NMDA Receptor
NMDA (N-methyl-D-aspartic acid) receptor glycine modulatory site. Cycloleucine is a selective NMDA receptor antagonist that targets the glycine site. It also inhibits S-adenosyl-methionine-mediated methylation and acts as a competitive inhibitor of ATP: L-methionine-S-adenosyl transferase.

Internal methylation of viral RNA in B77 transformed chick embryo fibroblasts is blocked by cycloleucine (4–40 mM; 3 h)[5]. More than 90% of B77 38S RNA subunits are prevented from forming the penultimate Gm and m6A by cycloleucine (40 mM; 24 h)[5]. ?The viability of human KB and mouse L1210s leukemia cell lines is inhibited by cytostatic (10 μg/mL)[5].

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Cycloleucine is a selective NMDA receptor antagonist that targets the glycine modulatory site. It acts as a competitive antagonist at this site with a Ki of 600 μM. In electrophysiological studies, it inhibited NMDA-evoked currents with an IC50 of 24 μM in the presence of 1 μM glycine. The inhibition was partially surmountable by glycine, indicating both competitive and non-competitive components. Cycloleucine did not affect kainic acid- or quisqualic acid-evoked currents, demonstrating its selectivity for the NMDA receptor. It also inhibits S-adenosyl-methionine-mediated methylation and acts as a competitive inhibitor of ATP: L-methionine-S-adenosyl transferase.

In rats, cycloleucine (0.5–4 μg; intracerebroventricular injection) prolongs open arm entry, increases open arm duration, and causes severe arrivals[3]. In mice infected with the Semliki Forest virus (SFV) strain A7(74) and normal mice, cycloleucine lowers the weights of the spleen and thymus[6].

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In vivo, cycloleucine has been shown to produce a brief depression of self-stimulation in animal models, but only after relatively large doses which were not antagonized by D-cycloserine. As an NMDA receptor antagonist, it may modulate glutamatergic neurotransmission and affect learning, memory, and other cognitive processes. However, detailed in vivo efficacy data are limited.

In vitro receptor binding assays for cycloleucine are performed to evaluate its affinity for the NMDA receptor glycine site. Radioligand binding studies using membrane preparations from brain tissue or cells expressing NMDA receptors are conducted. The compound's ability to displace a specific radiolabeled glycine site ligand, such as [3H]-MDL-105,519 or [3H]-glycine, is measured to calculate its Ki value (600 μM). Functional assays, such as electrophysiological recordings or calcium mobilization assays, are used to confirm its antagonist activity.

In vitro cell-based assays are conducted using cultured hippocampal neurons or cells expressing recombinant NMDA receptors. The cells are treated with NMDA and glycine in the presence or absence of varying concentrations of cycloleucine. The inhibition of NMDA-evoked currents is measured using whole-cell voltage-clamp techniques. The IC50 for the inhibition is determined from the concentration-response curves at different glycine concentrations. These assays confirm the compound's activity as a glycine site antagonist.

Animal/Disease Models: Male rats bilaterally cannulated into the nucleus accumbens septi (NAS)[3]
Doses: 1 µL of 0.5, 1.0, 2.0, 4 µg/µL
Route of Administration: Intracerebroventrical injection
Experimental Results: Increased time spent in the open arms and extreme arrivals at all doses. Increased open arm entries at the dose of 4 μg.

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In vivo animal studies for cycloleucine are conducted to evaluate its effects on NMDA receptor-mediated behaviors. The compound is typically administered via intraperitoneal or intracerebroventricular injection. Its effects on learning and memory, seizure susceptibility, and self-stimulation behaviors are assessed in rodent models. Dosing regimens are determined based on preliminary dose-response studies.

Absorption, Distribution and Excretion
Fifteen minutes after intravenous injection of [(14)C]-1-aminocyclopentanecarboxylic acid in rhesus monkeys, the (14)C levels in the liver and pancreas were 8 and 2 times higher than in the blood, respectively. Twenty-four hours later, the (14)C levels in these tissues were similar to those in the blood. When mice were administered cycloleucine, the accumulation levels of cycloleucine in tissues ranged from 0.02 to 1.29 mg/ml. The remaining 2% was largely bound to proteins. The drug concentration gradient showed accumulation in the kidneys, with higher accumulation in the spleen and pancreas. The distribution ratios in central nervous system tissues, kidneys, and spleen did not change with plasma cycloleucine concentrations or time from 4 to 40 days after administration. Five days after administration of cycloleucine to mice, the liver cycloleucine distribution ratio in the lowest dose group was significantly greater than 1 and changed with increasing plasma cycloleucine concentrations.

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Biological Half-Life
At doses of 0.4–0.5 mg/g, plasma concentrations on day 24 were not significantly different from those on day 1. At the highest doses (1–3 mg/g), plasma concentrations were almost identical on day 4. The plasma half-life is extremely long.

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No detailed pharmacokinetic data are publicly available for cycloleucine. As a small, polar amino acid analog, its oral bioavailability is likely limited, and it may not readily cross the blood-brain barrier. The compound's pharmacokinetic properties would be studied in preclinical models to guide in vivo experiments if needed.

No specific toxicity data are publicly available for cycloleucine. As an NMDA receptor antagonist, its toxicity profile would be expected to be related to its mechanism of action. Excessive NMDA receptor inhibition can impair synaptic plasticity and cognitive function. At high doses, it may cause psychotomimetic effects or other central nervous system side effects.

[1]. 1-Aminocyclobutane-1-carboxylate (ACBC): a specific antagonist of the N-methyl-D-aspartate receptor coupled glycine receptor. Eur J Pharmacol. 1989 Feb 28;161(2-3):281-2.

[2]. RNA methylation and control of eukaryotic RNA biosynthesis. Effects of cycloleucine, a specific inhibitor of methylation, on ribosomal RNA maturation. Eur J Biochem. 1977 Mar 15;74(1):19-29.

[3]. Effects of Cycloleucine in the Nucleus Accumbens Septi on the Elevated plus Maze Test in Rats. Neuropsychobiology. 2020;79(3):191-197.

[4]. Cytostatic activity in vitro of cycloleucine, aspartic acid and glutamic acid phosphonic analogues. Arch Immunol Ther Exp (Warsz). 1980;28(3):433-8.

[5]. Cycloleucine blocks 5'-terminal and internal methylations of avian sarcoma virus genome RNA. Biochemistry. 1978 Aug 22;17(17):3627-32.

[6]. The effect of cycloleucine on SFV A7(74) infection in mice. Br J Exp Pathol. 1987 Apr;68(2):225-35.

1-Aminocyclopentanecarboxylic acid is a non-protein α-amino acid formed by the substitution of cyclopentane at the 1-position with an amino and carboxyl group. It is an EC 2.5.1.6 (methionine adenosyltransferase) inhibitor. Cycloleucine is a non-metabolizable amino acid formed by the cyclization of leucine. It is also a specific and reversible nucleic acid methylation inhibitor, widely used in biochemical experiments. Cycloleucine has been reported to exist in Streptomyces hydrogenans, with relevant data available. Cycloleucine is a non-metabolizable synthetic amino acid formed by the cyclization of leucine and possesses immunosuppressive, antitumor, and cell-suppressive activities. Cycloleucine competitively inhibits methionine adenosyltransferase, leading to the inhibition of the synthesis of S-adenosylmethionine (SAM) from methionine and ATP, thereby inhibiting nucleic acid methylation and polyamine production; ultimately resulting in the arrest of RNA (and possibly less so DNA) biosynthesis and cell cycle progression. This substance is also a competitive inhibitor of the glycine regulatory site of the N-methyl-D-aspartate (NMDA) receptor. Cyclic leucine is an amino acid formed by the cyclization of leucine. It has cell-inhibiting, immunosuppressive and antitumor activities. Mechanism of action Synthetic amino acid, considered to act as a valine antagonist. Therapeutic use Veterinary experimental use: A single dose of antilymphocyte serum, cyclophosphamide and cycloleucine delayed the onset of hyperacute allergic encephalomyelitis in rats. Cyclic leucine and thiaprozil hydrochloride have been shown to have a synergistic effect. Veterinary experimental use: (11)C-Cyclic leucine was evaluated as a tumor scanning agent in 38 patients. Based on animal data, the total body radiation dose was estimated to be 0.01 RAD/UCI and the pancreatic radiation dose was less than 0.06 RAD/UCI. Animals 33/38 also underwent gallium citrate (67)GA scanning; the results showed that 19 scans were positive and 24 scans were positive.
Drug (Veterinary): Administration of a few millicuries of carboxyl-labeled (11)C-cycloleucine. Tissue distribution (750 microcuries, intravenous injection) in rats carrying Morris 5123C hepatomas suggests the compound has the potential as a tumor localizer for detecting human cancer.
Drug (Veterinary): Cycloleucine protected rats from seizures in a maximal electric shock test, but had no protective effect against pentylenetetrazol-induced seizures in mice.

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Pharmacodynamics
Cycloleucine exhibits cytosolic, immunosuppressive, and antitumor activities.

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Cycloleucine is a selective NMDA receptor antagonist that targets the glycine modulatory site. It acts as a competitive antagonist at this site with a Ki of 600 μM and inhibits NMDA-evoked currents with an IC50 of 24 μM. Cycloleucine also inhibits S-adenosyl-methionine-mediated methylation and acts as a competitive inhibitor of ATP: L-methionine-S-adenosyl transferase. It is used as a research tool to study the NMDA receptor and its role in various physiological and pathological processes. The compound is not approved for human therapeutic use.

C6H11NO2

129.16

129.078

52-52-8

2901

White to off-white solid powder

1.2±0.1 g/cm3

256.1±23.0 °C at 760 mmHg

330 °C, DECOMP

108.7±22.6 °C

0.0±1.1 mmHg at 25°C

1.522

-0.05

2

3

1

9

127

0

C1CCC(C1)(C(=O)O)N

NILQLFBWTXNUOE-UHFFFAOYSA-N

InChI=1S/C6H11NO2/c7-6(5(8)9)3-1-2-4-6/h1-4,7H2,(H,8,9)

1-aminocyclopentane-1-carboxylic acid

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)

H2O: 20 mg/mL (154.85 mM)
DMSO: < 1 mg/mL

Solubility in Formulation 1: 100 mg/mL (774.23 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

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