Technical Data

Molecular Weight 351.45 Storage powder


in solvent
CAS No. 872573-93-8 Synonyms N/A
Solubility (25°C) * In vitro DMSO 13 mg/mL warmed (36.98 mM)
Water Insoluble
Ethanol Insoluble
In vivo
* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
Chemical Name (Z)-5-(quinolin-6-ylmethylene)-2-(thiophen-2-ylmethylamino)thiazol-4(5H)-one

Biological Activity

Description RO-3306 is an ATP-competitive, and selective CDK1 inhibitor with Ki of 20 nM, >15-fold selectivity against a diverse panel of human kinases.
CDK1 [1]
(Cell-free assay)
PKCδ [1]
(Cell-free assay)
SGK [1]
(Cell-free assay)
ERK [1]
(Cell-free assay)
20 nM(Ki) 318 nM(Ki) 497 nM(Ki) 1980 nM(Ki)
In vitro RO-3306 inhibits CDK1/cyclin B1, CDK1/cyclin A, CDK2/cyclin E, and CDK4/cyclin D activity with Ki of 35 nM, 110 nM, 340 nM, and over 2000 nM, respectively. Treatment of HCT116, SW480, and HeLa cells with RO-3306 for 20 h leads to a complete block of the cell cycle in the G2/M phase. The proliferation of both HCT116 and SW480 is effectively blocked by RO-3306. RO-3306 appears to be more proapoptotic in cancer cells (HCT116 and SW480) than nontumorigenic cells (MCF 10A and MCF 12A).[1] RO-3306 effectively arrests oocyte maturation at a concentration of 10 μM.[2]
In vivo

Protocol (Only for Reference)

Kinase Assay: [1]

CDK assay The activity of CDK1 cyclin B1, CDK1 cyclin A, CDK2 cyclin E, and CDK4 cyclin D is measured by a homogeneous time-resolved fluorescence assay in a 96-well format. The assay buffer contained 25 mM Hepes, 6.25 mM MgCl2, 0.003% Tween 20, 0.3 mg/mL BSA, 1.5 mM DTT, and ATP as follows: 162 μM (CDK1), 90 mM (CDK2), or 135 μM (CDK4). CDK1 and CDK2 buffer contained 10 mM MgCl2. Test compounds are diluted in assay buffer to 3-fold their final concentration in 20 μL, and the reaction is started by the addition of a 40 μL assay buffer containing the pRB substrate (0.185μM). The plates are incubated at 37°C for 30 min with constant agitation, and the reaction is terminated by the addition of 15 μL of 1.6μM anti-phospho pRB antibody (Ser-780) in 25 mM Hepes, 24 mM EDTA, and 0.2 mg/mL BSA. After an additional 30 min of incubation with shaking, 15μL of 3nM Lance-Eu-W1024-labeledanti-rabbitIgG and 60 nM Alophycocyanin-conjugated anti-His-6 antibody in 25 mM Hepes, and 0.5 mg/mL BSA is added and incubated for 1 h. The plates are read in the Victor-V multi- label reader at excitation 340 nm and emission 615 nm and 665 nm. The IC50 values are calculated from the readings at 665 nm and normalized for Europium readings at 615 nm. Ki values are calculated according to the equation: Ki= IC50/(1 + S/Km ), where S is the ATP concentration in the assay and Km is the Michaelis-Menten constant for ATP. The inhibitory activity against the panel of kinases is determined by the IMAP assay technology.

Cell Assay: [3]

Cell lines MDA-MB-231 cell line
Concentrations 20 μM
Incubation Time 72 h
Method Log phases cells (25,000) are seeed in 96-well plates and incubated in a 37℃ incubator with CO2, After 24 h, different concentrations of RO-3306 are administered to determine the drug concentrations required to achieve a 50% growth inhibition (IC50). MTT (20 μL, 5mg/mL stock solution in saline) is added to each well and the cells are incubated for 4 h. Supernatants are removed and formazan crystals from viable cells are solubilized with 200 μL anhydrous DMSO. The absorbance is detected with a 550 model microplate reader at the 565 nm wavelength.


Customer Product Validation

Data from [Data independently produced by , , J Cell Biol, 2015, 209(2): 221-34 ]

HeLa cells were untreated or treated with either colcemid (10 μg/ml for 16 h), RO-3306 (9 μM for 16 h), or both colcemid and RO-3306, as indicated. Cells were then lysed, and proteins were detected by Western analysis.

Data from [Data independently produced by , , Arthritis Rheumatol, 2016, 68(5):1222-32.]

A, Expression of IFIT3 in HeLa cells pretreated with 5 μM RO-3306 for 12 hours and then stimulated with type I IFN (1,000 units/ml) for 8 hours. The cells were collected and total RNA was extracted for the detection of IFIT3 mRNA. Results are the relative expression levels of IFIT3 mRNA normalized to endogenous GAPDH mRNA levels. B, Expression of IFI27 in HeLa cells measured in the same samples as in A.

Ro-3306 has been referenced in 4 publications.



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Chemical Structure

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