Rabusertib (LY2603618) 911222-45-2_Chk_Cell Cycle_Signaling Pathways_Products

Rabusertib (also known as IC-83; LY-2603618; LY 2603618; IC83) is a novel, potent and selective Chk1 (cell cycle checkpoint kinase 2) inhibitor with potential antitumor activity. In a cell-free assay, it inhibits Chk1 with an IC50 of 7 nM and may have anti-tumor effects. By binding to and inhibiting the activity of chk2, LY2603618 may increase the antitumor efficacies of different chemotherapeutic agents by preventing DNA damage caused by DNA-damaging agents from being repaired. Double-strand breaks (DSBs) activate ATP-dependent serine-threonine kinase Chk2, which is an essential part of the DNA replication-monitoring checkpoint system and is overexpressed by a range of cancer cell types.

Physicochemical Properties

Molecular Formula

C18H22BRN5O3

Molecular Weight

436.3

Exact Mass

435.09

Elemental Analysis

C, 49.55; H, 5.08; Br, 18.31; N, 16.05; O, 11.00

CAS #

911222-45-2

Related CAS #

911222-45-2

PubChem CID

11955855

Appearance

White to off-white solid powder

Density

1.5±0.1 g/cm3

Boiling Point

503.1±50.0 °C at 760 mmHg

Flash Point

258.1±30.1 °C

Vapour Pressure

0.0±1.3 mmHg at 25°C

Index of Refraction

1.633

LogP

2.1

Hydrogen Bond Donor Count

Hydrogen Bond Acceptor Count

Rotatable Bond Count

Heavy Atom Count

Complexity

486

Defined Atom Stereocenter Count

SMILES

O(C1=CC(C)=C(Br)C=C1NC(=O)NC1N=CC(C)=NC=1)C[C@H]1OCCNC1

InChi Key

SYYBDNPGDKKJDU-ZDUSSCGKSA-N

InChi Code

InChI=1S/C18H22BrN5O3/c1-11-5-16(27-10-13-8-20-3-4-26-13)15(6-14(11)19)23-18(25)24-17-9-21-12(2)7-22-17/h5-7,9,13,20H,3-4,8,10H2,1-2H3,(H2,22,23,24,25)/t13-/m0/s1

Chemical Name

1-[5-bromo-4-methyl-2-[[(2S)-morpholin-2-yl]methoxy]phenyl]-3-(5-methylpyrazin-2-yl)urea

Synonyms

IC-83; LY2603618; IC 83; LY 2603618; IC83; LY-2603618

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

Chk1 (IC50 = 7 nM); Chk2 (IC50 = 12000 nM); PDK1 (IC50 = 893 nM); CAMK2 (IC50 = 1550 nM); VEGFR3 (IC50 = 2128 nM); MET (IC50 = 2200 nM); JNK1 (IC50 = 4930 nM); RSK2 (IC50 = 5700 nM); NTRK1 (IC50 = 12000 nM)

ln Vitro

Rabusertib (LY2603618) is a potent inhibitor of several Chk1 biological processes. In vitro tests of rabusertib (LY2603618) are conducted against a panel of 51 different protein kinases. Rabusertib (LY2603618) has an IC50 of 7 nM for Chk1, which makes it approximately 100 times more potent against Chk1 than it is against all other protein kinases tested (IC50=893 nM for PDK1, >1000 nM for the others). With an EC50 of 430 nM, rabusertib (LY2603618) successfully decreased Chk1 autophosphorylation. Rabusertib (LY2603618) effectively inhibited Chk1 in cells treated with DNA damaging agents, thereby abrogating the G₂/M DNA damage checkpoint. When cells were treated with Rabusertib (LY2603618), a cellular phenotype that was reported for Chk1 depletion by RNA interference (RNAi) was observed. When Rabusertib (LY2603618) inhibits intracellular Chk1, it leads to decreased DNA synthesis and increased H2A. X phosphorylation is a marker for early mitotic entry and DNA damage[1]. MTT assays show dose-dependent inhibition of cell growth in response to varying concentrations of Rabusertib (LY2603618) treatment of SK-N-BE(2) cells, with an IC50 of 10.81 µM[1].

ln Vivo

Rabusertib (LY2603618) at a single concurrent oral dosage of 200 mg/kg and 150 mg/kg (IP) of Gemcitabine are administered to mice containing Calu-6 xenografts. Rabusertib (LY2603618) at 200 mg/kg is enough to inhibit 85% of Chk1 autophosphorylation in vivo after two hours. Rabusertib (LY2603618), a selective chemical inhibitor of Chk1, supports the cited report by effectively reducing Gemcitabine-induced phosphorylation on Tlk serine 695[1].

Cell Assay

Cells are plated on 96-well tissue culture plates at a density of 2.5×10³ per well, and then incubated for one cell doubling (18–24 hours). The final concentration range of 1-1000 nM is covered by half-log steps when setting up gemcitabine dilutions. Rabusertib (LY2603618) is made by dilutions in DMSO to a final concentration of 5000×, and then a 1000-fold dilution in medium to produce 5× stocks that are added to wells. Rabusertib (LY2603618) is added about 24 hours after the addition of gemcitabine. Three copies of each combination are made. Following the addition of Rabusertib (LY2603618) and allowing two cell doublings, MTS/PMS reagent is added to each well in accordance with the manufacturer's instructions. A Spectra Max 250 spectrophotometer is used to measure absorbance at 490 nm, and GraphPad Prism 4.0 is used to analyze the results. Non-linear regression is used to fit dose-response curves, with the bottom fits being limited to 0% inhibition[1].

Animal Protocol

Mice: These investigations employ 26–28 g female Harlan athymic nude mice. Each subject animal's rear flank is subcutaneously injected with 1×10⁶ Calu-6 cells in a 1:1 mixture of serum-free growth medium and Matrigel to start the tumor's growth. The animals are randomly assigned to treatment groups based on body weight and tumor size once the tumor volumes have grown to a size of about 150 mm³. A total of two injections are given to each animal: one is an intraperitoneal injection of 150 mg/kg Gemcitabine or saline vehicle, and the other is an oral 200 μL dose of LY2603618 or Captisol vehicle.

References

[1]. Characterization and preclinical development of LY2603618: a selective and potent Chk1 inhibitor. Invest New Drugs. 2014 Apr;32(2):213-26.

[2]. Panobinostat synergistically enhances the cytotoxic effects of cisplatin, doxorubicin or etoposide on high-risk neuroblastoma cells. PLoS One. 2013 Sep 30;8(9):e76662.

Additional Infomation

1-[5-bromo-4-methyl-2-[[(2S)-2-morpholinyl]methoxy]phenyl]-3-(5-methyl-2-pyrazinyl)urea is a member of ureas.
Rabusertib has been used in trials studying the treatment of Cancer, Solid Tumors, Advanced Cancer, Pancreatic Neoplasms, and Non Small Cell Lung Cancer.
Rabusertib is an inhibitor of the cell cycle checkpoint kinase 2 (chk2) with potential chemopotentiating activity. Rabusertib binds to and inhibits the activity of chk2, which may prevent the repair of DNA caused by DNA-damaging agents, thus potentiating the antitumor efficacies of various chemotherapeutic agents. Chk2, an ATP-dependent serine-threonine kinase, is a key component in the DNA replication-monitoring checkpoint system and is activated by double-stranded breaks (DSBs); activated chk2 is overexpressed by a variety of cancer cell types.

Solubility Data

Solubility (In Vitro)

DMSO: ~13 mg/mL (~29.8 mM)

Water: <1 mg/mL

Ethanol: <1 mg/mL

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.73 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 (5.73 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 (5.73 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.

Solubility in Formulation 4: 2% DMSO +30% PEG400+0.5% Tween80+5% Propylene glycol : 30mg/mL

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

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.2920 mL	11.4600 mL	22.9200 mL
	5 mM	0.4584 mL	2.2920 mL	4.5840 mL
	10 mM	0.2292 mL	1.1460 mL	2.2920 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.

PeptideDB

Rabusertib (LY2603618) 911222-45-2

CAS No.: 911222-45-2

Physicochemical Properties

Biological Activity

Solubility Data