NMDA receptor

Mice with learning impairments caused by Aβ 25–35 exhibit anti-amnesic effects when given fluoroethylnormemantine (0.1–10 mg/kg; a single ip)[1]. In mice, Aβ 25-35-induced behavioral impairments, neuroinflammation, oxidative stress, apoptosis, and cell death are attenuated by fluoroethylnormemantine (0.1–10 mg/kg; ip once daily for 7 days)[1]. In rats, fluoroethylnormemantine (1–20 mg/kg; single injection) lowers fear behavior in cued fear conditioning (FC) and extinction training, as well as behavioral despair in the forced swim test (FST)[2].

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Given after stress, FluoroethylnormemantineFENM decreased behavioral despair and reduced perseverative behavior. When administered after re-exposure, FENM facilitated extinction learning. As a prophylactic, FENM attenuated learned fear and decreased stress-induced behavioral despair. FENM was behaviorally effective in both male and female mice. (R,S)-ketamine, but not FENM, increased expression of c-fos in vCA3. Both (R,S)-ketamine and FENM attenuated large-amplitude AMPA receptor–mediated bursts in vCA3, indicating a common neurobiological mechanism for further study. Conclusions Our results indicate that Fluoroethylnormemantine/FENM is a novel drug that is efficacious when administered at various times before or after stress. Future work will further characterize FENM’s mechanism of action with the goal of clinical development. [1]

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Results: Unlike memantine, Fluoroethylnormemantine/FENM did not produce nonspecific side effects and did not alter sensorimotor gating or locomotion. FENM decreased immobility in the forced swim test. Moreover, FENM robustly facilitated fear extinction learning when administered prior to either cued fear conditioning training or tone reexposure. Conclusions: These results suggest that FENM is a promising, novel compound that robustly reduces fear behavior and may be useful for further preclinical testing. [2]

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Results: Both Memantine and FluoroethylnormemantineFENM showed symptomatic anti-amnesic effects in Aβ 25-35-treated mice. Interestingly, FENM was not amnesic when tested alone at 10 mg/kg, contrarily to Memantine. Drugs injected once per day prevented Aβ 25-35-induced memory deficits, oxidative stress (lipid peroxidation, cytochrome c release), inflammation (interleukin-6, tumor necrosis factor-α increases; glial fibrillary acidic protein and Iba1 immunoreactivity in the hippocampus and cortex), and apoptosis and cell loss (Bcl-2-associated X/B-cell lymphoma 2 ratio; cell loss in the hippocampus CA1 area). However, FENM effects were more robust than observed with Memantine, with significant attenuations vs the Aβ 25-35-treated group. Conclusions: FENM therefore appeared as a potent neuroprotective drug in an AD model, with a superior efficacy compared with Memantine and an absence of direct amnesic effect at higher doses. These results open the possibility to use the compound at more relevant dosages than those actually proposed in Memantine treatment for AD [3].

Animal/Disease Models: Male Swiss CD-1 mice (7-9 weeks) were injected with Aβ25-35[1]
Doses: 0.1, 0.3, 1, 3, 10 mg/kg
Route of Administration: Ip 30 minutes before the behavioral tests
Experimental Results: Attenuated Aβ 25-35-induced spontaneous alternation deficit, passive avoidance deficit, and novel object exploration deficit.

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Fluoroethylnormemantine: FENM/Fluoroethylnormemantine was administered in a single dose at 10, 20, or 30 mg/kg of body weight. Saline, memantine (10 mg/kg), (R,S)-ketamine (30 mg/kg), or FENM (10, 20, or 30 mg/kg) was administered before or after contextual fear conditioning in 129S6/SvEv mice. Drug efficacy was assayed using various behavioral tests. Protein expression in the hippocampus was quantified with immunohistochemistry or Western blotting. In vitro radioligand binding was used to assay drug binding affinity. Patch clamp electrophysiology was used to determine the effect of drug administration on glutamatergic activity in ventral hippocampal cornu ammonis 3 (vCA3) 1 week after injection. [1]

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Researchers administered saline, FENM, or memantine prior to a number of behavioral assays, including paired-pulse inhibition, open field, light dark test, forced swim test, and cued fear conditioning in male Wistar rats. FENM/Fluoroethylnormemantine was administered in a single dose at 1, 3, 5, 10, or 20 mg/kg of body weight. [2]

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Swiss mice were treated intracerebroventricularly with aggregated Aβ 25-35 peptide and examined after 1 week in a battery of memory tests (spontaneous alternation, passive avoidance, object recognition, place learning in the water-maze, topographic memory in the Hamlet). Toxicity induced in the mouse hippocampus or cortex was analyzed biochemically or morphologically.
Researchers examined 2 effects of the drugs/Fluoroethylnormemantine. First, symptomatic effects were analyzed in Aβ 25-35-treated mice by injecting the drugs just before the behavioral tests. Second, the neuroprotection was analyzed by repeatedly o.d. injecting the mice for 1 week starting on the day of peptide injection. For symptomatic effects, drugs were injected only on day 8 after Aβ 25–35 injection, 30 minutes before the behavioral tests: spontaneous alternation, passive avoidance training, session 2 of the object recognition test or each water-maze training sessions (supplementary Figure 1a). A group was tested for spontaneous alternation, passive avoidance, and object recognition in series. As Memantine, and expectedly Fluoroethylnormemantine/FENM, has a short half-life in mice (<2 hours; Beconi et al., 2011), all the drug was excreted overnight. A separate group was trained in the Hamlet before Aβ 25–35 injection to assess topographic memory (supplementary Figure 1b). For neuroprotective effects, drugs were injected o.d. from day 1 to day 7 after Aβ 25–35 injection (supplementary Figure 1c), and mice were tested for spontaneous alternation, passive avoidance, and object recognition in series. They were killed at day 13 for immunochemistry (group A). A group of mice performed place learning in the water-maze, then were killed at day 16 and used for biochemical assays (group B). An additional series (group C) included mice killed at day 5 after Aβ 25–35 peptide injection and daily drug injections for assessing cytokine levels by enzyme-linked immuno-sorbent assays (ELISA).[3]

Recently, a novel NMDAR antagonist, Fluoroethylnormemantine (FENM), was derived from the NMDAR antagonist memantine. To determine its biodistribution and safety profile, FENM was developed into the radiolabeled compound [18F]-FENM. In rats, [18F]-FENM (44 ± 11 MBq) stabilized in the brain 40 minutes after injection, with 0.4% of the injected dose found in the brain. In rats anesthetized with isoflurane before [18F]-FENM injection, combined ex vivo autoradiography and immunohistochemical staining demonstrated a strong colocalization of NMDARs and [18F]-FENM binding, particularly in cortical regions and the hippocampus (HPC). Most interestingly, if rats were anesthetized with (R,S)-ketamine (80 mg/kg) immediately before [18F]-FENM injection, the [18F]-FENM autoradiographic signal no longer correlated with NMDAR staining, indicating that binding was disabled or blocked. [18F]-FENM was calculated to have a Ki of 3.5 μM, compared with (R,S)-ketamine, which exhibits a Ki of 0.53 μM. Furthermore, FENM was recently shown to facilitate extinction learning in male rats without altering sensorimotor behavior. However, it remains unknown whether FENM can be efficacious as a prophylactic or antidepressant. [1]

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Recently, a novel radiolabeled compound, [18F]- Fluoroethylnormemantine (FENM), was derived from memantine as a novel positron emission tomography (PET) tracer (Salabert et al., 2015, 2018). With a Ki of 3.510-6 M and high lipophilicity (logD = 1.93), [18F]-FENM stabilized 40 minutes after injection with approximately 0.4% of the original dose present in the brain. Combined ex vivo autoradiography and immunohistochemistry indicated that [18F]-FENM strongly colocalizes with NMDARs in the cortex and cerebellum. Intriguingly, this colocalization is blocked by injection of (R,S)-ketamine, suggesting that FENM exhibits a lower affinity to the NMDAR receptor than (R,S)-ketamine. Moreover, because both compounds bind to phencyclidine sites in the NMDAR channel pore, these data suggest that the compounds may also exert similar behavioral effects. However, while the antidepressant-like effects of FENM remain unknown, recent data indicate that FENM enhances cognitive function and exerts neuroprotective properties in a mouse model of AD (Couly et al., 2020). In this study, Couly and colleagues found that FENM reversed deficits in long-term memory, navigation, and place learning, and object recognition in a pharmacological model of AD. Interestingly, compared with the effects of memantine, the authors showed that FENM improved spatiotemporal orientation in the Hamlet test while memantine did not affect behavior. FENM’s behavioral actions were found to correspond with a reduction in inflammatory cytokines and neuronal cell loss in the hippocampus. Thus, although FENM shows potential for enhancing cognition and protecting age-related brain impairments, it is still unknown whether the drug can reverse stress-related maladaptive behaviors.[2]

[1]. Anti-Amnesic and Neuroprotective Effects of Fluoroethylnormemantine in a Pharmacological Mouse Model of Alzheimer's Disease. Int J Neuropsychopharmacol. 2021 Feb 15;24(2):142-157.

[2]. Fluoroethylnormemantine, a novel derivative of memantine, facilitates extinction learning without sensorimotor deficits. Int J Neuropsychopharmacol. 2021 Jul 14;24(6):519-531.

[3]. Fluoroethylnormemantine, a novel NMDA receptor antagonist, for the prevention and treatment of stress-induced maladaptive behavior. Biological Psychiatry. 2021 Feb 15;24(2):142-157.

Here, we characterized FENM, an NMDAR antagonist with antidepressant and prophylactic efficacy. We found that 1) FENM exhibits antidepressant-like properties in male and female mice exposed to stress, 2) FENM suppresses hyponeophagia in nonstressed male mice, 3) FENM attenuates fear when administered after extinction in male mice, 4) FENM is prophylactic when administered 1 week before exposure to stress in both sexes, and 5) FENM reduces large-amplitude AMPAR-mediated bursts in hippocampal vCA3 1 week after administration. [1]

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In this study, we aimed to test the behavioral effects of FENM administration on sensorimotor gating, avoidance behavior, behavioral despair, and fear behavior in rats. Our experiments show that FENM can effectively reduce behavioral despair as well as facilitate extinction learning without altering locomotion or sensorimotor behavior. These results show that FENM exerts antidepressant-like and fear-attenuating properties. FENM was also effective in attenuating learned fear when administered acutely prior to FC, indicating that it may also be leveraged as a resilience-enhancing prophylactic. Together, our findings indicate that FENM, a novel NMDAR antagonist, may be a suitable candidate for further preclinical testing in a variety of stress-related behavioral assays. [2]

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FENM appeared as a promising drug. Its effect must now be confirmed in transgenic mouse models of AD. Only repeated administration regimens in these chronic models will allow to determine if FENM is able to decrease the amyloid load and plaque formation in amyloid-based models or kinases activities and neurofibrillary tangles formation in tau-based models. This was previously described for Memantine (Wang et al., 2015) and several other drugs with similar symptomatic and neuroprotective profiles. The strength of FENM-induced neuroprotection must be investigated in similar transgenic models in the future, in parallel to the analysis of the drug mechanism of action, to establish the superiority of the molecule over Memantine and to determine whether the drug is a putative candidate for synergic combinations with current drugs under development. In conclusion, we described the symptomatic and neuroprotective efficacy of a novel Memantine derivative, FENM, in a pharmacological mouse model of AD. Comparison with its parent molecule revealed that FENM is more effective in preventing oxidative stress, apoptosis, and neuroinflammation and suggested that the molecule may not only be used as a potent PET radiotracer for NMDAR but also as a promising neuroprotective drug in AD. Moreover, the compound may be used at more relevant dosages than those actually proposed with the Memantine treatment. [3]

C12H20FN

197.29

197.157

C, 73.05; H, 10.22; F, 9.63; N, 7.10

1639210-26-6

Fluoroethylnormemantine hydrochloride;1639210-25-5

170907856

Colorless to light yellow ointment

3.2

1

2

2

14

237

2

C1[C@@H]2CC3(C[C@H]1CC(C2)(C3)N)CCF

HUYVZSFADWYSHD-ZYANWLCNSA-N

InChI=1S/C12H20FN/c13-2-1-11-4-9-3-10(5-11)7-12(14,6-9)8-11/h9-10H,1-8,14H2/t9-,10+,11?,12?

(5S,7R)-3-(2-fluoroethyl)adamantan-1-amine

Fluoroethylnormemantine; SCHEMBL16274382;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

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

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.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*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).

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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)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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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).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)