DREADD agonist 21 is a potent agonist (EC50=1.7 nM) of the human muscarinic acetylcholine M3 receptor (hM3Dq).

Physicochemical Properties

Molecular Formula	C17H18N4
Molecular Weight	278.35
Exact Mass	278.153
Elemental Analysis	C, 73.35; H, 6.52; N, 20.13
CAS #	56296-18-5
Related CAS #	DREADD agonist 21 dihydrochloride; 2250025-92-2
PubChem CID	11818276
Appearance	Light yellow to yellow solid powder
Density	1.3±0.1 g/cm3
Boiling Point	470.1±55.0 °C at 760 mmHg
Flash Point	238.1±31.5 °C
Vapour Pressure	0.0±1.2 mmHg at 25°C
Index of Refraction	1.699
LogP	0.98
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	3
Rotatable Bond Count	1
Heavy Atom Count	21
Complexity	388
Defined Atom Stereocenter Count	0
SMILES	N1(C2C3=C([H])C([H])=C([H])C([H])=C3N([H])C3=C([H])C([H])=C([H])C([H])=C3N=2)C([H])([H])C([H])([H])N([H])C([H])([H])C1([H])[H]
InChi Key	JCBYXNSOLUVGTF-UHFFFAOYSA-N
InChi Code	InChI=1S/C17H18N4/c1-2-6-14-13(5-1)17(21-11-9-18-10-12-21)20-16-8-4-3-7-15(16)19-14/h1-8,18-19H,9-12H2
Chemical Name	6-piperazin-1-yl-11H-benzo[b][1,4]benzodiazepine
Synonyms	DREADD-agonist-21; DREADD agonist 21; DREADD agonist-21
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	hM3Dq ( EC50 = 1.7 nM )
ln Vitro	DREADD Agonist 21 is a potent human muscarinic cardiac M3 receptor agonist (hM3Dq) agonist (EC50=1.7 nM) that does not activate the human M3 receptor (hM3). In addition to being inactive against hM3, DREADD agonist 21 is a potent full agonist of hM3Dq (EC50=1.7 nM). It is only 3.5-fold more selective for hM3Dq than H1 and 40-fold more selective than 5HT2A. 100 times more selective for 5HT2C than α1A. DREADD Agonist 21 has high affinity for 5HT2A and 5HT2C serotonin uptake, α1A initiator uptake and H1 histamine uptake, with Ki values of 66, 170, 280 and 6 nM respectively. [1]. DREADD agonist 21 potently activates hM1Dq, hM3Dq and hM4Di. DREADD agonist 21 binds hM1, hM4, hM1Dq, and hM4Di with pKi of 5.97, 5.44, 7.20, and 6.75, respectively. DREADD agonist 21 effectively activated hM3Dq in Chinese mouse hamster or (CHO) cells transfected cells in vitro, with a pEC50 of 8.48±0.05. DREADD Agonist 21 is a highly selective and potent muscarinic DREADD agonist (pEC50 for hM1Dq=6.54 and hM4Di=7.77 in pERK assay) [2].
ln Vivo	DREADD agonist 21 (0.3, 1.0 and 3.0 mg/kg; ip) activates neuronal hM3Dq in mice[2]. DREADD agonist 21 has excellent bioavailability, pharmacokinetic properties and brain penetration. 21 (0.1, 1 and 10 mg/kg; ip) exhibits 95.1% rabbit protein binding and 95% brain protein binding in mice [2].
References	[1]. The first structure-activity relationship studies for designer receptors exclusively activated by designer drugs. ACS Chem Neurosci. 2015 Mar 18;6(3):476-84. [2]. DREADD Agonist 21 Is an Effective Agonist for Muscarinic-Based DREADDs in Vitro and in Vivo. ACS Pharmacol Transl Sci. 2018 Sep 14;1(1):61-72.

Solubility Data

Solubility (In Vitro)

DMSO: ~56 mg/mL (~201.2 mM) Ethanol: ~56 mg/mL

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (8.98 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 (8.98 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.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.5926 mL	17.9630 mL	35.9260 mL
	5 mM	0.7185 mL	3.5926 mL	7.1852 mL
	10 mM	0.3593 mL	1.7963 mL	3.5926 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.