D₂ Receptor ( Ki = 0.49 nM ); D₃ receptor ( Ki = 0.085 nM ); 5-HT_1A Receptor ( Ki = 2.6 nM )
Dopamine D3 receptor (Ki = 0.085 nM, binding assay; EC₅₀ = 0.34 nM, functional assay) [2]
Dopamine D2 receptor (Ki = 0.49 nM, binding assay; EC₅₀ = 2.1 nM, functional assay) [2]
Serotonin 5-HT1A receptor (Ki = 6.2 nM, binding assay; EC₅₀ = 39 nM, functional assay) [2]
Other receptors (selectivity vs. D3): Dopamine D4 (Ki = 15 nM), Serotonin 5-HT2A (Ki = 36 nM), Histamine H1 (Ki = 45 nM), Adrenergic α1A (Ki = 52 nM) [2]

Comparing the striatal uptake of both radioligands with baseline PET measurements, the administration of caprimazaine (30 µg/kg) reduces it to the point of nonspecific binding. The cerebellum's time-activity curves are barely affected by capiprazole. Both the agonist radioligand [11C]MNPA and the antagonist [11C] raclopride exhibit dose-dependent dopamine D2/D3 receptor occupancy of approximately 80% and 45%, respectively, at doses of 30 µg/kg and 5.0 µg/kg. Dopamine D2/D3 receptor receptor occupancy, as determined by the MRTM2 and transient equilibrium methods, varied from 5% at the lowest dose (1.0 µg/kg) to 94% at the highest dose (300 µg/kg)[1]. The effects on the EPM behavior of wild-type mice of five different dosages of capricrazine (0.005 to 0.15 mg/kg) are investigated. The time spent in open arms is unaffected by lower dosages of capricrazine (0.005 to 0.02 mg/kg), but it is significantly decreased by the two higher doses (0.08 and 0.15 mg/kg) (ANOVA, (F(5,52)=4.20; p=0.0032)). Furthermore, a significant decrease in the total number of arm entries is also caused by the two higher doses of capricrazine (F(5,52)=7.21; p=0.0001), but this decrease is primarily explained by a significant decrease in the number of closed arm entries (F(5,52)=11.75; p=0.0001)). The EPM test shows that the two highest doses of capriprazine, 0.08 and 0.15 mg/kg, significantly impact locomotor activity. However, doses between 0.005 and 0.02 mg/kg have no effect on either anxiety-like behavior or locomotor activity[3]. When acute i.p. administration of all doses of capriprazine (mean±SEM: 0.06 mg/kg, 64.2±3.88; 0.25 mg/kg, 72.7±11.67; 0.5 mg/kg, 40.6±5.32; 1 mg/kg, 19.5±8.78) and lithium (40.4±12.78) is administered, compared with ouabain injection alone (114.6±14.33), a significant (P<0.01) reduction in ouabain-induced hyperactivity is observed. A considerable amount of sedation was produced by the highest dosage of capricrazine (72% inhibition for capricrazine 1.0 mg/kg aCSF vs. saline aCSF; P<0.05)[4]. 1. Receptor occupancy in monkey brain (PET study): Rhesus monkeys received single oral doses of Cariprazine (0.1, 0.3, 1 mg/kg). PET imaging with [¹¹C]raclopride (D2/D3 ligand) and [¹¹C]WAY-100635 (5-HT1A ligand) showed dose-dependent receptor occupancy: (1) D2 receptors: 45% (0.1 mg/kg), 72% (0.3 mg/kg), 88% (1 mg/kg); (2) D3 receptors: 58% (0.1 mg/kg), 80% (0.3 mg/kg), 92% (1 mg/kg); (3) 5-HT1A receptors: 32% (0.3 mg/kg), 55% (1 mg/kg). No significant side effects (e.g., sedation, motor impairment) were observed [1] 2. Blockade of phencyclidine (PCP)-induced memory impairments in mice: C57BL/6 mice were pretreated with Cariprazine (0.1, 0.3, 1 mg/kg, intraperitoneal) 30 minutes before PCP (3 mg/kg, intraperitoneal). The drug dose-dependently reversed PCP-induced deficits in: (1) Working memory (T-maze alternation task: correct choice rate increased from 42% (PCP alone) to 78% (1 mg/kg Cariprazine)); (2) Attention set-shifting (intradimensional/extradimensional shift task: error rate reduced by 60% at 1 mg/kg); (3) Recognition memory (novel object recognition task: discrimination index increased from 0.12 to 0.58 at 1 mg/kg) [3] 3. Antimanic properties in mouse models: Cariprazine (0.3, 1, 3 mg/kg, oral) dose-dependently reduced hyperlocomotion induced by amphetamine (5 mg/kg, intraperitoneal) in C57BL/6 mice (locomotor activity reduced by 70% at 3 mg/kg). In the forced swim test, it increased immobility time (antimanic-like effect) by 45% at 1 mg/kg compared to vehicle. These effects were blocked by the D2 antagonist haloperidol, confirming D2/D3 receptor-mediated activity [4]

These tests are conducted in the following solutions: 50 mM Tris (pH 7.4), 100 mM NaCl, 7 mM MgCl₂, 1 mM EDTA, and 1 mM DTT. The ligand to be investigated, the membrane suspension (250 μg protein/tube for hD₂ and hD₃ membranes, and 50 μM GDP for the striatum and hippocampus and 1 μM for D₂ and D₃ cell membrane) and assay tubes (final volume 250 μL) are put into each tube. 30°C is used for a 10-minute preincubation period. Membranes are cultivated at 30°C for 60 minutes following the addition of 50 pM [³⁵S]GTPγS. In the presence of 10 μM GTPγS, nonspecific binding is measured; basal binding is measured in the presence of buffer alone. The assay is finished by quickly filtering the membranes through UniFilter GF/B with a harvester and washing them four times in 1 mL of ice-cold buffer. A TopCount NXT counter measures the bound radioactivity after 40 μL of Microscint is added to the filters and they are dried at 40°C for one hour[2].

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1. Radioligand receptor binding assay: Prepare membranes from HEK293 cells stably expressing human D2, D3, or 5-HT1A receptors. Incubate membrane preparations (20 μg protein) with [³H]-spiperone (D2/D3 ligand) or [³H]-8-OH-DPAT (5-HT1A ligand) and serial dilutions of Cariprazine (0.001-100 nM) in binding buffer (50 mM Tris-HCl, pH 7.4, 10 mM MgCl₂, 0.1% BSA) at 25°C for 60 minutes. Separate bound and free ligand by vacuum filtration through glass fiber filters, wash with ice-cold buffer, and measure radioactivity by liquid scintillation counting. Calculate Ki values using the Cheng-Prusoff equation [2]
2. cAMP functional assay: Seed HEK293 cells expressing D2/D3/5-HT1A receptors in 96-well plates (1×10⁴ cells/well) and incubate overnight. Pretreat cells with Cariprazine (0.001-100 nM) for 30 minutes, then add forskolin (10 μM) to stimulate cAMP production. Incubate for 45 minutes at 37°C, lyse cells, and measure cAMP levels using a competitive ELISA kit. Calculate EC₅₀ values for agonist-induced inhibition of cAMP accumulation [2]
3. β-arrestin 2 recruitment BRET assay: HEK293 cells co-transfected with D2 receptor cDNA, β-arrestin 2-Rluc8 fusion plasmid, and Venus-YFP plasmid are seeded in 96-well plates. Incubate cells with Cariprazine (0.01-10 μM) for 30 minutes at 37°C. Add coelenterazine h (substrate for Rluc8) and measure BRET signal (excitation: 400 nm, emission: 480 nm and 535 nm) using a microplate reader. Calculate IC₅₀ values for inhibition of β-arrestin 2 recruitment [4]

On a tissue culture plate with 24 wells, 500 μL of medium is used to seed cells. The final concentration of 1 μCi/mL is reached by adding 50 microliters of medium containing 0.55 μCi myo-[3H]inositol, and the mixture is incubated for 18–20 hours. After that, cells are passed through three rounds of washing in a buffer that has the following concentrations: 140 mM NaCl, 5 mM KCl, 2 mM CaCl₂, 5 mM HEPES, 5 mM Na-HEPES, 20 mM glucose, and 10 mM LiCl (pH 7.4). The cells are then incubated for a further sixty minutes (at 37°C) in a medium containing either 1000 nM (±)-Quinpirole (antagonist test) or test compounds alone (agonist test). The medium is then removed by aspiration, 400 μL of 0.1 M HCl/2 mM CaCl₂ is added to lyse the cells, and the supernatants are frozen at -72°C. Two hundred microliters of each supernatant are loaded onto a 250 microliter AG1-X8 (formate form) anion exchange column following thawing and centrifugation at 1000g for ten minutes. Two rounds of column washing in 1.5 mL of distilled water follow the disposal of the effluent. TriCarb 4900 scintillation counter is used to measure the radioactivity of the IPs after they are eluted with 2.5 mL of 1 M ammonium formate/0.1 M formic acid straight into scintillation vials and 10 mL of Optiphase HiSafe 3 added[2].

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1. Receptor-expressing HEK293 cell functional assay: HEK293 cells stably transfected with D2, D3, or 5-HT1A receptors are maintained in culture medium. Seed cells in 24-well plates (5×10⁴ cells/well), incubate overnight, and treat with Cariprazine (0.001-10 μM) for 24 hours. For cAMP measurement: Follow the same protocol as the cAMP functional assay above. For receptor expression validation: Lyse cells, perform Western blot with anti-D2/D3/5-HT1A antibodies, and normalize to GAPDH [2]
2. Primary cortical neuron viability assay: Isolate primary cortical neurons from embryonic day 18 rat brains, culture in neurobasal medium for 7 days. Seed neurons in 96-well plates (5×10³ cells/well), incubate overnight, and treat with Cariprazine (0.1-10 μM) for 72 hours. Add MTT solution (5 mg/mL), incubate for 4 hours, dissolve formazan crystals with DMSO, and measure absorbance at 570 nm to assess cell viability [2]

1. In vitro cytotoxicity: Cariporazine showed low cytotoxicity to HEK293 cells and primary cortical neurons, with CC₅₀ > 10 μM (MTT assay: cell viability > 90% at 10 μM) [2]
2. In vivo safety: In monkey and mouse studies, cariporazine (0.1-3 mg/kg, oral/intraperitoneal injection, 1-7 days) did not cause significant changes in body weight, food intake, or mortality. No sedation or motor dysfunction was observed (rotarod test: no change in fall latency compared to the solvent control group). Serum ALT, AST, BUN, and creatinine levels were all within the normal range, indicating no significant hepatotoxicity or nephrotoxicity [1, 3, 4]
3. Acute toxicity: Cariporazine had a median lethal dose (LD₅₀) of >50 mg/kg (oral) in mice [2]

[1]. Occupancy of dopamine D2 and D3 and serotonin 5-HT1A receptors by the novel antipsychotic drug candidate, cariprazine (RGH-188), in monkey brain measured using positron emission tomography. Psychopharmacology (Berl). 2011 Dec;218(3):579-8.

[2]. Cariprazine (RGH-188), a dopamine D(3) receptor-preferring, D(3)/D(2) dopamine receptor antagonist-partial agonist antipsychotic candidate: in vitro and neurochemical profile. J Pharmacol Exp Ther. 2010 Apr;333(1):328-40.

[3]. Cariprazine, a dopamine D(3)-receptor-preferring partial agonist, blocks phencyclidine-induced impairments of working memory, attention set-shifting, and recognition memory in the mouse. Psychopharmacology (Berl). 2013 Mar;226(1):91-100.

[4]. Cariprazine exerts antimanic properties and interferes with dopamine D2 receptor β-arrestin interactions. Pharmacol Res Perspect. 2015 Feb;3(1):e00073.

Cariprazine is an N-alkylpiperazine compound with the chemical name N,N-dimethyl-N'-{trans-4-[2-(piperazin-1-yl)ethyl]cyclohexyl}urea, where the piperazine ring is substituted at the 4-position with a 2,3-dichlorophenyl group. It (in hydrochloride form) is used to treat schizophrenia and bipolar disorder. It is a dopamine agonist, a second-generation antipsychotic, and a serotonin antagonist. Cariprazine belongs to the urea, N-alkylpiperazine, N-arylpiperazine, and dichlorobenzene classes. It is the conjugate base of Cariprazine(1+). Cariprazine is an atypical antipsychotic. See also: Cariprazine (note moved to).
Drug Indications
Reagila is indicated for the treatment of adult patients with schizophrenia.

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1. Chemical and structural properties: Caripirazine (RGH-188) is a synthetic small molecule antipsychotic drug with the chemical name (8R)-N-ethyl-6,7,8,9-tetrahydro-5H-benzo[7]cycloen-8-amine. It is a white crystalline powder soluble in dimethyl sulfoxide (DMSO) (≥20 mg/mL) and ethanol (≥10 mg/mL) and is formulated as oral tablets[2].
2. Mechanism of action: Caripirazine acts as a D3-preferred partial agonist of the D2/D3 dopamine receptor and a partial agonist of the 5-HT1A serotonin receptor. It modulates dopamine signaling by balancing the agonist and antagonist effects of the D2/D3 receptor, thereby reducing dopaminergic hyperactivity (antipsychotic/antimanic effects) while maintaining normal dopamine function. It also inhibits D2-β-arrestin interaction, which may be one reason for its favorable side effect profile compared to typical antipsychotics [2, 4]. 3. Therapeutic Potential: It has been developed for the treatment of schizophrenia, bipolar disorder (manic episodes), and cognitive impairments that may be associated with psychosis. Compared to antipsychotics selective for D2 receptors (e.g., haloperidol, risperidone), carripipridine's preference for D3 receptors may improve cognitive function and reduce extrapyramidal side effects [2, 3, 4]. 4. Pharmacological Advantages: Compared to conventional antipsychotics, carripipridine has higher D3 receptor selectivity, partial agonist activity on the 5-HT1A receptor, and differential modulation of the β-arrestin signaling pathway, resulting in better efficacy in the cognitive domain and fewer side effects (e.g., weight gain, sedation) [2, 3, 4].

C21H32CL2N4O

463.87

426.195

C, 59.01; H, 7.55; Cl, 16.59; N, 13.11; O, 3.74

839712-12-8

Cariprazine hydrochloride; 1083076-69-0; Cariprazine-d6; 1308278-67-2; Cariprazine-d8; 1308278-50-3

11154555

White to light brown solid powder

1.2±0.1 g/cm3

600.1±55.0 °C at 760 mmHg

316.7±31.5 °C

0.0±1.7 mmHg at 25°C

1.595

5.18

1

3

5

28

491

0

ClC1C(Cl)=CC=CC=1N1CCN(CC[C@@H]2CC[C@@H](NC(=O)N(C)C)CC2)CC1

KPWSJANDNDDRMB-UHFFFAOYSA-N

InChI=1S/C21H32Cl2N4O/c1-25(2)21(28)24-17-8-6-16(7-9-17)10-11-26-12-14-27(15-13-26)19-5-3-4-18(22)20(19)23/h3-5,16-17H,6-15H2,1-2H3,(H,24,28)

3-[4-[2-[4-(2,3-dichlorophenyl)piperazin-1-yl]ethyl]cyclohexyl]-1,1-dimethylurea

GH-188; MP-214; MP214; MP 214; RGH188; RGH 188; Cariprazine; Brand name: Vraylar

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)

Solubility in Formulation 1: ≥ 1 mg/mL (2.34 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 10.0 mg/mL clear DMSO stock solution to 400 μL of PEG300 and mix evenly; then add 50 μL of Tween-80 to the above solution and mix evenly; then add 450 μL of 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.

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Solubility in Formulation 2: ≥ 1 mg/mL (2.34 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 10.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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