4-Br-Bnlm is a selective inhibitor of glucose-regulated protein (Grp94) with EC50 of 0.96 µM. 4-Br-Bnlm reduces the levels of misfolding of mutant and wild-type cardiac proteins in cells. 4-Br-Bnlm promotes the clearance of toxic myocardin and reduces myocardial toxicity.

Physicochemical Properties

Molecular Formula	C20H18BRCLN2O4
Molecular Weight	465.72
Exact Mass	464.013
CAS #	1654775-71-9
PubChem CID	127049159
Appearance	Typically exists as solid at room temperature
LogP	4.9
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	5
Rotatable Bond Count	7
Heavy Atom Count	28
Complexity	520
Defined Atom Stereocenter Count	0
InChi Key	FHDUIAYTJZAXNW-UHFFFAOYSA-N
InChi Code	InChI=1S/C20H18BrClN2O4/c1-28-20(27)18-14(19(22)16(26)10-15(18)25)6-7-17-23-8-9-24(17)11-12-2-4-13(21)5-3-12/h2-5,8-10,25-26H,6-7,11H2,1H3
Chemical Name	methyl 2-[2-[1-[(4-bromophenyl)methyl]imidazol-2-yl]ethyl]-3-chloro-4,6-dihydroxybenzoate
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 Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.
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	EC50: 0.96 µM (glucose-regulated protein 94, Grp94)[1][2]
ln Vitro	In human embryonic kidney (HEK) cells, 4-Br-Bnlm (0-100 µM; 24 h) suppresses the production of the 1477N mutant cardiac protein [1]. The toxicity and levels of mutant myocardin in primary human trabecular meshwork (HTM) cells are reduced by 4-Br-Bnlm (30 µM; 24 h) [1]. In a dose-dependent manner, 4-Br-Bnlm (3, 10 and 30 µM; 24 h) suppresses myocardin production in myosin cells [1]. In HTM cells, 4-Br-Bnlm (30 µM and 100 µM; 24 h) does not cause a heat shock response [2].
ln Vivo	In the Tg-MYOCY437H mouse model, 4-Br-Bnlm (300 µM; eye drops; once daily for 12 weeks) decreased intraocular pressure and removed mutant cardiomyopathy within trabecular meshwork (TM) cells. The photonegative reaction (PhNR) defect is improved by 4-Br-Bnlm[2].
Cell Assay	Cell Viability Assay[1] Cell Types: Primary human HTM cells. Tested Concentrations: 30 µM. Incubation Duration: 24 h. Experimental Results: Dramatically diminished the level of Y437H mutant myocilin without affecting WT myocilin and the RFP vector in HEK cells. Western Blot Analysis [1] Cell Types: HEK cells. Tested Concentrations: 0, 1, 3, 10, 30 and 100 µM. Incubation Duration: 24 h. Experimental Results: Dramatically diminished 1477N mutant myocilin in HEK cells expression without causing the degradation of other Hsp90-dependent clients Akt and Ras.
Animal Protocol	Animal/Disease Models: Tg-MYOCY437H mice[2]. Doses: 300 µM. Route of Administration: Eye drops; one time/day for 12 weeks. Experimental Results: Inhibited mutant myocilin expression and restored the PhNR deficits.
References	[1]. Stothert AR, et al, Fontaine SN, Crowley VM, Mishra S, Blagg BS, Lieberman RL, Dickey CA. Exploiting the interaction between Grp94 and aggregated myocilin to treat glaucoma. Hum Mol Genet. 2014 Dec 15;23(24):6470-80. [2]. Stothert AR, et al. Isoform-selective Hsp90 inhibition rescues model of hereditary open-angle glaucoma. Sci Rep. 2017 Dec 20;7(1):17951.

Solubility Data

Solubility (In Vitro)

DMSO: 33.33 mg/mL (71.57 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.37 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 (5.37 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.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.1472 mL	10.7361 mL	21.4721 mL
	5 mM	0.4294 mL	2.1472 mL	4.2944 mL
	10 mM	0.2147 mL	1.0736 mL	2.1472 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.