Monastrol is an allosteric inhibitor of the mitotic kinesin Eg5 that exhibits an antiproliferative effect against several cell lines. As a kinesin-5(KIF11) cell-permeable small molecule inhibitor, monastrol may have anticancer properties. Additionally, it exhibits no selectivity but has a stronger antiproliferative effect on MCF-7 tumor cells when compared to non-tumor HB4a cells. Centrosome duplication and cell cycle progression through the S and G2 phases are not inhibited by metronidazole. Furthermore, monastrol-induced mitotic arrest is quickly reversible. In Xenopus egg extracts, monastrol also prevents the formation of bipolar spindles. While monostrol has no effect on microtubules in interphase cells or microtubule polymerization in vitro, it stops cells in mitosis when they form monoastral spindles, which are made up of a radial array of microtubules encircled by a ring of chromosomes.

Physicochemical Properties

Molecular Formula	C14H16N2O3S
Molecular Weight	292.35
Exact Mass	292.088
Elemental Analysis	C, 57.52; H, 5.52; N, 9.58; O, 16.42; S, 10.97
CAS #	329689-23-8
Related CAS #	(S)-Monastrol;254753-54-3
PubChem CID	2987927
Appearance	White to off-white solid powder
Density	1.3±0.1 g/cm3
Boiling Point	441.3±55.0 °C at 760 mmHg
Flash Point	220.7±31.5 °C
Vapour Pressure	0.0±1.1 mmHg at 25°C
Index of Refraction	1.648
LogP	2.18
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	4
Heavy Atom Count	20
Complexity	436
Defined Atom Stereocenter Count	0
SMILES	O=C(C1=C(C)NC(NC1C2=CC=CC(O)=C2)=S)OCC
InChi Key	LOBCDGHHHHGHFA-UHFFFAOYSA-N
InChi Code	InChI=1S/C14H16N2O3S/c1-3-19-13(18)11-8(2)15-14(20)16-12(11)9-5-4-6-10(17)7-9/h4-7,12,17H,3H2,1-2H3,(H2,15,16,20)
Chemical Name	ethyl 4-(3-hydroxyphenyl)-6-methyl-2-sulfanylidene-3,4-dihydro-1H-pyrimidine-5-carboxylate
Synonyms	(±)-Monastrol; Monastrol
HS Tariff Code	2934.99.9001
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)

Biological Activity

Targets	Eg5 (IC50 = 14 μM)
ln Vitro	Monastrol does not prevent centrosome duplication or cell cycle progression through the S and G2 phases. Furthermore quickly reversible is the mitotic arrest brought on by monastrol. Monastrol also prevents the formation of bipolar spindles in extracts from Xenopus eggs. Microtubules in interphase cells and microtubule polymerization in vitro are unaffected by monoastrrol, which stops cells in mitosis and creates monoastral spindles made of a radial array of microtubules encircled by a ring of chromosomes[1]. Monastrol exposure for a few hours increases the number and rate of axon growth in cultured sympathetic neurons. After more time, the axons' total lengths become indistinguishable from controls. A comparable transient rise in axonal growth rate is observed in sensory neurons. Nonetheless, extended exposure causes shorter axons, indicating that sensory neurons may be more vulnerable to the drug's harmful effects. Even so, the cultures' general health is still far stronger than that of cultures that were given the medication taxol, which is frequently used in anti-cancer therapy[2]. Monastrol causes apoptosis and mitotic arrest in HeLa cells by activating the spindle checkpoint[3].
ln Vivo
Enzyme Assay	Monastrol is an allosteric mitotic kinesin Eg5 inhibitor that has antiproliferative properties against a variety of cell lines. As a kinesin-5(KIF11) cell-permeable small molecule inhibitor, monastrol may have anticancer properties.
Cell Assay	BS-C-1 cells that are growing exponentially are cultivated for 16 hours in normal growth medium with 2 mM thymidine in order to perform the double thymidine arrest. Following this, the cells are placed into regular growth medium and given a 9-hour dose of 24 μM deoxycytidine. For 16 hours, the cells were kept in a serum-free medium with 2 millimolar thymidine as part of the second thymidine block. The cells are then released into a regular growth medium that contains 0.1% DMSO or 100 μM monastrol in addition to 24 μM deoxycytidine. In order to evaluate the reversibility of the monastrol and nocodazole treatment effect, plated BS-C-1 cells on coverslips are treated for 4 hours in normal growth medium supplemented with either 100 μM monastrol or 2 μM nocodazole, and subsequently released back into normal medium. Coverslips are processed for immunofluorescence and the cells in interphase or mitosis are counted and sorted at various time intervals.
Animal Protocol
References	[1]. J Cell Biol . 2000 Sep 4;150(5):975-88. [2]. Cell Motil Cytoskeleton . 2004 May;58(1):10-6. [3]. Mol Cancer Ther . 2006 Oct;5(10):2580-91. [4]. Science . 1999 Oct 29;286(5441):971-4.
Additional Infomation	Monastrol is a racemate comprising equimolar amounts of R- and S-monastrol. It has a role as an antineoplastic agent, an EC 3.5.1.5 (urease) inhibitor, an antileishmanial agent and an antimitotic. It contains a (R)-monastrol and a (S)-monastrol.

Solubility Data

Solubility (In Vitro)

DMSO: ~58 mg/mL (~198.4 mM) Water:<1 mg/mL Ethanol: ~58 mg/mL (~198.4 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (8.55 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 (8.55 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 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. Solubility in Formulation 3: ≥ 2.5 mg/mL (8.55 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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.4206 mL	17.1028 mL	34.2056 mL
	5 mM	0.6841 mL	3.4206 mL	6.8411 mL
	10 mM	0.3421 mL	1.7103 mL	3.4206 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.