RPI‐194 is a Novel Troponin Activator that Increases the Calcium Sensitivity of Striated Muscle Contraction
Bibliographic record
Abstract
Introduction Systolic heart failure is a cardiac disease in which the contractile function of heart muscle becomes weakened and unable to pump blood with adequate force. As of yet, no existing positive inotrope is able to lower the mortality rate in systolic heart failure. Cardiac troponin complex (cTn) is a trimeric complex consisting of troponin C (cTnC), troponin I (cTnI) and troponin T (cTnT). cTnC regulates blood supply to the heart by turning muscle contraction on and off in a calcium‐dependent manner. More precisely, the regulatory N‐domain of cTnC (cNTnC) binds the switch region of cTnI to activate contraction. Drug‐like molecules need to bind to this cNTnC‐cTnI switch region interface and modulate the pumping activity of the heart. Our objective was to identify a small molecule that has the ability to stabilize the activated conformation of cTn by binding to this interface and thereby enhance cardiac muscle contraction. Hypothesis A cardiac troponin activator could potentially compensate the impaired contractile function of the heart in the treatment of systolic heart failure Methods A total of 47 new compounds were designed using existing knowledge of calcium sensitizers. The compounds were screened against a recombinantly purified cNTnC‐cTnI chimeric construct, named gChimera, using nuclear magnetic resonance and fluorescence spectroscopy. We identified a novel small molecule cardiac troponin activator, RPI‐194, and measured its binding affinity to recombinantly purified cNTnC and gChimera. We also investigated the activity of RPI‐194 in rat skinned cardiac muscle trabeculae, isolated mouse cardiomyocytes, and isolated working mouse hearts. RPI‐194 activity was also tested in skinned skeletal muscle fibers from rats as both cardiac and slow skeletal muscle share the same isoform of cTnC. Results We demonstrated that a small molecule troponin activator RPI‐194 binds to gChimera with a K D of 12‐24 µM and is able to stabilize activated cTn. It also slowed down the rate of calcium release from reconstituted cTn. RPI‐194 showed increasing calcium sensitivity of isometric contraction in skinned cardiac muscle trabeculae, as well as in slow and fast skeletal muscle fibers, suggesting it to be a calcium sensitizer, with cross‐reactivity within striated muscle. Contrarily, RPI‐194 was unable to slow down calcium release from isolated cNTnC. It also reduced the velocity of unloaded shortening in skeletal muscle fibers, suggesting that it slows the rate of actin‐myosin cross‐bridging, although no effect on myosin ATPase activity was found. RPI‐194 decreased the velocity and amplitude of contractions in isolated cardiomyocytes but contractility was preserved in isolated working hearts. Conclusions RPI‐194 is a small molecule troponin activator that acts as a calcium sensitizer in striated muscle. Because of the isoform sharing of cTnC in cardiac and slow skeletal muscle, it is nearly impossible to develop a cardiac‐specific troponin activator. Whole animal model studies are needed to determine the impact of RPI‐194 on the different striated muscle types and whether this would be acceptable for therapeutic intervention.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.002 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".