AR-M 1000390 hydrochloride (the hydrochloric acid of ARM-390; AR-M1000390) is a potent, highly selective agonist of δ opioid receptor that can induce insulin depletion in the rat and RINm5F cells. The δ opioid receptor was activated with an EC50 of 7.2±0.9 nM. After seven days of administration, AR-M100390 (600 micromol/kg) produced vacuolation in the rat pancreatic beta-cell, which was linked to insulin deficiency and hyperglycemia. The specific inhibition of rat insulin2 mRNA transcription in vivo was the cause of the insulin loss caused by AR-M100390. Hyperglycemia and insulin deficiency were reversible. Using the rat pancreatic beta-cell line RINm5F, the effects of AR-M100390 were replicated. This compound inhibited intracellular insulin content and secretion without compromising cell survival. Insulin loss in vitro was also reversible and caused by specific inhibition of insulin2 mRNA transcription. It is possible that the effects were not mediated by the delta-opioid receptor because pretreatment of cells with the pertussis toxin or the delta-opioid antagonist naltrindole did not reverse the loss of insulin in AR-M100390-treated cells. In RINm5F cells, AR-M100390 inhibited KCl-mediated calcium mobilization, indicating that L-type calcium channels present in these cells as well as in pancreatic beta-cells may contribute to this compound's partial inhibition of insulin secretion. In conclusion, research conducted both in vivo and in vitro indicates that AR-M100390'sinhibitionof insulin is caused by a combination of inhibitions of insulin release and/or synthesis.

Physicochemical Properties

Molecular Formula	C23H29CLN2O
Molecular Weight	384.95
Exact Mass	384.196
Elemental Analysis	C, 71.76; H, 7.59; Cl, 9.21; N, 7.28; O, 4.16
CAS #	209808-47-9
Related CAS #	209808-47-9 (HCl);
PubChem CID	76848958
Appearance	White to off-white solid powder
LogP	5.484
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	2
Rotatable Bond Count	5
Heavy Atom Count	27
Complexity	472
Defined Atom Stereocenter Count	0
SMILES	O=C(N(CC)CC)C1=CC=C(/C(C2=CC=CC=C2)=C3CCNCC/3)C=C1.Cl
InChi Key	OTXTZCLQEGSAMW-UHFFFAOYSA-N
InChi Code	InChI=1S/C23H28N2O.ClH/c1-3-25(4-2)23(26)21-12-10-19(11-13-21)22(18-8-6-5-7-9-18)20-14-16-24-17-15-20;/h5-13,24H,3-4,14-17H2,1-2H3;1H
Chemical Name	N,N-diethyl-4-[phenyl(piperidin-4-ylidene)methyl]benzamide;hydrochloride
Synonyms	ARM-390 HCl; AR M1000390; ARM 390; ARM1000390 HCl; ARM390; AR-M1000390; ARM-390 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	δ opioid receptor ( EC50 = 7.2±0.9 nM )
ln Vitro	In vitro activity: AR-M 1000390 (Compound 6a) demonstrates a very high degree of selectivity over both the ε and δ opioid receptors (IC50=7470±606 nM and 3800±172 nM, respectively), with binding affinities (IC50) for the δ opioid receptor of 0.87±0.23 nM[1]. Insulin levels secreted and intracellular are measured after 16–24 hours of treatment with AR-M 1000390 (AR-M100390) and Cyclizine in RINm5F cells. With a maximal inhibition of approximately 90% at the highest concentration tested (10 μM), AR-M 1000390 mediates a dose-dependent decrease in insulin content[2].
ln Vivo	Rats are given 5, 100, and 600 μmol/kg of AR-M 1000390 (AR-M100390) for three or seven days; another group of rats is given 600 μmol/kg of the compound and is given fourteen days to recuperate. After seven days of dosing, the rat pancreatic β-cell experiences vacuolation due to AR-M 1000390 (600 μmol/kg), which is linked to insulin depletion and hyperglycemia. Rat pancreatic β-cell vacuolation is induced by 600 μmol/kg of AR-M 1000390, and this vacuolation is comparable to that observed with cyclizine and cyproheptadine[2].
Enzyme Assay	AR-M 1000390 hydrochloride (the hydrochloric acid of ARM-390) is a potent, highly selective δ opioid receptor agonist with an EC50 of 7.2±0.9 nM.
Cell Assay	RINm5F cells are cultivated in 24-well plates and subjected to serum-free medium containing vehicle (DMSO), 10 μM AR-M 1000390 (AR-M100390), and 10 μM Cyclizine. Following treatment, the cells are washed with phosphate-buffered saline and kept at -80°C until further examination. The RNeasy purification system is used to isolate RNA after treating it with DNAse[2].
Animal Protocol	Rats: AR-M 1000390 (AR-M100390) or vehicle (saline) is administered to six Han Wistar rats per treatment group for seven days in doses of 5, 100, or 600 μmol/kg/day. Fourteen days are spent in recovery after a seven-day treatment with 600 μmol/kg/day in a different group of rats. Three days are spent administering 600 μmol/kg/day to a different group. Day 2, 4, 8, and 22 blood samples are collected for measurements of insulin, lipids, and glucose. Days 4 and 8 see the collection of blood samples for measurements of the AR-M 1000390 concentration. The pancreas is removed and processed for histopathology, insulin immunohistochemistry, and insulin mRNA analyses after the animals are put to sleep with CO2 on days 4, 8, and 22[2].
References	[1]. N,N-Diethyl-4-(phenylpiperidin-4-ylidenemethyl)benzamide: a novel, exceptionally selective, potent delta opioid receptor agonist with oral bioavailability and its analogues. J Med Chem. 2000 Oct 19;43(21):3895-905. [2]. Mechanistic investigation of N,N-diethyl-4-(phenyl-piperidin-4-ylidenemethyl)-benzamide-inducedinsulin depletion in the rat and RINm5F cells. Toxicol Sci. 2008 Sep;105(1):221-9.

Solubility Data

Solubility (In Vitro)

DMSO: ≥ 150 mg/mL (390 mM) Water: N/A Ethanol: N/A

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (6.49 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 (6.49 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 (6.49 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	2.5977 mL	12.9887 mL	25.9774 mL
	5 mM	0.5195 mL	2.5977 mL	5.1955 mL
	10 mM	0.2598 mL	1.2989 mL	2.5977 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.