response to submaximal rates of nerve stimulation in situ producing significantly higher absolute forces at low isokinetic velocities, whereas there was no difference in force at the highest velocities. Overall muscle power was increased and the findings are consistent with a lack of effect on crossbridge kinetics. In conclusion, CK-2066260 acts as a fast skeletal troponin activator that may be used to increase muscle force and power in conditions of muscle weakness.