BI 2536

Catalog No.S1109

BI2536 is a potent Plk1 inhibitor with IC50 of 0.83 nM in a cell-free assay. It shows 4- and 11-fold greater selectivity against Plk2 and Plk3. Phase 2.

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BI 2536 Chemical Structure

BI 2536 Chemical Structure
Molecular Weight: 521.66

Validation & Quality Control

Product Use Citation(50)

Customer Product Validation(10)

Quality Control & MSDS

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Product Information

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  • Research Area
  • Inhibition Profile
  • BI 2536 Mechanism

Product Description

Biological Activity

Description BI2536 is a potent Plk1 inhibitor with IC50 of 0.83 nM in a cell-free assay. It shows 4- and 11-fold greater selectivity against Plk2 and Plk3. Phase 2.
Targets PLK1 [1]
(Cell-free assay)
PLK2 [1]
(Cell-free assay)
PLK3 [1]
(Cell-free assay)
PI3Kα [1]
(Cell-free assay)

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IC50 0.83 nM 3.5 nM 9.0 nM 2.407 μM
In vitro BI 2536 blocks the activities of Plk2 and Plk3 to a slightly lesser extent with IC50 of 3.5 nM and 9.0 nM, respectively. In HeLa cells, BI 2536 treatment ranging from 10-100 nM leads to the blocking of the recruitment of γ-tubulin and phosphorylation of Apc6 at mitotic centrosomes, inhibition of cohesin release from chromosome arms, induction of monopolar spindles, as well as a range of other mitotic processes that are known to depend on Plk1. BI 2536 treatment leads to the HeLa cells arrested in G2/M, subsequently a sub-G1 DNA peak indicative of DNA breakdown and apoptosis, and accumulated cleaved PARP p85 fragments in a concentration-dependent manner. BI 2536 inhibits the growth of a panel of 32 human cancer cell lines with EC50 of 2-25 nM, while blocking the proliferation of exponentially growing hTERT-RPE1, human umbilical vein endothelial cells (HUVECs), and normal rat kidney (NRK) cells with EC50 of 12-31 nM. [1] Plk1 inhibition by BI 2536 reduces the growth and viability of anaplastic thyroid carcinoma (ATC) cells such as CAL62, OCUT-1, SW1736, 8505C, and ACT-1 with EC50 values of 1.4-5.6 nM. [2]
Cell Data
Cell LinesAssay TypeConcentrationIncubation TimeFormulationActivity DescriptionPMID
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... Click to View More Cell Line Experimental Data

In vivo BI 2536 given i.v. once or twice per week is highly efficacious in diverse xenograft models with acceptable tolerability by inhibiting cell proliferation through a mitotic arrest, and subsequently induction of tumor-cell death. Administration of BI 2536 at 50 mg/kg once or twice per week significantly inhibits growth of HCT 116 xenografts with T/C of 15% and 0.3%, respectively. BI 2536 treatment twice-weekly also leads to excellent tumor-growth in BxPC-3 and A549 models with T/C of 5% and 14%, respectively. [1]
Features The first potent and selective Plk1 inhibitor that induces all hallmarks of Plk1 inhibition.

Protocol(Only for Reference)

Kinase Assay: [1]

Plk1 in vitro kinase assay Recombinant human Plk1 (residues 1-603) is expressed as N-terminal, GST-tagged fusion protein with a baculoviral expression system and purified by affinity chromatography with Glutathione-agarose. Enzyme activity assays for Plk1 are performed in the presence of serially diluted BI 2536 with 20 ng of recombinant kinase and 10 μg casein from bovine milk as the substrate. Kinase reactions are performed in a final volume of 60 μL 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). Reactions are terminated by the addition of 125 μL of ice-cold 5% TCA. After transfer of the precipitates to Multi-Screen mixed ester cellulose filter plates, plates are washed with 1% TCA and quantified radiometrically. Dose-response curve is used for calculating IC50 value.

Cell Assay: [1]

Cell lines HeLa, A43, SKOV-3, HT-29, K562, A549, Saos-2, MCF7, HCT116, COLO 205, Hep G2, Raji and PC-3 cells, etc.
Concentrations Dissolved in DMSO, final concentrations ~1 μM
Incubation Time 24 and 72 hours
Method Cells are exposed to various concentrations of BI 2536 for 24, and 72 hours. Cell growth is assessed by the measurement of Alamar Blue dye conversion in a fluorescence spectrophotometer. For determining the DNA content of the cultures, cell suspensions are fixed in 80% ethanol, treated for 5 minutes with 0.25% Triton X-100 in PBS, and incubated with 0.1% RNase and 10 μg/mL propidium iodide (PI) in PBS for 20 minutes at RT. Cell-cycle profiles are determined by flow cytometric analysis.

Animal Study: [1]

Animal Models Female BomTac:NMRI-Foxn1nu mice injected subcutaneously with HCT 116, NCI-H460, or A549 cells
Formulation Formulated in hydrochloric acid (0.1 N), and diluted with 0.9% NaCl
Dosages ~50 mg/kg
Administration Injection i.v. once or twice per week

Conversion of different model animals based on BSA (Value based on data from FDA Draft Guidelines)

SpeciesMouseRatRabbitGuinea pigHamsterDog
Weight (kg)0.020.151.80.40.0810
Body Surface Area (m2)0.0070.0250.150.050.020.5
Km factor36128520
Animal A (mg/kg) = Animal B (mg/kg) multiplied by  Animal B Km
Animal A Km

For example, to modify the dose of resveratrol used for a mouse (22.4 mg/kg) to a dose based on the BSA for a rat, multiply 22.4 mg/kg by the Km factor for a mouse and then divide by the Km factor for a rat. This calculation results in a rat equivalent dose for resveratrol of 11.2 mg/kg.

Rat dose (mg/kg) = mouse dose (22.4 mg/kg) ×  mouse Km(3)  = 11.2 mg/kg
rat Km(6)
1

References

[1] Steegmaier M, et al. Curr Biol, 2007, 17(4), 316-322.

[2] Nappi TC, et al. Cancer Res, 2009, 69(5), 1916-1923.

Clinical Trial Information( data from http://clinicaltrials.gov, updated on 2016-04-16)

NCT Number Recruitment Conditions Sponsor
/Collaborators
Start Date Phases
NCT00526149 Completed Breast Cancer|Endometrial Cancer|Head and Neck Cancer|Melanoma (Skin)|Ovarian Cancer|Sarcoma European Organisation for Research and Treatment of Cance  ...more European Organisation for Research and Treatment of Cancer - EORTC July 2007 Phase 2
NCT02215044 Terminated Pancreatic Neoplasms Boehringer Ingelheim June 2007 Phase 1
NCT00412880 Completed Carcinoma, Small Cell Boehringer Ingelheim January 2007 Phase 2
NCT02211833 Completed Carcinoma, Non-Small-Cell Lung Boehringer Ingelheim October 2006 Phase 1
NCT00701766 Completed Leukemia, Myeloid, Acute Boehringer Ingelheim October 2006 Phase 2

view more

Chemical Information

Download BI 2536 SDF
Molecular Weight (MW) 521.66
Formula

C28H39N7O3

CAS No. 755038-02-9
Storage 3 years -20℃powder
6 months-80℃in solvent
Synonyms N/A
Solubility (25°C) * In vitro DMSO 21 mg/mL (40.25 mM)
Ethanol 100 mg/mL (191.69 mM)
Water <1 mg/mL (<1 mM)
In vivo 30% PEG400/0.5% Tween80/5% propylene glycol 15 mg/mL
* <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 (R)-4-(8-cyclopentyl-7-ethyl-5-methyl-6-oxo-5,6,7,8-tetrahydropteridin-2-ylamino)-3-methoxy-N-(1-methylpiperidin-4-yl)benzamide

Customer Product Validation (10)


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Rating
Source Cancer Cell 2010 18, 641–654. BI 2536 purchased from Selleck
Method Immunohistochemical analysis
Cell Lines SCID mice
Concentrations
Incubation Time
Results Control tumors grow exponentially, whereas genetic ablation of Cdc20 results in complete tumor regression in about 10 days. In comparison, treatment with taxol, vincristine, and BI2536 only induces partial responses ( Figure C). In agreement with the data observed in vitro, immunohistochemical analysis of samples taken 3 days after the first treatment indicates lack of proliferation (as measured by BrdU incorporation) and massive apoptosis in Cdc20 null cells, whereas all other samples maintain significant levels of proliferation and reduced apoptotic markers ( Figure D)

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Rating
Source Nat Cell Biol 2011 13, 1265-71. BI 2536 purchased from Selleck
Method cold-stable assay, Immunological methods
Cell Lines RPE1 cells
Concentrations 40 nM
Incubation Time 45 min
Results To examine whether B56-PP2A attenuates the signalling of kinetochore kinases other than Aurora B, we examined Plk1. Remarkably, wash-in of the Plk1 inhibitor BI2536 also restored K-fibres in B56-PP2A-siRNA cells.

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Rating
Source Cancer Res 2011 71, 1385-1395. BI 2536 purchased from Selleck
Method Western blot
Cell Lines NB TICs
Concentrations 10-100 nM
Incubation Time 16/24 h
Results The cyclin-dependent kinase inhibitor p21, a newly identified substrate of PLK1, also accumulated in BI 2536-treated NB TICs (Fig. C). Finally, PLK1 kinase activity was inhibited following treatment of NB TICs with BI 2536, as demonstrated by anti-PLK1 immunoprecipitiation followed by in vitro PLK1 kinase assay ( Fig. D)

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Rating
Source Cancer Res 2011 71, 1385-1395. BI 2536 purchased from Selleck
Method Trypan blue exclusion, Western blot
Cell Lines NB TICs
Concentrations 10/30/100 nmol/L
Incubation Time 24/48/72 h
Results We observed a significant difference in viable cell numbers in BI2536-treated samples, as compared with DMSO (vehicle)-treated cells, at 24 hours posttreatment. The difference became more pronounced with time, such that at 72 hours there were only approximately 20% viable cells following treatment with 10 nmol/L BI 2536 and approximately 2% to 5% viable cells following treatment with 30 and 100 nmol/L as compared with DMSO-treated cells(Fig. A). At 24 hours, BI 2536-treated cells gradually accumulated cleaved PARP, in contrast to DMSO-treated cells, suggesting induction of apoptosis ( Fig. B).

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Rating
Source Cancer Res 2011 71, 1385-1395. BI 2536 purchased from Selleck
Method Tumor growth assay
Cell Lines NOD/SCID mice
Concentrations 12.5-25 mg/kg
Incubation Time 0-24 d
Results As shown in Figure A,tumor growth was significantly inhibited following administration of the third cycle of BI 2536 therapy. When mice were administered 25 mg/ kg weekly on 2 consecutive days for a total of only 2 cycles, tumor growth was inhibited after the second cycle ( Fig. B)

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Rating
Source Cancer Res 2011 71, 1385-1395. BI 2536 purchased from Selleck
Method Tumor growth assay
Cell Lines NOD/SCID mice
Concentrations 12.5 mg/kg
Incubation Time 0-60 d
Results We examined a combination of a low dose of BI 2536 (12.5 mg/kg) with a low dose of irinotecan (10 mg/kg), a topoisomerase inhibitor with demonstrated activity in phase II clinical trials in relapsed NB. Single-agent treatment with low-dose of either BI 2536 or irinotecan was comparably efficacious, resulting in significant growth inhibition, as compared with vehicle-treated animals.

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Rating
Source J Invest Dermatol 2011 131, 1886–1895. BI 2536 purchased from Selleck
Method MTT assay
Cell Lines M14 cell lines, WM-115 cell lines
Concentrations 1-20 nM
Incubation Time
Results Specific inhibition of Plk-1 using the commercially available inhibitor BI 2536 led to a dose- and time-dependent decrease in human melanoma cell viability.

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Rating
Source J Biol Chem 2012 287(21), 17088-99. BI 2536 purchased from Selleck
Method Cell death assay
Cell Lines HeLa cells
Concentrations 0 -120 nM
Incubation Time 72 h
Results In HeLa cells lacking GRK5, there was an~4.5-fold decrease in the EC50 of BI 2536-mediated cell death (control shRNA, 18.9 ?.2 nM; GRK5 shRNA, 4.1?.1 nM, n = 6, p < 0.05) (Fig. A). Similar to BI 2536, cells lacking GRK5 were ~4.5-fold more sensitive to GSK461364 compared with control cells (EC50 for control shRNA cells, 17.0 ?4.4 nM;EC50 for GRK5-shRNA cells, 3.8 ?0.7 nM, n = 4, p < 0.05).

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Rating
Source J Biomol Screen 2013 18(1), 54-66. BI 2536 purchased from Selleck
Method 3D Tumor Growth Assay
Cell Lines PC-3M cells
Concentrations 0-10uM
Incubation Time 7 d
Results Reduction of colony size was observed for a number of established anticancer drugs, and IC 50 values for those reference compounds were determined based on the average area size of the cluster. IC 50 values were determined in two independent experiments and were quite reproducible.

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Rating
Source J Biomol Screen 2013 18(1), 54-66. BI 2536 purchased from Selleck
Method 3D Tumor Growth Assay
Cell Lines PC-3M cells
Concentrations 10uM
Incubation Time 7 d
Results Reduction of colony size was observed for a number of established anticancer drugs, and IC 50 values for those reference compounds were determined based on the average area size of the cluster. IC 50 values were determined in two independent experiments and were quite reproducible.

Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

Tel: +1-832-582-8158 Ext:3

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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
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