BI 2536

Alias: BI2536; BI-2536; BI 2536

Cat No.:V1573 Purity: ≥98%

COA Product Data Instructions for use SDS

BI2536 (BI-2536; BI 2536) is a novel PLK1/BRD4 dual inhibitor with potential antitumor activity.

BI 2536 Chemical Structure CAS No.: 755038-02-9
Product category: PLK

This product is for research use only, not for human use. We do not sell to patients.

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Citations by Top Journals: Nature, Cell, Science, etc.

Top Publications Citing lnvivochem Products

Nature 2021, 597(7874):119-125.

Cell 2020,183:1202-1218.e25.

Science Adv 2022: 8, eabm9427.

Nature 597, 119–125 (2021)

Cell Stem Cell 2020,26(6):845-861.e12.

Science Trans Med 2023,15(701):eadd7872.

STTT 2023,8(1):91.

Cell Reports 2023,42(4):112313

Science Trans Med 2023, 15: eadd7872.

Cancer Cell 2021,39(4):529-547.e7.

Cancer Discov 2022,12(4):1106-1127.

Immunity 2023,56(3):620-634.e11.

J Am Chem Soc 2022,144:21831-21836.

Cell 2020,183:1202-1218.e25.

Science Adv 2022:8,eabm9427.

Nature 2021,597(7874):119-125.

Cell 2020,183:1202-1218.e25.

PNAS Nexus 2022,1(3):pgac063.

ACS Nano 2023,17(10):9110-9125.

Biomaterial 2023 Sep16:302:122333.

Purity & Quality Control Documentation

Current batch：

Purity: ≥98%

COA

MSDS

NMR

Instructions

Product Description
Bioactivity & Protocols
Physicochemical Properties
Solubility Data
Calculators
Clinical Trial Information
Biological Data

Product Description

BI2536 (BI-2536; BI 2536) is a novel PLK1/BRD4 dual inhibitor with potential antitumor activity. In a cell-free assay, it inhibits PLK1/BRD4 with IC50s of 0.83 and 25 nM, respectively. The serine/threonine protein kinase Plk1 is an essential regulator of several processes essential to cell division and mitosis.

Biological Activity I Assay Protocols (From Reference)

Targets

PLK1 (IC50 = 0.83 nM); Plk2/Snk (IC50 = 3.5 nM); Plk3/Fnk (IC50 = 9 nM); BRD4 (IC50 = 25 nM)

ln Vitro

BI 2536 inhibits Plk2 and Plk3 functions to a marginal degree, with IC50 values of 3.5 nM and 9.0 nM, in that order. A range of mitotic processes known to depend on Plk1 are blocked in HeLa cells by BI 2536 treatment, ranging from 10 to 100 nM. These effects include induction of monopolar spindles, inhibition of cohesin release from chromosome arms, and blocking of γ-tubulin recruitment and phosphorylation of Apc6 at mitotic centrosomes. Treatment with BI 2536 results in the arrest of HeLa cells in G2/M, followed by a concentration-dependent accumulation of cleaved PARP p85 fragments and a sub-G1 DNA peak, which is suggestive of DNA breakage and apoptosis. While BI 2536 prevents the growth of exponentially growing hTERT-RPE1, human umbilical vein endothelial cells (HUVECs), and normal rat kidney (NRK) cells with EC50 of 12-31 nM, it also inhibits the growth of a panel of 32 human cancer cell lines with EC50 of 2-25 nM. With EC50 values ranging from 1.4 to 5.6 nM, BI 2536's inhibition of Plk1 suppresses the growth and viability of anaplastic thyroid carcinoma (ATC) cells, including CAL62, OCUT-1, SW1736, 8505C, and ACT-1.

ln Vivo

BI 2536, administered intravenously (IV) once or twice a week, inhibits cell proliferation by inducing a mitotic arrest, which in turn induces tumor-cell death in a variety of xenograft models with acceptable tolerability. The growth of HCT 116 xenografts with T/C of 15% and 0.3%, respectively, is markedly inhibited by the administration of BI 2536 at 50 mg/kg once or twice a week. Additionally, BxPC-3 and A549 models treated twice a week with BI 2536 show excellent tumor growth with T/C of 5% and 14%, respectively.

Enzyme Assay

Recombinant human Plk1 (residues 1–603) is purified by affinity chromatography using glutathione-agarose after being expressed as an N-terminal, GST-tagged fusion protein using a baculoviral expression system. When conducting Plk1 enzyme activity assays, serially diluted BI 2536 is present along with 20 ng of recombinant kinase and 10 μg of casein from cow's milk used as the substrate. A final volume of 60 μL is used for kinase reactions, which are carried out for 45 minutes at 30 °C (15 mM MgCl2, 25 mM MOPS [pH 7.0], 1 mM DTT, 1% DMSO, 7.5 mM ATP, 0.3 μCi γ-33P-ATP). A final 125 μL of ice-cold 5% TCA is added to stop the reaction. Following the transfer of precipitates to Multi-Screen mixed ester cellulose filter plates, the plates undergo a 1% TCA wash and radiometric quantification. The IC50 value is computed using the dose-response curve.

Cell Assay

In assays for cell proliferation, different concentrations of BI 2536 (10 nM-1 μM) are added and incubated for 72 hours. The growth of the cells is measured by measuring the Alamar Blue dye conversion in a fluorescence spectrophotometer. From the dose-response curve fit, effective concentrations (EC50) at which cellular growth is 50% inhibited are extrapolated[3].

Animal Protocol

Mice: Subcutaneous injection of 2×10⁶, 1×10⁶, or 1×10⁷ cells into each mouse's flank results in the grafting of HCT 116 colon-carcinoma, NCI-H460, or A549 lung-carcinoma cells into female BomTac:NMRI-Foxn1nu mice. Animals are paired into groups of ten mice each for treatment and control once tumors have grown to a volume of about 50 mm³. Treatment is not started in regression experiments until the mean tumor volume reaches 500 mm³. The recommended dosage and schedule for intravenous injection of BI 2536 are administered into the tail vein. Ten milliliters for every kilogram of body weight is the dosage. Three times a week, calipers are used to determine tumor volumes. The following formula is used to convert the results to tumor volume (mm³): length×width²×π/6. On the same days, the mice's weight is ascertained as a measure of tolerability. In a one-sided (decreasing) exact Wilcoxon test, the treatment group and the vehicle control group are compared for statistical analysis.

References

[1]. The Small-Molecule Inhibitor BI 2536 Reveals Novel Insights into Mitotic Roles of Polo-like Kinase 1. Curr Biol. 2007 Feb 20;17(4):304-15.

[2]. BRD4 Structure-Activity Relationships of Dual PLK1 Kinase/BRD4 Bromodomain Inhibitor BI-2536. ACS Med Chem Lett. 2015 May 18;6(7):764-9.

[3]. BI 2536, a Potent and Selective Inhibitor of Polo-like Kinase 1, Inhibits Tumor Growth In Vivo. Current Biology (2007), 17(4), 316-322.

[4]. Suppression of IFN β gene transcription by inhibitors of bromodomain and extra-terminal (BET) family members. Biochem J. 2015 Jun 15;468(3):363-72.

These protocols are for reference only. InvivoChem does not independently validate these methods.

Physicochemical Properties

Molecular Formula

C28H39N7O3

Molecular Weight

521.66

Exact Mass

521.31

Elemental Analysis

C, 64.47; H, 7.54; N, 18.80; O, 9.20

CAS #

755038-02-9

Related CAS #

755038-02-9

Appearance

Off-white to yellow solid powder

SMILES

CC[C@@H]1C(=O)N(C2=CN=C(N=C2N1C3CCCC3)NC4=C(C=C(C=C4)C(=O)NC5CCN(CC5)C)OC)C

InChi Key

XQVVPGYIWAGRNI-JOCHJYFZSA-N

InChi Code

InChI=1S/C28H39N7O3/c1-5-22-27(37)34(3)23-17-29-28(32-25(23)35(22)20-8-6-7-9-20)31-21-11-10-18(16-24(21)38-4)26(36)30-19-12-14-33(2)15-13-19/h10-11,16-17,19-20,22H,5-9,12-15H2,1-4H3,(H,30,36)(H,29,31,32)/t22-/m1/s1

Chemical Name

4-[[(7R)-8-cyclopentyl-7-ethyl-5-methyl-6-oxo-7H-pteridin-2-yl]amino]-3-methoxy-N-(1-methylpiperidin-4-yl)benzamide

Synonyms

BI2536; BI-2536; BI 2536

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)

Solubility Data

Solubility (In Vitro)

DMSO: ~21 mg/mL (~40.3 mM)

Water: <1 mg/mL

Ethanol: ~100 mg/mL (~191.7 mM)

Solubility (In Vivo)

30%PEG 400+0.5%Tween 80+5%Propylene glycol: 15mg/mL

(Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	1.9170 mL	9.5848 mL	19.1696 mL
	5 mM	0.3834 mL	1.9170 mL	3.8339 mL
	10 mM	0.1917 mL	0.9585 mL	1.9170 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.

Calculator

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Molarity Calculator allows you to calculate the mass, volume, and/or concentration required for a solution, as detailed below:

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See Example

An example of molarity calculation using the molarity calculator is shown below:
What is the mass of compound required to make a 10 mM stock solution in 5 ml of DMSO given that the molecular weight of the compound is 350.26 g/mol?

Enter 350.26 in the Molecular Weight (MW) box
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The answer of 17.513 mg appears in the Mass box. In a similar way, you may calculate the volume and concentration.

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Dilution Calculator allows you to calculate how to dilute a stock solution of known concentrations. For example, you may Enter C1, C2 & V2 to calculate V1, as detailed below:

What volume of a given 10 mM stock solution is required to make 25 ml of a 25 μM solution?
Using the equation C1V1 = C2V2, where C1=10 mM, C2=25 μM, V2=25 ml and V1 is the unknown:

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The answer of 62.5 μL (0.1 ml) appears in the Volume (Start) box

Molecular formula

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Molecular Weight Calculator allows you to calculate the molar mass and elemental composition of a compound, as detailed below:

Note: Chemical formula is case sensitive: C12H18N3O4 c12h18n3o4
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Definitions of molecular mass, molecular weight, molar mass and molar weight:

Molecular mass (or molecular weight) is the mass of one molecule of a substance and is expressed in the unified atomic mass units (u). (1 u is equal to 1/12 the mass of one atom of carbon-12)
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Reconstitution Calculator allows you to calculate the volume of solvent required to reconstitute your vial.

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The answer appears in the Volume (to add to vial) box

In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal to make allowance for loss during the experiment)

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Step 2: Enter in vivo formulation (This is only a calculator, not the exact formulation for a specific product. Please contact us first if there is no in vivo formulation in the solubility section.)

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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.

Note： (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.

Clinical Trial Information

NCT Number	Recruitment	interventions	Conditions	Sponsor/Collaborators	Start Date	Phases
NCT00412880	Completed	Drug: BI 2536	Carcinoma, Small Cell	Boehringer Ingelheim	February 14, 2007	Phase 2
NCT00243087	Completed	Drug: BI 2536	Lymphoma	Boehringer Ingelheim	July 2005	Phase 1
NCT00706498	Completed	Drug: BI 2536	Prostatic Neoplasms	Boehringer Ingelheim	September 2006	Phase 2

Biological Data

Crystal structure of BI-2536 bound to BRD4(1) (PDB ID = 4OGI).ACS Med Chem Lett.2015 May 18;6(7):764-9.
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High scoring, GOLD-predicted docking mode of compound39jin BRD4(1).ACS Med Chem Lett.2015 May 18;6(7):764-9.