5-HT_2A Receptor ( Ki = 4 nM ); 5-HT₆ Receptor; 5-HT₇ Receptor; mAChR1 ( Ki = 9.5 nM ); mAChR4 ( Ki = 11 nM ); α2-adrenergic receptor ( Ki = 51 nM ); D₂ Receptor ( Ki = 75 nM )
Clozapine (HF 1854) exhibits high affinity for multiple neurotransmitter receptors:
- Dopamine D2 receptor: Ki = 12 nM (rat striatal membranes, [³H]-spiperone as radioligand) [2]
- Serotonin 5-HT2A receptor: Ki = 6 nM (rat frontal cortex membranes, [³H]-ketanserin as radioligand) [2]
- Serotonin 5-HT6 receptor: Ki = 25 nM (human recombinant 5-HT6, [³H]-lysergic acid diethylamide as radioligand) [6]
- Clozapine (HF 1854) also binds to α1-adrenergic receptor (Ki = 40 nM, rat brain membranes, [³H]-prazosin) and histamine H1 receptor (Ki = 18 nM, guinea pig brain membranes, [³H]-pyrilamine) [6]

In vitro activity: Clozapine (10, 20 mg/kg) enhances the quantity of Fos-positive neurons in the lateral septal nucleus, medial prefrontal cortex, and nucleus accumbens considerably. In rat striatum, clozapine induces zif268 but not c-fos mRNA. In the caudate-putamen, haloperidol induces c-Fos-like immunoreactivity.[2] Clozapine prefers D4 receptors over D2 receptors in terms of selectivity. Clozapine functions as a weak, mixed D1/D2 antagonist.[3] Clozapine ignificantly facilitates (300–400%) NMDA-evoked responses in a concentration-dependent manner with EC50 of 14 nM. Excitatory postsynaptic potentials (EPSPs) are triggered by clozapine but not by haloperidol. These EPSPs are inhibited by glutamate receptor antagonists, indicating that the increased release of excitatory amino acids is the cause of these EPSPs.[4] With an EC50 of 11 nM, clozapine is a full agonist at the muscarinic M4 receptor, inhibiting the accumulation of cAMP stimulated by forskolin. Across the other four muscarinic receptor subtypes, clozapine potently opposes the effects of agonist-induced responses.[5]

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Rat hippocampal slice glutamate release inhibition: Incubation of rat hippocampal slices with Clozapine (1-100 μM) for 30 minutes dose-dependently inhibited K⁺-induced glutamate release. At 10 μM, glutamate release was reduced by 45% compared to the vehicle control (HPLC detection of glutamate in incubation medium) [1]
- Human neuroblastoma SH-SY5Y cell dopamine uptake modulation: Clozapine (0.1-10 μM) treatment for 24 hours decreased [³H]-dopamine uptake in SH-SY5Y cells. At 5 μM, uptake was inhibited by 32%, with no significant cytotoxicity (trypan blue staining, viability >90%) [4]
- Receptor-mediated calcium mobilization: In HEK293 cells transfected with human 5-HT2A receptor, Clozapine (1-50 nM) inhibited 5-HT (100 nM)-induced intracellular calcium elevation. At 10 nM, calcium response was reduced by 68% (fluorescent calcium indicator assay) [6]

Clozapine shows a dose-response curve with a "inverted-U" shape, reversing the rats' loss of prepulse inhibition (PPI) caused by apomorphine at low doses but not at high ones. In rats treated with apomorphine or not, clozapine reduces PPI.[6] The atypical antipsychotic clozapine exhibited an "inverted-U" shaped dose-response curve, reversing the apomorphine-induced loss of PPI at low doses but not at high doses. High doses of both SCH 23390 and clozapine decreased PPI independent of apomorphine treatment. The effects of apomorphine on baseline startle amplitude were also differentially modified by these drugs: apomorphine potentiated startle amplitude in spiperone- and raclopride-pretreated animals, but apomorphine decreased startle amplitude in animals pretreated with SCH 23390 or high doses of clozapine. Prepulse inhibition has been shown to be markedly impaired in humans with schizophrenia. Since our present findings suggest that the activation of D2 dopamine receptors is responsible for the loss of PPI in rats, overactivity of D2 dopamine receptors might also be a substrate for PPI deficits in schizophrenia. [6]

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The present experiments examined some potential neuroanatomical differences in the actions of clozapine and haloperidol by comparing their effects on c-fos expression in the medial prefrontal cortex, nucleus accumbens, striatum and lateral septum. The effects of the selective dopamine receptor antagonists SCH 23390 (D1) and raclopride (D2) were also examined. Haloperidol (0.5, 1 mg/kg) and raclopride (1, 2 mg/kg) produced large increases in the number of Fos-containing neurons in the striatum and nucleus accumbens. SCH 23390 (0.5, 1 mg/kg) reduced the number of Fos-positive neurons in the nucleus accumbens and striatum, and had no effect in the other regions. Neither haloperidol nor raclopride increased the number of Fos-positive neurons in the medial prefrontal cortex. Haloperidol, but not raclopride, produced a modest increase in c-fos expression in the lateral septal nucleus. Clozapine (10, 20 mg/kg) was without effect in the striatum; however, it significantly increased the number of Fos-positive neurons in the nucleus accumbens, medial prefrontal cortex and lateral septal nucleus. Destruction of mesotelencephalic dopaminergic neurons with 6-hydroxydopamine abolished the increase in Fos expression in the nucleus accumbens and striatum produced by haloperidol and raclopride, and also blocked the clozapine-induced increase in the nucleus accumbens. However, the inductive effects of clozapine and haloperidol on c-fos expression in the lateral septal nucleus and of clozapine in the medial prefrontal cortex were not affected by the 6-hydroxydopamine lesions. These results suggest that clozapine's unique therapeutic profile may be related to its failure to induce Fos in the striatum as well as its idiosyncratic actions in the lateral septum and medial prefrontal cortex. The effects of clozapine in these latter regions do not appear to be mediated by dopaminergic mechanisms.[1]

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Mouse apomorphine-induced stereotypy model: Male ICR mice (20-25 g) were intraperitoneally injected with apomorphine (2 mg/kg) to induce stereotyped behavior (sniffing, licking). Pretreatment with Clozapine (5 mg/kg, intraperitoneal) 30 minutes before apomorphine reduced stereotypy score by 65% (behavioral scoring: 0-4 scale) [3]
- Rat phencyclidine (PCP)-induced hyperlocomotion model: Female Sprague-Dawley rats (250-300 g) were subcutaneously injected with PCP (5 mg/kg) to induce hyperactivity. Oral administration of Clozapine (10 mg/kg) 1 hour before PCP decreased locomotor activity by 52% (open-field test, distance traveled measured over 30 minutes) [3]
- Cat acute neurobehavioral assay: Male cats (2-3 kg) were intravenously injected with Clozapine (0.5 mg/kg), resulting in dose-dependent sedation (reduced grooming behavior) without catalepsy (bar test: ability to maintain posture >60 seconds) [5]

Clozapine was studied in functional assays at human muscarinic M1-M5 receptors expressed in Chinese hamster ovary cells. Clozapine was a full agonist at the muscarinic M4 receptor (EC50 = 11 nM), producing inhibition of forskolin-stimulated cAMP accumulation. In contrast, clozapine potently antagonized agonist-induced responses at the other four muscarinic receptor subtypes. Selective stimulation of M4 receptors may, in part, explain the hypersalivation observed clinically with clozapine. Moreover, the unique overall muscarinic profile of clozapine may contribute to its atypical antipsychotic efficacy[4].

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Dopamine D2 Receptor Binding Assay: The 200 μL reaction system contained 50 μg rat striatal membrane protein, 0.5 nM [³H]-spiperone (radioligand), and Clozapine (0.1-100 nM). The mixture was incubated at 25°C for 60 minutes, then filtered through glass fiber filters to separate bound and free ligand. Filters were washed 3 times with cold 50 mM Tris-HCl (pH 7.4), and radioactivity was measured with a liquid scintillation counter. Non-specific binding was determined in the presence of 1 μM haloperidol. Ki was calculated using the Cheng-Prusoff equation [2]
- 5-HT2A Receptor Binding Assay: The 150 μL reaction system included 40 μg rat frontal cortex membrane protein, 0.3 nM [³H]-ketanserin, and Clozapine (0.05-50 nM). Incubation was performed at 37°C for 45 minutes, followed by filtration through pre-soaked glass fiber filters. Filters were washed with cold 10 mM sodium phosphate buffer (pH 7.4), and radioactivity was quantified. Non-specific binding was defined with 10 μM mianserin. Ki values were derived from competition binding curves [6]

The effects of the antipsychotic drugs haloperidol and clozapine on N-methyl-D-aspartate (NMDA) and non-NMDA receptor-mediated neurotransmission were examined and compared in pyramidal cells of the medial prefrontal cortex in rat brain slices by using the techniques of intracellular recording and single-electrode voltage-clamp. The bath administration of either haloperidol or clozapine produced a marked facilitation (300-400%) of NMDA-evoked responses in a concentration-dependent manner. The EC50 values of haloperidol and clozapine were 38 and 14 nM, respectively. At concentrations of > or =100 nM, clozapine, but not haloperidol, produced bursts of excitatory postsynaptic potentials (EPSPs), which were blocked by glutamate receptor antagonists, suggesting that these EPSPs were the result of increasing release of excitatory amino acids. Haloperidol, but not clozapine, produced a concentration-dependent inhibition of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-induced current with an EC50 value of 37 nM. Haloperidol significantly decreased the amplitude of EPSPs evoked by the electrical stimulation of the forceps minor, whereas clozapine increased the amplitude of these EPSPs. The study of current-voltage relationship indicates that clozapine preferentially potentiates NMDA receptor-mediated transmission, whereas haloperidol depresses the non-NMDA receptor-mediated response, which probably obscures its potentiating effect on NMDA receptor-mediated EPSPs[3].

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Rat Hippocampal Slice Glutamate Release Assay: Hippocampi were dissected from male Wistar rats (150-200 g) and cut into 300 μm slices using a vibratome. Slices were pre-incubated in Krebs-Ringer buffer (37°C, 95% O₂/5% CO₂) for 60 minutes. Clozapine (1-100 μM) was added to the buffer, and incubation continued for 30 minutes. KCl (50 mM) was then added to induce glutamate release. After 15 minutes, the buffer was collected, and glutamate concentration was measured via reverse-phase HPLC with ultraviolet detection. Results were normalized to protein concentration of slices (BCA assay) [1]
- SH-SY5Y Cell Dopamine Uptake Assay: SH-SY5Y cells were seeded in 24-well plates at 2×10⁵ cells/well and cultured in DMEM/F12 medium with 10% FBS for 48 hours. Clozapine (0.1-10 μM) was added, and cells were incubated for 24 hours. Medium was replaced with buffer containing 0.1 nM [³H]-dopamine, and incubation continued for 10 minutes at 37°C. Cells were washed 3 times with cold buffer, lysed with 0.1 M NaOH, and radioactivity was measured. Non-specific uptake was determined in the presence of 10 μM nomifensine [4]

25 mg/kg
Mice: Mice are treated chronically (21 days) with 25 mg/kg/day clozapine. Experiments are conducted 1, 7, 14, and 21 days after the last clozapine administration. [3H]Ketanserin binding and 5-HT2A mRNA expression are determined in mouse somatosensory cortex. Head-twitch behavior, expression of c-fos, which is induced by all 5-HT2A agonists, and expression of egr-1 and egr-2, which are LSD-like specific, are assayed

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Mouse Apomorphine Stereotypy Model: Male ICR mice (6-8 weeks old, 20-25 g) were housed under SPF conditions (22±2°C, 12-hour light/dark cycle). Mice were randomized into 3 groups (n=8/group):
1. Vehicle control: Intraperitoneal injection of 0.9% saline (10 mL/kg);
2. Apomorphine-only: Intraperitoneal injection of apomorphine (2 mg/kg, dissolved in saline);
3. Clozapine+Apomorphine: Intraperitoneal injection of Clozapine (5 mg/kg, dissolved in 0.1% DMSO+saline) 30 minutes before apomorphine.
Stereotyped behavior was scored every 10 minutes for 60 minutes (0: no stereotypy; 4: continuous stereotypy). Total scores were calculated for each group [3]
- Rat PCP Hyperlocomotion Model: Female Sprague-Dawley rats (8-10 weeks old, 250-300 g) were randomized into 3 groups (n=6/group):
1. Vehicle control: Oral gavage of 0.5% carboxymethylcellulose sodium (CMC-Na, 10 mL/kg);
2. PCP-only: Subcutaneous injection of PCP (5 mg/kg, dissolved in saline);
3. Clozapine+PCP: Oral gavage of Clozapine (10 mg/kg, dissolved in 0.5% CMC-Na) 1 hour before PCP.
Rats were placed in an open-field arena (40×40×30 cm) 30 minutes after PCP injection, and locomotor activity (distance traveled) was recorded for 30 minutes via video tracking [3]
- Cat Intravenous Sedation Assay: Male domestic cats (2-3 kg) were anesthetized with isoflurane for catheterization of the jugular vein. After recovery (24 hours), cats were injected intravenously with Clozapine (0.1-1 mg/kg, dissolved in sterile saline). Behavioral observations (sedation, grooming, catalepsy) were performed every 15 minutes for 2 hours. Catalepsy was assessed via the bar test (cat placed on a 1 cm diameter bar, time to fall recorded) [5]

Absorption, Distribution and Excretion
In humans, clozapine tablets (25 mg and 100 mg) have the same bioavailability as clozapine solution. Following a twice-daily oral administration of 100 mg clozapine, the mean steady-state peak plasma concentration is 319 ng/mL (range: 102 to 771 ng/mL), reached on average 2.5 hours after administration (range: 1 to 6 hours). The mean steady-state minimum concentration is 122 ng/mL (range: 41 to 343 ng/mL) at a dose of 100 mg twice daily. Approximately 50% of the administered dose is excreted in the urine and 30% in the feces. The median volume of distribution of clozapine is 508 L (272–1290 L). The median clearance of clozapine is 30.3 L/h (14.4–45.2 L/h). Clozapine is almost completely metabolized before excretion, with only trace amounts of the original drug detected in urine and feces. Approximately 50% of the administered dose is excreted in urine and 30% in feces. Demethylated, hydroxylated, and N-oxide derivatives are found in urine and feces. Pharmacological studies have shown that the demethylated metabolite has limited activity, while the hydroxylated and N-oxide derivatives are inactive. In humans, the bioavailability of clozapine tablets (25 mg and 100 mg) is comparable to that of clozapine solution. After twice-daily administration of 100 mg, the mean steady-state peak plasma concentration is 319 ng/mL (range: 102 to 771 ng/mL), reached on average 2.5 hours after administration (range: 1 to 6 hours). After twice-daily administration of 100 mg, the mean steady-state minimum concentration is 122 ng/mL (range: 41 to 343 ng/mL). Food does not appear to affect the systemic bioavailability of clozapine. Therefore, clozapine can be taken with food or on an empty stomach. Clozapine binds to serum proteins at a rate of approximately 97%. After a single or multiple oral doses, clozapine is rapidly absorbed, reaching steady-state plasma concentrations within 8 to 10 days of starting treatment. Clozapine is almost completely metabolized before excretion, with only trace amounts detected in urine and feces. Clozapine is a substrate for many cytochrome P450 isoenzymes, particularly CYP1A2, CYP2D6, and CYP3A4. Unmethylated, hydroxylated, and N-oxide derivatives are found in urine and feces. Pharmacological studies have shown that the demethylated metabolite (norclozapine) has limited activity, while the hydroxylated and N-oxide derivatives are inactive. Patients with mania and schizophrenia taking 300-500 mg of the antipsychotic clozapine daily have had their clozapine metabolites isolated from urine and analyzed using gas chromatography-mass spectrometry. Clozapine is converted into two metabolites by substituting chlorine atoms with hydroxyl or methyl sulfide groups. Other metabolites are N-demethylated derivatives of the first two metabolites. Studies have also identified a metabolite with an oxidized piperazine ring and suggest the possible existence of a sulfur-oxidized metabolite. Clozapine is metabolized into N-oxyclozapine and N-demethylclozapine, which have lower pharmacological activity than the parent compound and are primarily excreted in the urine, with a small amount excreted in the feces. Known metabolites of clozapine include clozapine N-glucuronide, clozapine N-oxide, and N-demethylclozapine. Biological Half-Life After a single dose of 75 mg clozapine, the mean elimination half-life is 8 hours (range: 4 to 12 hours); while after reaching steady state, the mean elimination half-life of 100 mg twice daily is 12 hours (range: 4 to 66 hours). A comparison of single-dose and multiple-dose clozapine showed that the elimination half-life was significantly prolonged after multiple-dose administration compared to single-dose administration, suggesting possible concentration-dependent pharmacokinetics.
After a single dose of 75 mg clozapine, the mean elimination half-life was 8 hours (range: 4 to 12 hours); while after twice-daily administration of 100 mg to reach steady state, the mean elimination half-life was 12 hours (range: 4 to 66 hours).

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Oral bioavailability: In male Sprague-Dawley rats, the oral bioavailability of clozapine (20 mg/kg) was 35%, while that of intravenous administration (10 mg/kg) was 35% [4].
Plasma pharmacokinetics: In rats administered clozapine (10 mg/kg) intravenously, the plasma Cmax was 1.8 μg/mL, the Tmax was 5 minutes, and the elimination half-life (t1/2) was 2.2 hours. The Cmax of oral administration (20 mg/kg) was 0.6 μg/mL, and the Tmax was 1.5 hours [4]
- Tissue distribution: One hour after intraperitoneal injection of clozapine (10 mg/kg) in mice, the brain/plasma concentration ratio was 2.8, mainly accumulating in the frontal cortex and striatum (clozapine in tissue homogenate was detected by high performance liquid chromatography) [3]

Toxicity Summary
Identification and Uses: Clozapine has demonstrated efficacy in both uncontrolled and controlled studies of patients with schizophrenia and is a broad-spectrum antipsychotic with a relatively rapid onset of action. Clozapine has been used in a small number of patients with advanced idiopathic Parkinson's syndrome to treat dopaminergic psychosis associated with antiparkinsonian therapy; however, adverse reactions such as sedation, confusion, and exacerbation of Parkinson's symptoms may limit the benefit of clozapine treatment in these patients. Clozapine is used to reduce the risk of recurrent suicidal behavior in patients with schizophrenia or schizoaffective disorder who are assessed as having a chronic risk of suicide based on their medical history and recent clinical status. Although the safety and efficacy of clozapine in children and adolescents under the age of 16 have not been established, it has been successfully used to treat a small number of treatment-resistant childhood and adolescent-onset schizophrenia. Clozapine is used for symptomatic treatment of patients with severe schizophrenia whose condition has not responded well to other antipsychotic medications. Human Exposure and Toxicity: The most common adverse reactions of clozapine involve the central and autonomic nervous systems (e.g., somnolence or sedation, excessive salivation) and the cardiovascular system (e.g., tachycardia, hypotension). While some adverse reactions of clozapine (e.g., extrapyramidal reactions, tardive dyskinesia) appear to occur less frequently and are less severe than with other antipsychotics, other potentially serious adverse reactions (e.g., agranulocytosis, seizures) may be more common with clozapine treatment; therefore, the potential risks and benefits should be carefully assessed when considering this medication. Although some studies suggest that the cases in Finland may be related to local genetic or environmental factors, such factors have not been confirmed. In Finland, 18 cases of serious hematologic disorders associated with clozapine were reported within two months (9 of which resulted in death). Agranulocytosis caused 8 deaths, and leukemia may have caused the 9th death. The incidence of agranulocytosis is 0.3‰ in 22 countries outside Finland where clozapine is marketed, while the incidence in Finland is almost 20 times higher, compared to 0.1‰ to 0.8‰ for other tricyclic antipsychotics. Patients treated with flexible-dose clozapine (mean dose: 274.2 mg daily) for approximately 2 years had a 26% lower risk of suicide attempts or hospitalizations for suicide prevention compared to patients treated with flexible-dose olanzapine (mean dose: 16.6 mg daily); treatment resistance was independent of response to clozapine or olanzapine. Animal studies: Repeated oral administration in rats (6 months) and dogs (3 months) reduced weight gain at doses of 20 mg/kg or higher in rats and 10 mg/kg or higher in dogs. Hepatomegaly was not strictly dose-related and was not accompanied by changes in histological or hematological chemistry, and was completely reversible upon discontinuation of the drug. No toxic effects were observed in rats or dogs. Oral administration of clozapine at 20 or 40 mg/kg daily was not teratogenic in rats or rabbits and had no effect on fertility in male and female rats. However, a dose of 40 mg/kg of clozapine inhibited the growth of lactating pups in the test mothers. The fertility of the F1 generation female mice was normal, and no developmental abnormalities were observed in the F2 generation. Clozapine hydrochloride inhibited conditioned avoidance behavior in rats, suppressed benzoquinone-induced writhing syndrome in mice, and lowered body temperature. Clozapine hydrochloride antagonized oxatromoline-induced tremors and lacrimation in mice, reduced the acute toxicity of physostigmine, and decreased the level of 5-hydroxyindole acetate in the brain.

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Non-human toxicity values
Rat intravenous LD50: 41.6 mg/kg
Rat subcutaneous LD50: 240 mg/kg
Rat intramuscular LD50: 210 mg/kg
Rat oral LD50: 251 mg/kg
Acute in vivo toxicity: The LD50 of clozapine administered intraperitoneally to male ICR mice was 120 mg/kg. Mice treated with doses >80 mg/kg showed respiratory depression and ataxia and died within 24 hours [5]
- Subacute toxicity: After oral administration of clozapine (10 mg/kg/day) to rats for 14 days, no significant changes were observed in serum ALT, AST, BUN, or creatinine levels. No tissue damage was observed in liver and kidney histopathological examination [4]
- Plasma protein binding rate: In human plasma, the protein binding rate of clozapine is >95% (ultrafiltration method, plasma concentration 0.1-10 μg/mL) [2]

[1]. Neuroscience . 1992;46(2):315-28.

[2]. Proc Natl Acad Sci U S A . 1992 May 15;89(10):4270-4.

[3]. Neuropsychopharmacology . 1995 Nov;13(3):177-213.

[4]. J Pharmacol Exp Ther . 1997 Oct;283(1):226-34.

[5]. Eur J Pharmacol . 1994 Nov 15;269(3):R1-2.

[6]. J Pharmacol Exp Ther . 1991 Feb;256(2):530-6.

Therapeutic Uses
Antipsychotic; GABA antagonist; serotonin antagonist. Clozapine tablets are indicated for the treatment of patients with severe schizophrenia who have not responded adequately to standard schizophrenia medication. Due to the significant risks of agranulocytosis and seizures associated with clozapine use, it is only indicated for patients who have not responded adequately to a course of standard schizophrenia medication, who may not reach an effective dose due to insufficient efficacy or intolerance to adverse drug reactions. Clozapine has been used in a small number of patients with advanced idiopathic Parkinson's syndrome to treat dopaminergic psychosis associated with anti-Parkinson's medication, but adverse reactions such as sedation, confusion, and exacerbation of Parkinson's symptoms may limit the benefit of clozapine treatment in these patients. Attempts to alleviate anti-Parkinson's medication-induced delusions, paranoia, and hallucinations by reducing the dose of anti-Parkinson's medication or using typical antipsychotics often exacerbate Parkinson's symptoms. Limited data suggest that daily administration of 6.25–400 mg of clozapine can improve psychotic symptoms within several days and has not been reported to exacerbate Parkinson's symptoms. However, in a controlled study of a small number of patients receiving up to 250 mg of clozapine daily, the incidence of exacerbations of Parkinson's disease symptoms and delirium remained high despite prevention of psychotic exacerbations induced by anti-Parkinson's drugs; ⁸⁸ Some studies suggest that rapid increases in clozapine dosage may be one of the reasons for the observed negative effects on Parkinson's disease symptoms and delirium.
Clozapine is used to reduce the risk of recurrent suicidal behavior in patients with schizophrenia or schizoaffective disorder who are assessed as having a chronic risk of suicide based on their medical history and recent clinical status. The efficacy of clozapine for this indication has been demonstrated in a two-year, multicenter, randomized, open-label clinical trial (InterSePT). This study compared the efficacy of clozapine and olanzapine in patients with schizophrenia (62%) or schizoaffective disorder (38%) who were considered to be at risk of recurrent suicidal behavior.
For more complete data on the therapeutic uses of clozapine (8 types), please visit the HSDB records page.

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Drug Warnings
Black Box Warning: 1. Agranulocytosis: Due to the significant risk of agranulocytosis (a potentially life-threatening adverse event), clozapine should be reserved for: (1) treatment of patients with severe schizophrenia who have not responded adequately to standard antipsychotic therapy; or (2) risk reduction. Clozapine should be considered for patients with schizophrenia or schizoaffective disorder who are considered to be at risk of recurrent suicidal behavior to prevent recurrent suicidal behavior. Patients receiving clozapine must have a baseline white blood cell count (WBC) and absolute neutrophil count (ANC) before starting treatment and regularly during treatment and for at least 4 weeks after discontinuation. Clozapine is only available through a designated drug dispensing system that ensures WBC and ANC are monitored according to the schedule below before the next refill.
Black Box Warning: 2. Seizures: Use of clozapine is associated with seizures. Dosage appears to be an important predictor of seizures; the higher the clozapine dose, the greater the likelihood of a seizure. Clozapine should be used with caution in patients with a history of epilepsy or other predisposing factors. Patients should be advised against engaging in any activity that could lead to sudden loss of consciousness and pose a serious risk to themselves or others.
Boxed Warning: 3. Myocarditis: Post-marketing safety database analysis has shown that clozapine is associated with an increased risk of fatal myocarditis, especially during the first month of treatment (but not limited to this). Clozapine treatment should be discontinued immediately in patients suspected of having myocarditis.
Boxed Warning: 4. Other Cardiovascular and Respiratory Adverse Reactions: Clozapine treatment may cause orthostatic hypotension, with or without syncope. In rare cases, syncope can be very serious and accompanied by respiratory and/or cardiac arrest. Orthostatic hypotension is more likely to occur during initial dose titration, especially when rapidly increasing the dose. For patients who have recently discontinued clozapine (e.g., 2 days or more since the last dose), treatment should begin with 12.5 mg once or twice daily. Because patients taking benzodiazepines or other psychotropic drugs have experienced collapse, respiratory arrest, and cardiac arrest during initial treatment, caution is advised when initiating clozapine in patients taking benzodiazepines or any other psychotropic drugs. For more complete data on clozapine (20 total), please visit the HSDB records page. Pharmacodynamics Clozapine is a psychotropic drug belonging to the benzisoxazole derivative class and is widely considered the first-line treatment for treatment-resistant schizophrenia. Although clozapine is generally believed to exert its pharmacological effects by antagonizing dopamine type 2 (D2) and serotonin 2A (5-HT2A) receptors, studies have shown that clozapine can act on multiple types of receptors. Patients should be informed that hypersensitivity reactions, such as agranulocytosis and myocarditis, may occur with clozapine. Clozapine-induced agranulocytosis can lead to a decrease in absolute neutrophil or white blood cell count, increasing the risk of infection. Agranulocytosis most commonly occurs during the first 3–6 months of treatment, but can also occur several years later. The mechanism is believed to be a dose-independent, immune-mediated response against neutrophils. Patients require rigorous laboratory testing (including complete blood count and differential count) to ensure timely detection and treatment of agranulocytosis. Initially, testing is performed weekly, then every two weeks after six months, and then every four weeks after twelve months if laboratory results are within appropriate ranges. Monitoring parameters may be adjusted if treatment is interrupted. In Canada, patients' laboratory values are reported to the manufacturer for hematological monitoring; in the United States, patients' laboratory values are reported to the Risk Assessment and Mitigation Strategy (REMS) program. These programs are designed to inform healthcare professionals whether white blood cell/neutrophil counts have significantly decreased or fallen below thresholds. Patients entering the “red” zone (white blood cell count <2 × 10⁹/L or absolute neutrophil count <1.5 × 10⁹/L) should generally not be treated with clozapine again. Clozapine-induced myocarditis is a hypersensitivity reaction that typically occurs in the third week of clozapine treatment, affecting approximately 2% of clozapine patients. Monitor patients for troponin, C-reactive protein, and electrocardiogram at baseline and on day 28 of treatment. Follow appropriate follow-up guidelines based on the patient's laboratory results. If myocarditis occurs, the patient should not be treated with clozapine again. Mechanism of Action: Clozapine (HF 1854) is an atypical antipsychotic whose therapeutic effect is achieved by balancing dopamine D2 receptor antagonism (reducing positive symptoms of schizophrenia) and serotonin 5-HT2A receptor antagonism (improving negative symptoms). It can also modulate other receptors (α1-adrenergic receptors, H1 receptors) to reduce side effects such as extrapyramidal symptoms [3,6]. Treatment Potential: Clozapine is indicated for treatment-resistant schizophrenia (patients unresponsive to typical antipsychotics). In a rat model of PCP (a preclinical model of schizophrenia), clozapine showed efficacy at doses of 5–10 mg/kg without inducing rigidity (a hallmark of extrapyramidal side effects) [3,5]. Chemical properties: Clozapine (HF 1854) is soluble in DMSO (10 mg/mL) and slightly soluble in physiological saline (0.5 mg/mL). It is stable for 24 hours at 4°C in aqueous solution at pH 7.4 [4].

C18H19CLN4

326.82

326.129

C, 66.15; H, 5.86; Cl, 10.85; N, 17.14

5786-21-0

Clozapine-d8; 1185053-50-2; Clozapine-d4; 204395-52-8; Clozapine N-oxide; 34233-69-7

135398737

White to light yellow solid powder

1.3±0.1 g/cm3

489.2±55.0 °C at 760 mmHg

182-185°C

249.6±31.5 °C

0.0±1.2 mmHg at 25°C

1.681

2.36

1

3

1

23

446

0

ClC1C([H])=C([H])C2=C(C=1[H])N=C(C1=C([H])C([H])=C([H])C([H])=C1N2[H])N1C([H])([H])C([H])([H])N(C([H])([H])[H])C([H])([H])C1([H])[H]

QZUDBNBUXVUHMW-UHFFFAOYSA-N

InChI=1S/C18H19ClN4/c1-22-8-10-23(11-9-22)18-14-4-2-3-5-15(14)20-16-7-6-13(19)12-17(16)21-18/h2-7,12,20H,8-11H2,1H3

3-chloro-6-(4-methylpiperazin-1-yl)-11H-benzo[b][1,4]benzodiazepine

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: ≥ 2.5 mg/mL (7.65 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.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (7.65 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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (7.65 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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 5%DMSO + Corn oil: 5.0mg/ml (15.30mM)

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