RO5461111 is a specific, orally bioactive Cathepsin S antagonist (IC50= 0.4 nM, human Cathepsin S; 0.5 nM, murine Cathepsin S). RO5461111 effectively inhibits antigen-specific T cell and B cell activation. RO5461111 has an improvement effect on lung inflammation and lupus nephritis.

Physicochemical Properties

Molecular Formula	C27H24F6N4O4S
Molecular Weight	614.559286117554
Exact Mass	614.142
CAS #	1252637-46-9
PubChem CID	59543651
Appearance	White to off-white solid powder
LogP	3.6
Hydrogen Bond Donor Count	1
Hydrogen Bond Acceptor Count	12
Rotatable Bond Count	6
Heavy Atom Count	42
Complexity	1250
Defined Atom Stereocenter Count	2
SMILES	CC1=NC=CC(=C1)C2=CC(=C(C=C2)S(=O)(=O)[C@@H]3C[C@H](N(C3)C(=O)C4(CC4)C(F)(F)F)C(=O)NC5(CC5)C#N)C(F)(F)F
InChi Key	CDBYZSRUFZXRLK-QUCCMNQESA-N
InChi Code	InChI=1S/C27H24F6N4O4S/c1-15-10-17(4-9-35-15)16-2-3-21(19(11-16)26(28,29)30)42(40,41)18-12-20(22(38)36-24(14-34)5-6-24)37(13-18)23(39)25(7-8-25)27(31,32)33/h2-4,9-11,18,20H,5-8,12-13H2,1H3,(H,36,38)/t18-,20+/m1/s1
Chemical Name	(2S,4R)-N-(1-cyanocyclopropyl)-4-[4-(2-methylpyridin-4-yl)-2-(trifluoromethyl)phenyl]sulfonyl-1-[1-(trifluoromethyl)cyclopropanecarbonyl]pyrrolidine-2-carboxamide
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	human Cathepsin S 0.4 nM (IC50) murine Cathepsin S 0.5 nM (IC50)
ln Vitro	In mouse (A20) or human (RAJI) cell lines, RO5461111 (16 h) is cytoactive and dramatically lowers splenic dendritic cell activation, as well as the subsequent growth and activation of CD4 T cells and CD4/CD8 double negative T cells[1].
ln Vivo	Following sheep IgG immunization, RO5461111 (0.1–100 mg/kg; oral; single dosage) prevents T cell priming and anti-sheep IgG in BALB/c mice[1]. In MRL-Fas(lpr) mice, RO5461111 (1.31 mg/mouse or 30 mg/kg; oral; 8 weeks) decreases hypergammaglobulinemia, lupus autoantibody product F, and disturbs the germinal center (e.g., diminishes CXCL12 expression). Moreover, RO5461111 can lessen lupus nephritis and lessen lung inflammation[1].
Animal Protocol	Animal/Disease Models: Female MRL-Fas(lpr) mice (12weeks old; with proteinuria/serum creatinine levels up)[1] Doses: 262.5 mg/kg chow; or 5 mg/mouse with 1.31 mg/mouse Route of Administration: Oral gavage; 8 weeks Experimental Results: decreased the activation and expansion of spleen dendritic cells, CD4, double-negative T cells and plasma cells. decreased plasma levels of IL-10 and TNF-α.
References	[1]. Cathepsin S inhibition suppresses systemic lupus erythematosus and lupus nephritis because cathepsin S is essential for MHC class II-mediated CD4 T cell and B cell priming. Ann Rheum Dis. 2015 Feb;74(2):452-63. [2]. Preparation of proline dipeptidyl nitrile derivatives as cathepsin, particularly cathepsin S and L, inhibitors: United States, US20100267722. 2010-10-21.

Solubility Data

Solubility (In Vitro)

DMSO: 100 mg/mL (162.72 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 1.67 mg/mL (2.72 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 16.7 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: ≥ 1.67 mg/mL (2.72 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 16.7 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	1.6272 mL	8.1359 mL	16.2718 mL
	5 mM	0.3254 mL	1.6272 mL	3.2544 mL
	10 mM	0.1627 mL	0.8136 mL	1.6272 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.