RO2959 mono HCl is a potent and specific CRAC channel inhibitor (antagonist) with IC50 of 402 nM. RO2959 mono HCl is a potent blocker of storage calcium entry (SOCE) mediated by Orai1/Stim1 channels with IC50 of 25 nM. RO2959 mono HCl is also a potent inhibitor of human IL-2 production, effectively blocking T cell receptor-triggered gene expression and T cell function pathways.

Physicochemical Properties

Molecular Formula	C21H20CLF2N5OS
Molecular Weight	463.931208610535
Exact Mass	463.104
CAS #	2309172-44-7
Related CAS #	RO2959 hydrochloride;1219927-22-6
PubChem CID	45274292
Appearance	Light brown to brown solid powder
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	8
Rotatable Bond Count	4
Heavy Atom Count	31
Complexity	655
Defined Atom Stereocenter Count	0
SMILES	Cl.S1C(C)=CN=C1N1CC(C2C=NC(=CN=2)NC(C2C(=CC=CC=2F)F)=O)=C(C)CC1
InChi Key	GMLSDODALBRLPE-UHFFFAOYSA-N
InChi Code	InChI=1S/C21H19F2N5OS.ClH/c1-12-6-7-28(21-26-8-13(2)30-21)11-14(12)17-9-25-18(10-24-17)27-20(29)19-15(22)4-3-5-16(19)23;/h3-5,8-10H,6-7,11H2,1-2H3,(H,25,27,29);1H
Chemical Name	2,6-difluoro-N-[5-[4-methyl-1-(5-methyl-1,3-thiazol-2-yl)-3,6-dihydro-2H-pyridin-5-yl]pyrazin-2-yl]benzamide;hydrochloride
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, avoid exposure to moisture.
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	CRAC channel 402 nM (IC50) Orai1/Stim1 channels 25 nM (IC50) IL-2
ln Vitro	RO2959 has an IC50 of 25 nM for Orai1 and 530 nM for Orai3 inhibition. With an IC50 value of 265 nM, RO2959 inhibits store operated calcium entry (SOCE) in activated CD4+T lymphocytes[1]. Strong SOCE inhibitor RO2959 inhibits both IP3-dependent current in CRAC-expressing RBL-2H3 cells and CHO cells that exhibit stable human Orai1 and Stim1 expression, as well as SOCE in human primary CD4+ T cells stimulated by TCR activation or thapsigargin therapy. When TCR stimulation or mixed lymphocyte reaction (MLR) is used to stimulate T cells, RO2959 totally blocks both cytokine generation and T cell proliferation[1].
References	[1]. Characterization of a Novel CRAC Inhibitor That Potently Blocks Human T Cell Activation and Effector Functions. Mol Immunol. 2013 Jul;54(3-4):355-67. [2]. Gastrodin Inhibits Store-Operated Ca 2+ Entry and Alleviates Cardiac Hypertrophy. Front Pharmacol. 2017 Apr 25;8:222.

Solubility Data

Solubility (In Vitro)

DMSO: 25 mg/mL (53.89 mM) H2O: < 0.1 mg/mL

Solubility (In Vivo)

Solubility in Formulation 1: 2.08 mg/mL (4.48 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 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: ≥ 2.08 mg/mL (4.48 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 20.8 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: ≥ 2.08 mg/mL (4.48 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 20.8 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	2.1555 mL	10.7775 mL	21.5550 mL
	5 mM	0.4311 mL	2.1555 mL	4.3110 mL
	10 mM	0.2155 mL	1.0777 mL	2.1555 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.