BRD-5648 (also known as (R)-BRD-0705) is an inactive (R)-enantiomer of BRD-0705. BRD-0705 is a potent GSK3α inhibitor (IC50: 66 nM; Kd: 4.8 μM). It can be used for acute myeloid.

Physicochemical Properties

Molecular Formula	C20H23N3O
Molecular Weight	321.416124582291
Exact Mass	321.1841
Elemental Analysis	C, 74.74; H, 7.21; N, 13.07; O, 4.98
CAS #	2056261-42-6
Related CAS #	BRD0705;2056261-41-5;(Rac)-BRD0705;1597440-03-3
PubChem CID	136980414
Appearance	Light yellow to khaki solid powder
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	3
Rotatable Bond Count	2
Heavy Atom Count	24
Complexity	561
Defined Atom Stereocenter Count	1
InChi Key	NCKLQXXBRWCYMA-HXUWFJFHSA-N
InChi Code	InChI=1S/C20H23N3O/c1-4-20(13-8-6-5-7-9-13)14-12-21-23-18(14)22-15-10-19(2,3)11-16(24)17(15)20/h5-9,12H,4,10-11H2,1-3H3,(H2,21,22,23)/t20-/m1/s1
Chemical Name	(4R)-4-ethyl-7,7-dimethyl-4-phenyl-1,6,8,9-tetrahydropyrazolo[3,4-b]quinolin-5-one
Synonyms	BRD 5648; BRD5648; BRD-5648; (R)-BRD-0705; (R)-BRD0705; (R)-BRD 0705
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	BRD5648, the inactive enantiomer of BRD0705, does not induce changes in enzyme phosphorylation or total β-catenin protein stabilization[1].
ln Vitro	BRD5648, the inactive enantiomer of BRD0705, does not induce changes in enzyme phosphorylation or total β-catenin protein stabilization[1]. BRD5648 did not induce changes in GSK3α/β enzyme phosphorylation (Tyr279/216) or total β-catenin protein stabilization in U937 AML cells when treated at a concentration of 10 µM for 24 hours, supporting its designation as an inactive control compound. [1]
Enzyme Assay	A mobility shift microfluidic assay was used to determine the inhibitory activity of compounds against GSK3α and GSK3β. Purified kinase was incubated with the test compound in the presence of ATP (at or below the Km) and a fluorescent peptide substrate for 60 minutes at room temperature. The reaction was terminated by EDTA addition, and substrate and product were separated electrophoretically. Fluorescence intensity was measured to calculate percent conversion and derive IC50 values. BRD5648 was tested using a 12-point dose curve with 3-fold serial dilution starting from 33 µM, and was found to be relatively inactive. [1]
Cell Assay	For Western blot analysis of GSK3 phosphorylation and β-catenin levels, U937 AML cells were treated with BRD5648 (10 µM) or other compounds for 24 hours. Cells were lysed, and proteins were resolved by gel electrophoresis, transferred to membranes, and probed with specific antibodies against phospho-GSK3α/β (Tyr279/216), total GSK3α/β, and β-catenin. BRD5648 treatment showed no effect on these markers. [1]
References	[1]. Exploiting an Asp-Glu "switch" in glycogen synthase kinase 3 to design paralog-selective inhibitors for use in acute myeloid leukemia. Sci Transl Med. 2018 Mar 7;10(431). pii: eaam8460.
Additional Infomation	BRD5648 is the enantiomer (inverted quaternary center stereochemistry) of the GSK3α-selective inhibitor BRD0705. Its inactivity demonstrates the limited three-dimensional space and strict stereochemical requirements within the ATP-binding domain of GSK3 kinases for inhibitor binding. [1] BRD5648 serves as a negative control compound in the study to validate the on-target activity of the active enantiomer BRD0705. [1]

Solubility Data

Solubility (In Vitro)

DMSO: ~300 mg/mL (~933.4 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 7.5 mg/mL (23.33 mM) (saturation unknown) in 10% DMSO + 40% PEG300 +5% Tween-80 + 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 75.0 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.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.1112 mL	15.5560 mL	31.1119 mL
	5 mM	0.6222 mL	3.1112 mL	6.2224 mL
	10 mM	0.3111 mL	1.5556 mL	3.1112 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.