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Purity: ≥98%
Olomoucine, a purine derivative, is a novel and potent CDK (cyclin-dependent kinases) inhibitor that competes for the ATP binding site of the kinase and induces G arrest.. Olomoucine has been demonstrated to reversibly stop cycling Arabidopsis cells in late G1 and G2, as well as differentiated Petunia cells that have been induced to divide at the G1 phase.
| Targets |
cdk2-cyclin A (IC50 = 7 μM); cdk2-cyclin E (IC50 = 7 μM); cdk5-p35 (IC50 = 25 μM)
p34cdc2/cyclin B kinase (IC₅₀: 7 µM) p33cdc2/cyclin A kinase (IC₅₀: 7 µM) p33cdc2/cyclin E kinase (IC₅₀: 7 µM) p33cdk5/p35 kinase (IC₅₀: 3 µM) ERK1/MAP kinase (and its starfish homologue p44mpk) (IC₅₀: 25 µM) ASK-γ (plant homologue of GSK-3) (IC₅₀: 130 µM) |
|---|---|
| ln Vitro |
Olomoucine suppresses the kinases CDK2 and CDC2, with IC50 values of 7 μM for CDC2/cyclin B, 7 μM for CDK2/cyclin A, 7 μM for CDK2/cyclin E, 3 μm for CDK5/p35, and 25 μM for ERK1/p44 MAPK, respectively[1].
Olomoucine is a non-competitive inhibitor of histone H and a competitive inhibitor of ATP at concentrations of 0, 5, 10, 15, and 25 μM[1]. Olomoucine (0-1000 μM) causes a Gl arrest akin to interleukin-2 deprivation and suppresses DNA synthesis in interleukin-2-stimulated T lymphocytes (CTLL-2 cells)[2]. Olomoucine (0-100 μM) inhibits the MB65 cell line's Gl/S transition in non-small cell lung cancer[2]. Olomoucine (0-150 μM) inhibits the Rdditapes oocytes' prophase/metaphase transition[2]. Olomoucine has been shown to inhibit the survival of tumor cells, with IC50s of 32.35 μM for dog melanoma, 42.15 μM for mouse B16 melanoma, and 82.30 μM for human melanoma, respectively[3]. Olomoucine specifically inhibits the kinase activity of cyclin-dependent kinases cdc2, cdk2, cdk5, and ERK1/MAP kinase at micromolar concentrations, while showing little to no effect on cdk4, cdk6, and many other protein kinases (including PKC isoforms, cAMP-dependent kinase, cGMP-dependent kinase, tyrosine kinases). It acts as a competitive inhibitor for ATP binding and a non-competitive inhibitor for the protein substrate histone H1. Olomoucine inhibits in vitro M-phase-promoting factor (MPF) activity in metaphase-arrested Xenopus egg extracts, preventing chromosome decondensation and nuclear envelope assembly at concentrations of 10 µM and above. Olomoucine inhibits in vitro DNA synthesis in Xenopus interphase egg extracts, with significant inhibition observed at 10-15 µM. Olomoucine inhibits the replication licensing factor activity in Xenopus egg extracts, an essential factor ensuring DNA is replicated only once per cell cycle, with significant inhibition at 10-15 µM. Among 81 purine derivatives tested, only those substituted at C2, N6, and N9 positions (like olomoucine) showed strong inhibitory effects on p34cdc2/cyclin B kinase. Substitution at the N7 position dramatically reduces activity. |
| ln Vivo |
Olomoucine (8 mg/kg; i.v.; once daily; 7 d) causes tumor cells to undergo apoptosis without causing any side effects on the third day following treatment[3].
It has been discovered that, in comparison to single dosage administration, cassette dosing underestimates the Cmax and overestimates the AUC[4]. Olomoucine inhibits the G2/M transition in starfish oocytes in vivo, induced by the hormone 1-methyladenine, with an IC₅₀ of 30 µM. Olomoucine reduces the in vivo tyrosine dephosphorylation of p34cdc2 in starfish oocytes, a step normally catalyzed by the cdc25 phosphatase and preceding cdc2 kinase activation at the G2/M transition. |
| Enzyme Assay |
Kinase activities were assayed at 30°C in triplicate in a buffer containing sn-glycerol 2-phosphate, p-nitrophenyl phosphate, Mops, EGTA, MgCl₂, dithiothreitol, sodium vanadate, NaF, phenylphosphate, and protease inhibitors (leupeptin, aprotinin, soybean trypsin inhibitor, benzamidine).
For p34cdc2/cyclin B kinase assay, the enzyme purified from starfish oocytes was incubated with 1 mg/ml histone H1 and 15 µM [γ-³²P]ATP in a final volume of 30 µl for 10 min. The reaction was stopped by spotting aliquots onto phosphocellulose paper, followed by washing in phosphoric acid and scintillation counting. Kinetic analysis for inhibition mechanism involved running initial-rate experiments with non-saturating substrate concentrations (ATP or histone H1). Double-reciprocal plots were used to determine the type of inhibition and apparent inhibition constants (Kᵢ). Assays for other kinases (e.g., cdk2/cyclin A/E, cdk5/p35, MAPK, PKC isoforms, tyrosine kinases) followed similar principles with their respective specific substrates (histone H1, myelin basic protein, casein, retinoblastoma protein, angiotensin II, etc.), appropriate co-factors, and detection methods (phosphocellulose filter binding, SDS-PAGE/autoradiography, densitometry). |
| Animal Protocol |
Dog with spontaneous melanoma (oral and maxillofacial tumors)[3]
8 mg/kg Intravenous injection; once daily; for 7 days Starfish (Marthasterias glacialis) oocyte maturation assay: G2-arrested oocytes were treated for 15 minutes with increasing concentrations of Olomoucine (dissolved in DMSO, final concentration not specified for vehicle) prior to addition of 1 µM 1-methyladenine to induce maturation. Germinal vesicle breakdown was scored after 30 minutes. For analysis of p34cdc2 tyrosine phosphorylation, oocytes were treated or not treated with 100 µM Olomoucine for 15 minutes prior to 1-methyladenine addition. At various times, extracts were prepared and p34cdc2 was affinity-purified for subsequent Western blot analysis with anti-phosphotyrosine antibodies. |
| References |
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| Additional Infomation |
Olomoucin is a 9H-purine with (2-hydroxyethyl)nitroso, benzylnitroso, and methyl groups substituted at positions 2, 6, and 9, respectively. It is a cyclin-dependent kinase inhibitor, belonging to EC 2.7.11.22 (cyclin-dependent kinase) inhibitors. It belongs to the 2,6-diaminopurine and ethanolamine classes. Olomoucin [2-(2-hydroxyethylamino)-6-benzylamino-9-methylpurine] is a purine derivative discovered during testing of compounds associated with the nonspecific kinase inhibitors N⁶-dimethylaminopurine and N⁶-(Δ²-isopentenyl)adenine. Its unique selectivity divides cyclin-dependent kinases into two subfamilies: cdc2/cdk2/cdk5 (sensitive) and cdk4/cdk6 (low-sensitivity). This specificity is thought to be due to differences in ATP-binding pockets. This study suggests that olomoxin may serve as a lead compound for designing more effective and selective CDK inhibitors, which could potentially be used as antimitotic and antitumor drugs. Furthermore, due to its inhibitory spectrum (CDK5/p35, ERK1, GSK-3), olomoxin may be relevant to Alzheimer's disease research, as the hyperphosphorylation of tau protein by these kinases is associated with Alzheimer's disease. This compound was initially described as an inhibitor of radish cotyledon cytokinin 7-glucosyltransferase.
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| Molecular Formula |
C15H18N6O
|
|---|---|
| Molecular Weight |
298.34302
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| Exact Mass |
298.154
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| Elemental Analysis |
C, 60.39; H, 6.08; N, 28.17; O, 5.36
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| CAS # |
101622-51-9
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| Related CAS # |
101622-51-9
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| PubChem CID |
4592
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| Appearance |
White to off-white solid powder
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| Density |
1.4±0.1 g/cm3
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| Boiling Point |
579.6±60.0 °C at 760 mmHg
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| Melting Point |
120-130 °C
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| Flash Point |
304.3±32.9 °C
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| Vapour Pressure |
0.0±1.7 mmHg at 25°C
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| Index of Refraction |
1.691
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| LogP |
-0.08
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
6
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| Rotatable Bond Count |
6
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| Heavy Atom Count |
22
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| Complexity |
338
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| Defined Atom Stereocenter Count |
0
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| SMILES |
CN1C=NC2=C(NCC3=CC=CC=C3)NC(=NCCO)N=C21
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| InChi Key |
GTVPOLSIJWJJNY-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C15H18N6O/c1-21-10-18-12-13(17-9-11-5-3-2-4-6-11)19-15(16-7-8-22)20-14(12)21/h2-6,10,22H,7-9H2,1H3,(H2,16,17,19,20)
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| Chemical Name |
2-[[6-(benzylamino)-9-methylpurin-2-yl]amino]ethanol
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| Synonyms |
Olomoucine
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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: ~66.7 mg/mL (~223.5 mM)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (8.38 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 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 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: 2.5 mg/mL (8.38 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (8.38 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.3519 mL | 16.7594 mL | 33.5188 mL | |
| 5 mM | 0.6704 mL | 3.3519 mL | 6.7038 mL | |
| 10 mM | 0.3352 mL | 1.6759 mL | 3.3519 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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