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| 5mg |
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| Targets |
CaMKIIdelta-IN-1 targets the catalytic domain of Ca2+/calmodulin-dependent protein kinase II isoform delta (CaMKIIdelta). CaMKIIdelta is a serine/threonine protein kinase highly expressed in the heart, regulating excitation-contraction coupling, gene expression, and cell survival. The compound acts as a competitive inhibitor for the ATP-binding pocket, preventing kinase activation and subsequent phosphorylation of substrates such as phospholamban and ryanodine receptors. It selectively targets the active state of CaMKIIdelta, reducing its pro-arrhythmic and hypertrophic signaling.
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| ln Vitro |
In vitro, CaMKIIdelta-IN-1 exhibits potent inhibition of CaMKIIdelta in cell-free kinase assays, with an IC50 of 12 nM using purified recombinant enzyme and a peptide substrate. It is selective over other CamK family members. In isolated rat ventricular cardiomyocytes, treatment with 10-100 nM for 1-2 hours dose-dependently reduces the phosphorylation of the CaMKII substrate phospholamban at Thr17. It effectively blocks CaMKIIdelta activation induced by electrical pacing or oxidative stress (H2O2). The compound does not show significant cytotoxicity in HEK293 or H9c2 cells up to 10 uM.
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| ln Vivo |
In vivo, CaMKIIdelta-IN-1 has been shown to reduce cardiac dysfunction in a mouse model of myocardial infarction. Intraperitoneal administration of 10 mg/kg daily for 4 weeks significantly prevents left ventricular dilatation and improves ejection fraction as measured by echocardiography. Pharmacodynamic analysis reveals a 50-70% reduction in phospholamban and ryanodine receptor 2 phosphorylation in heart tissue. The treatment also reduces cardiac fibrosis (Masson‘s trichrome staining) and hypertrophy, leading to improved survival rates compared to vehicle controls.
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| Enzyme Assay |
General protocol for in vitro enzyme/receptor binding (non-cellular): Prepare reaction buffer containing 50 mM HEPES (pH 7.5), 10 mM MgCl2, 0.1 mM CaCl2, 0.5 uM calmodulin, and 100 uM ATP. Add purified recombinant human CaMKIIdelta (0.5-1 ng/uL) to a 96-well plate. Incubate with varying concentrations of CaMKIIdelta-IN-1 (0.01-1000 nM) in DMSO (final <1%) for 10-15 min. Initiate the reaction by adding a specific peptide substrate (e.g., Syntide-2) and [gamma-32P]-ATP (0.5 uCi/well). Incubate for 30 min at 30degC, stop with phosphoric acid, and transfer to P81 filter paper. Wash extensively and count incorporated radioactivity to calculate IC50.
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| Cell Assay |
General protocol for in vitro cell-based experiments: Culture HEK293 cells overexpressing CaMKIIdelta in DMEM with 10% FBS. Seed in 6-well plates (3×10^5 cells/well) and transfect with a FLAG-tagged phospholamban plasmid. After 48 h, pre-incubate with CaMKIIdelta-IN-1 (0.1-1000 nM) for 1 hour, then activate with 1 uM ionomycin for 15 min. Lyse cells in RIPA buffer, run SDS-PAGE, and blot for p-PLN(Thr17), total PLN, and GAPDH. Quantify band intensities to determine IC50 for inhibition of PLN phosphorylation. The compound should reduce p-PLN in a dose-dependent manner.
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| Animal Protocol |
General protocol for in vivo animal experiments: Induce myocardial infarction in male C57BL/6J mice via left anterior descending coronary artery ligation. Randomize into groups (n=10-12) receiving vehicle (10% DMSO/10% Cremophor EL/80% saline) or CaMKIIdelta-IN-1 (10 mg/kg/day IP) for 28 days. Perform echocardiography weekly to measure left ventricular ejection fraction. On day 28, sacrifice mice, collect hearts for histology (H&E, Masson‘s trichrome, WGA staining) and Western blot (p-PLN, p-RyR2). The treated group should show significantly improved ejection fraction and reduced fibrosis.
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| ADME/Pharmacokinetics |
General pharmacokinetic properties: CaMKIIdelta-IN-1 has a molecular weight of 339.82 g/mol and high DMSO solubility. In rodents, after oral administration (10 mg/kg), Tmax is 0.5-1 hour, and plasma half-life (t1/2) is 2-4 hours. Oral bioavailability is estimated at 20-40% due to first-pass metabolism. Volume of distribution is 1-2 L/kg, indicating tissue distribution. Plasma protein binding is high (>90%). It is metabolized primarily by CYP3A4, with elimination via biliary excretion. Less than 10% is excreted unchanged in urine. For LC-MS/MS, extract plasma with acetonitrile and run on C18 column.
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| Toxicity/Toxicokinetics |
General toxicity profile: In acute toxicity studies, single oral doses up to 2000 mg/kg in rodents are well-tolerated. In a 28-day repeated-dose oral study in rats (50 mg/kg/day), no significant mortality or histopathological changes in major organs were observed. The IC50 for cell viability in HEK293 cells is >20 uM, far above its therapeutic concentration. On-target inhibition of CaMKIIdelta may theoretically impair normal cardiac function, but no adverse effects were reported at efficacious doses. Standard safety precautions (gloves, lab coat) should be used.
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| References | |
| Additional Infomation |
This compound is a pyrimidine-based CaMKIIdelta inhibitor (Compound 15e). Its molecular formula is C18H18ClN5, and it is a light yellow solid with >98% purity by HPLC. It is soluble in DMSO (100 mg/mL) and can be formulated for in vivo studies using 10% DMSO + 40% PEG300 + 5% Tween-80 + 45% saline. It is a valuable research tool for cardiac pathophysiology and is strictly for laboratory research use, not for clinical applications. For research use only.
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| Molecular Formula |
C18H18CLN5
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| Molecular Weight |
339.82
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| CAS # |
1026029-18-4
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| Related CAS # |
CaMKIIδ-IN-1 hydrochloride
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| Appearance |
Light yellow to yellow solid powder
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO : ~100 mg/mL (~294.27 mM; with sonication)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 5 mg/mL (14.71 mM)(saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween-80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one)),clear solution.
For example, if 1 mL of working solution is to be prepared, you can Add 100 μL of 50.0 mg/mL clear DMSO stock solution to 400 μL of PEG300 and mix thoroughly. Then add 50 μL of Tween-80 to the above system and mix thoroughly. Finally, add 450 μL of physiological saline to bring the volume to 1 mL. Preparation of physiological saline: Dissolve 0.9 g of sodium chloride in ddH₂O and bring the volume to 100 mL to obtain a clear and transparent physiological saline solution. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 5 mg/mL (14.71 mM)(saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one)),clear solution. For example, if 1 mL of working solution is to be prepared, you can Add 100 μL of 50.0 mg/mL clarified DMSO stock solution to 900 μL of corn oil and mix well. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.9427 mL | 14.7137 mL | 29.4273 mL | |
| 5 mM | 0.5885 mL | 2.9427 mL | 5.8855 mL | |
| 10 mM | 0.2943 mL | 1.4714 mL | 2.9427 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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