| Size | Price | |
|---|---|---|
| 500mg | ||
| 1g | ||
| Other Sizes |
| Targets |
Amitifadine primarily targets the human serotonin transporter (SERT), norepinephrine transporter (NET), and dopamine transporter (DAT), functioning as a “triple” reuptake inhibitor (SNDRI) with a serotonin‑preferring profile. Its inhibition potencies (IC₅₀) for the uptake of the three monoamines in HEK‑293 cells expressing human recombinant transporters are 12 nM for serotonin, 23 nM for norepinephrine, and 96 nM for dopamine. The corresponding binding affinities (Kᵢ) at the transporters are 99 nM for SERT, 262 nM for NET, and 213 nM for DAT. The in vivo pharmacological ratio for monoamine elevation is approximately 1:2:8.
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| ln Vitro |
Amitifadine is a potent inhibitor of human SERT, NET, and DAT in vitro. Using HEK‑293 cells stably expressing the human recombinant transporters, the compound exhibits IC₅₀ values of 12 nM for inhibiting serotonin uptake, 23 nM for norepinephrine uptake, and 96 nM for dopamine uptake. The binding affinities (Kᵢ) to the three transporters are 99 nM (SERT), 262 nM (NET), and 213 nM (DAT). This profile establishes amitifadine as a serotonin‑preferring triple reuptake inhibitor, with a potency ratio for monoamine uptake inhibition of approximately 1:2:8 (5‑HT:NE:DA). The major metabolite of amitifadine, the 2‑lactam compound, only weakly inhibits monoamine uptake, suggesting that the parent compound is the primary active moiety.
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| ln Vivo |
Amitifadine (10 mg/kg, i.p.) produces a marked and persistent increase in extracellular concentrations of serotonin, norepinephrine, and dopamine in the prefrontal cortex of freely moving rats, as measured by in vivo microdialysis. It also elevates extracellular dopamine levels in the striatum and in the nucleus accumbens, a core region of the brain’s reward circuitry. Consistent with its reuptake inhibition mechanism, amitifadine also significantly decreases the extracellular levels of monoamine metabolites (5‑HIAA, DOPAC, HVA) in these brain regions. Importantly, across a broad dose range, amitifadine does not increase locomotor activity or induce stereotypical behaviors typically associated with psychostimulants, which is an advantage for a compound that enhances dopamine transmission. A 10 mg/kg dose of amitifadine significantly reduced nicotine self‑administration in rats, an effect that persisted over two weeks of chronic treatment, suggesting potential for smoking‑cessation therapy.
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| Enzyme Assay |
The in vitro potency of amitifadine is determined using radioligand uptake inhibition assays in HEK‑293 cells expressing human recombinant SERT, NET, or DAT. Cells are plated in multi‑well plates and pre‑incubated with varying concentrations of the test compound. The uptake reaction is initiated by adding a low concentration (nanomolar range) of the respective radiolabeled neurotransmitter substrate ([³H]5‑HT for SERT, [³H]NE for NET, or [³H]DA for DAT). After a brief incubation (typically 5‑15 minutes) at 37 °C, the reaction is terminated by rapid aspiration and washing with ice‑cold buffer. The cells are then lysed, and the accumulated radioactivity in the lysate is quantified by liquid scintillation counting. Non‑specific uptake is determined in the presence of a high concentration of a specific reuptake inhibitor (e.g., fluoxetine for SERT, desipramine for NET, or GBR‑12909 for DAT). The percentage of inhibition at each compound concentration is calculated, and IC₅₀ values are derived by non‑linear regression analysis of the concentration‑inhibition curves.
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| Cell Assay |
Cell‑based assays for amitifadine utilize the same HEK‑293 cell lines stably expressing the human recombinant SERT, NET, or DAT as described in the enzyme assay section. These functional uptake assays are specifically designed to measure the compound’s ability to block the transporter‑mediated influx of radiolabeled neurotransmitters into living cells, which is the primary pharmacological action of the molecule. The protocol involves exposing the cells to the test compound and the radiolabeled substrate simultaneously. After the uptake period, the reaction is stopped by rapid washing to remove extracellular radioactivity, and the cells are lysed. The intracellular radioactivity (a direct measure of uptake) is then quantified by liquid scintillation counting. This method directly measures the functional antagonism of the transporters.
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| Animal Protocol |
Rat in Vivo Microdialysis: To assess the neurochemical effects of amitifadine, male Sprague‑Dawley rats are anesthetized, and guide cannulae are stereotaxically implanted into target brain regions (e.g., prefrontal cortex, striatum, or nucleus accumbens). After recovery, a microdialysis probe is inserted, and the animal is perfused with artificial cerebrospinal fluid (aCSF) at a constant flow rate (e.g., 1.5 μL/min). Following a stabilization period, baseline dialysate samples are collected every 20 minutes. Amitifadine is then administered intraperitoneally (i.p.) at a dose of 10 mg/kg, and sample collection continues for several hours. The concentrations of monoamines (5‑HT, NE, DA) and their metabolites (5‑HIAA, DOPAC, HVA) in the dialysate are analyzed by high‑performance liquid chromatography (HPLC) with electrochemical detection (ECD). Locomotor Activity Test: Sprague‑Dawley rats are placed in photocell‑equipped activity cages, and baseline locomotor activity is recorded. After a habituation period, amitifadine is administered i.p., and ambulatory counts are recorded to assess any stimulant or depressant effects on spontaneous motor activity over a broad dose range.
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| ADME/Pharmacokinetics |
In preclinical studies, amitifadine is rapidly absorbed following oral administration, with a plasma t_max of 0.7-1.2 hours. The elimination half-life (t₁/₂) is approximately 3.3-4.4 hours. Both C_max and AUC values increase in a dose-proportional manner across the dose range studied (5-150 mg). At a single dose of 10 mg, the average C_max was 78 ng/mL, and doses above this level result in plasma concentrations that exceed the in vitro IC₅₀ values for monoamine uptake inhibition. The compound shows no remarkable accumulation following 10 days of once‑daily dosing, with steady‑state being achieved within approximately one week. In humans, a single 8 mg dose of amitifadine hydrochloride produced a C_max of 2.02 ng/mL (18.4% CV) with a median T_max of 10 hours, and a mean half-life of 61.3 hours (34.2% CV).
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| Toxicity/Toxicokinetics |
In clinical trials for MDD, amitifadine has been shown to be safe and well‑tolerated. In a phase II proof‑of‑concept trial, the estimated effect size (Cohen‘s d) for amitifadine compared to placebo was −0.601 on the MADRS scale, indicating clinically meaningful antidepressant activity. Notably, amitifadine was well tolerated without causing weight gain or sexual dysfunction, which are common adverse effects limiting adherence to standard first‑line antidepressants. In a comprehensive phase IIb/IIIa safety and efficacy trial (TRIADE study) involving a total safety population of 342 patients with treatment‑resistant MDD, amitifadine demonstrated a favorable safety profile, with no significant safety signals concerning cardiovascular events, hepatotoxicity, or serious adverse events reported. The most common adverse events, if any, were likely mild and transient, consistent with its mechanism of action.
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| References |
[1]. Antidepressant-like actions of DOV 21,947: a "triple" reuptake inhibitor. Eur J Pharmacol. 2003 Feb 14;461(2-3):99-104.
[2]. Amitifadine, a triple monoamine re-uptake inhibitor, reduces nicotine self-administration in female rats. Eur J Pharmacol. 2015 Jun 20;764:30-37. |
| Additional Infomation |
Drug Indication
DOV 21947 has been studied for the treatment of depression. Mechanism of Action DOV 21947 possesses antidepressant-like properties and potent analgesic activity. It inhibits the reuptake of three neurotransmitters closely associated with depression—serotonin, norepinephrine, and dopamine—therefore its overall efficacy may be superior to currently available antidepressants. Amitifadine is part of the 3-azabicyclo[3.1.0]hexane class and is structurally related to the prototype triple reuptake inhibitor DOV-216,303. The synthesis of amitifadine has been described in several patents and involves a stereospecific route using (S)-epichlorohydrin to establish the cis-cyclopropane stereochemistry. The discovery that a triple reuptake inhibitor might improve outcomes in SSRI‑nonresponsive patients originated from the National Institute of Mental Health’s large‑scale STAR*D (Sequenced Treatment Alternatives to Relieve Depression) study, where combining serotonergic, noradrenergic, and dopaminergic mechanisms was associated with improved outcomes. The advanced phase IIb/IIIa TRIADE trial employed a novel clinical trial design, the Sequential Parallel Comparison Design (SPCD), to minimize the high placebo response rate that has plagued many depression trials, and also used the SAFER criteria to ensure diagnostic and treatment‑history accuracy. A previous phase II study in MDD patients indicated that the effect size for amitifadine was superior to that observed for citalopram, an active comparator, in the same trial. Beyond depression, preclinical evidence suggests amitifadine may have potential as a smoking‑cessation agent, as it significantly reduced nicotine self‑administration in an animal model. |
| Molecular Formula |
C11H11NCL2
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|---|---|
| Molecular Weight |
228.118
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| Exact Mass |
227.026
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| Elemental Analysis |
C, 57.92; H, 4.86; Cl, 31.08; N, 6.14
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| CAS # |
410074-73-6
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| Related CAS # |
410074-74-7 (HCl);410074-73-6;
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| PubChem CID |
11658655
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| Appearance |
Typically exists as solid at room temperature
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| Density |
1.4±0.1 g/cm3
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| Boiling Point |
324.4±37.0 °C at 760 mmHg
|
| Flash Point |
150.0±26.5 °C
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| Vapour Pressure |
0.0±0.7 mmHg at 25°C
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| Index of Refraction |
1.622
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| LogP |
2.57
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| Hydrogen Bond Donor Count |
1
|
| Hydrogen Bond Acceptor Count |
1
|
| Rotatable Bond Count |
1
|
| Heavy Atom Count |
14
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| Complexity |
245
|
| Defined Atom Stereocenter Count |
2
|
| SMILES |
C1=CC(=C(C=C1[C@]23C[C@@H]3CNC2)Cl)Cl
|
| InChi Key |
BSMNRYCSBFHEMQ-KCJUWKMLSA-N
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| InChi Code |
InChI=1S/C11H11Cl2N/c12-9-2-1-7(3-10(9)13)11-4-8(11)5-14-6-11/h1-3,8,14H,4-6H2/t8-,11+/m1/s1
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| Chemical Name |
(1R,5S)-1-(3,4-dichlorophenyl)-3-azabicyclo[3.1.0]hexane
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| Synonyms |
(-)-Amitifadine; DOV-21,947; EB-1010; DOV-21947; EB 1010; DOV21,947; EB1010; 8WR4Q5ZGU3; DOV-102677; DOV-102,677; 500733-69-7
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| HS Tariff Code |
2934.99.9001
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| 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)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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|---|---|
| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 4.3837 mL | 21.9183 mL | 43.8366 mL | |
| 5 mM | 0.8767 mL | 4.3837 mL | 8.7673 mL | |
| 10 mM | 0.4384 mL | 2.1918 mL | 4.3837 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.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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