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AC480 (BMS-599626)

Name: AC480 (BMS-599626)
Brand: Selleck Chemicals
SKU: S1056
Rating: 5 (10 reviews)

Catalog No.S1056 Purity:99.08%

AC480 (BMS-599626) is a selective and efficacious inhibitor of HER1 and HER2 with IC50 of 20 nM and 30 nM, ~8-fold less potent to HER4, >100-fold to VEGFR2, c-Kit, Lck, MET etc. Phase 1.

AC480 (BMS-599626) Chemical Structure

CAS No. 714971-09-2

Purity & Quality Control

Batch: Purity: 99.08%

99.08

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Signaling Pathway

HER2 Signaling Pathway

Biological Activity

Description

AC480 (BMS-599626) is a selective and efficacious inhibitor of HER1 and HER2 with IC50 of 20 nM and 30 nM, ~8-fold less potent to HER4, >100-fold to VEGFR2, c-Kit, Lck, MET etc. Phase 1.

Targets

HER1 ^[1]	HER2 ^[1]	HER4 ^[1]
20 nM	30 nM	190 nM

In vitro
In vitro	BMS-599626 also inhibits the related receptor HER4, but with reduced potency with IC50 of 190 nM. BMS-599626 is identified as an ATP-competitive inhibitor for HER1 and as an ATP-noncompetitive inhibitor for HER2 with K_i of 2 nM and 5 nM, respectively. BMS-599626 inhibits the proliferation of tumor cells expressing high levels of HER1 and/or HER2, including Sal2, BT474, N87, KPL-4, HCC202, HCC1954, HCC1419, AU565, ZR-75-30, MDA-MB-175, GEO, and PC9 cells with IC50 of 0.24 μM, 0.31 μM, 0.45 μM, 0.38μM, 0.94 μM, 0.34 μM, 0.75 μM, 0.63 μM, 0.51 μM, 0.84 μM, 0.90 μM and 0.34 μM, respectively. While the proliferation of the ovarian tumor cell line A2780 and MRC5 fibroblasts, neither of which expresses HER1 or HER2, are not inhibited significant by BMS-599626. ^[1] A recent study shows that BMS-599626 significantly enhances the radiosensitivity of HN-5 cells expressing both EGFR and Her2 cell, by promoting cycle redistribution and inhibiting DNA repair. ^[2]
Kinase Assay	Protein kinase assays
Kinase Assay	The entire cytoplasmic sequences of HER1, HER2, and HER4 are expressed as recombinant proteins in Sf9 insect cells. HER1 and HER4 are expressed as fusion proteins with glutathione-S-transferase and are purified by affinity chromatography on glutathione-S-Sepharose. HER2 is subcloned into the pBlueBac4 vector and expressed as an untagged protein using an internal methionine codon (M687) for translation initiation. The truncated HER2 protein is isolated by chromatography on a column of DEAE-Sepharose equilibrated in a buffer that contains 0.1 M NaCl, and the recombinant protein is eluted with a buffer containing 0.3 M NaCl. For the HER kinase assays, reaction volumes are 50 μL and contains 10 ng of glutathione-S-transferase fusion protein or 150 ng of partially purified HER2. The mixtures also contains 1.5 μM poly(Glu/Tyr) (4:1), 1 μM ATP, 0.15 μCi [γ-³³P]ATP, 50 mM Tris-HCl (pH 7.7), 2 mM DTT, 0.1 mg/mL bovine serum albumin, and 10 mM MnCl₂. Reactions are allowed to proceed at 27°C for 1 hour and are terminated by the addition of 10 μL of a stop buffer (2.5 mg/mL bovine serum albumin and 0.3 M EDTA), followed by a 108-μL mixture of 3.5 mM ATP and 5% trichloroacetic acid. Acid-insoluble proteins are recovered on GF/C Unifilter plates with a Filtermate harvester. Incorporation of radioactive phosphate into the poly(Glu/Tyr) substrate is determined by liquid scintillation counting. Percent inhibition of kinase activity is determined by nonlinear regression analyses and data are reported as the inhibitory concentration required to achieve 50% inhibition relative to control reactions (IC50). Data are the averages of triplicate determinations. All other tyrosine kinases are also assayed using poly(Glu/Tyr) as a substrate. Kinetics of HER1 and HER2 inhibition are determined in reaction mixtures that contains varying concentrations of ATP and BMS-599626.
Cell Research	Cell lines	Sal2, BT474, N87, KPL-4, HCC202, HCC1954, HCC1419, AU565, ZR-75-30, MDA-MB-175, GEO, PC9, A2780 and MRC5
	Concentrations	0-10 μM
	Incubation Time	72 hours
	Method	All cell lines are maintained in RPMI 1640 supplemented with 10% fetal bovine serum, 100 units/mL penicillin, and 100 μg/mL streptomycin. Cells are plated at 1,000 per well in 96-well plates and are cultured for 24 hours before BMS-599626 is added. BMS-599626 is diluted in culture medium such that the final concentrations of DMSO are ≤ 1%. Following the addition of BMS-599626, the cells are cultured for an additional 72 hours before cell viability is determined by measuring the conversion of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide dye with the CellTiter96 kit. For some cell lines, there is a lack of a correlation between 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide dye metabolism and cell number, and a thymidine uptake assay is used to measure proliferation of these cell lines. Cells are plated in 96-well plates and treated with compounds as above. At the end of the 72-hour incubation, cells are pulsed with [₃H]thymidine (0.4 μCi/well) for 3 hours before they are harvested. Cells are digested with 2.5% trypsin for 10 minutes at 37 °C and are harvested by filtration using a Packard Filtermate Harvester and GF/C Unifilter plates. Incorporation of radioactive thymidine into nucleic acids is determined by liquid scintillation counting.

In Vivo
In vivo	In vivo, oral administration of BMS-599626 results in a dose-dependent inhibition of Sal2 tumor growth at doses ranging from 60 mg/kg to 240 mg/kg, yielding a potent antitumor activity in a human breast tumor KPL-4 xenograft at its maximum tolerated dose of 180 mg/kg, and also has similar antitumor activity in other HER2 amplified xenograft models, as well as other HER1-overexpressing xenograft models. ^[1]
Animal Research	Animal Models	SAL2 murine salivary gland tumor, N87 human gastric carcinoma, BT474 human breast tumor, A549 human non–small-cell lung tumor, and GEO human colon tumor are maintained and passaged in athymic female nude mice.
	Dosages	≤240 mg/kg
	Administration	Administered via p.o.

NCT Number	Recruitment	Conditions	Sponsor/Collaborators	Start Date	Phases
Clinical Trial Information (data from https://clinicaltrials.gov, updated on 2024-05-22)
NCT01245543	Withdrawn	Solid Tumors	Daiichi Sankyo	November 2010	Phase 1
NCT00979173	Completed	Glioma	Annick Desjardins\|Ambit Biosciences Corporation\|Duke University	November 2009	Phase 1
NCT00093730	Completed	Unspecified Adult Solid Tumor Protocol Specific	Jonsson Comprehensive Cancer Center\|National Cancer Institute (NCI)	August 2004	Phase 1
NCT00095537	Completed	Cancer\|Metastases	Bristol-Myers Squibb	March 2004	Phase 1

References

Chemical Information & Solubility

Molecular Weight	530.55	Formula	C₂₇H₂₇FN₈O₃
CAS No.	714971-09-2	SDF	Download AC480 (BMS-599626) SDF
Smiles	CC1=C2C(=NC=NN2C=C1NC(=O)OCC3COCCN3)NC4=CC5=C(C=C4)N(N=C5)CC6=CC(=CC=C6)F
Storage (From the date of receipt)	3 years-20°Cpowder	Storage of Stock Solutions Aliquot the stock solutions to avoid repeated freeze-thaw cycles	1 year-80°Cin solvent
Storage (From the date of receipt)	3 years-20°Cpowder		1 month-20°Cin solvent

In vitro
Batch:

DMSO : 113 mg/mL ( (212.98 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Ethanol : 20 mg/mL

Water : Insoluble

Molecular Weight Calculator

In vivo Batch: Add solvents to the product individually and in order.				In vivo Formulation Calculator
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.
Clear solution	30%PEG400 1 0.5%Tween80 2 5%propylene glycol Validated by Selleck labs. Should you need adjustments to this formulation, contact our sales team for custom testing.	30.000mg/ml (56.55mM)	Taking the 1 mL working solution as an example, add 300 μL of 100 mg/ml clarified PEG400 stock solution to 5 μL of Tween80, mix evenly to clarify it; add 50 μL Propylene glycol to the above system, mix evenly to clarify it; then continue to add 645 μL ddH2O to adjust the volume. to 1 mL. The mixed solution should be used immediately for optimal results.
Homogeneous suspension	CMC-NA	≥5mg/ml	Taking the 1 mL working solution as an example, add 5 mg of this product to 1 ml of CMC-Na solution, mix evenly to obtain a homogeneous suspension with a final concentration of 5 mg/ml.

Preparing Stock Solutions

Molarity Calculator

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Dilution Calculator Molecular Weight Calculator

In vivo Formulation Calculator (Clear solution)

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

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Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid 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 master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH₂O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such
as vortex, ultrasound or hot water bath can be used to aid dissolving.

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.

Handling Instructions

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