A Comparative Study of the Pharmacokinetics of Clozapine N-Oxide and Clozapine N-Oxide Hydrochloride Salt in Rhesus Macaques.

Translating chemogenetic techniques from nonhuman primates to potential clinical applications has been complicated in part due to in vivo conversion of the chemogenetic actuator, clozapine N-oxide (CNO), to its pharmacologically active parent compound, clozapine, a ligand with known side effects, including five boxed warnings from the Food and Drug Administration. Additionally, the limited solubility of CNO requires high concentrations of potentially toxic detergents such as dimethylsulfoxide (DMSO). To address these concerns, pharmacokinetic profiling of commercially available CNO in DMSO (CNO-DMSO, 10% v/v DMSO in saline) and a water-soluble salt preparation (CNO-HCl, saline) was conducted in rhesus macaques. A time course of blood plasma and cerebrospinal fluid (CSF) concentrations of CNO and clozapine was conducted (30-240 minutes post-administration) following a range of doses (3-10 mg/kg, i.m. and/or i.v.) of CNO-DMSO or CNO-HCl. CNO-HCl resulted in 6- to 7-fold higher plasma concentrations of CNO compared to CNO-DMSO, and relatively less clozapine (3%-5% clozapine/CNO in the CNO-DMSO group and 0.5%-1.5% clozapine/CNO in the CNO-HCl group). Both groups had large between-subjects variability, pointing to the necessity of performing individual CNO pharmacokinetic studies prior to further experimentation. The ratio of CNO measured in the CSF was between 2% and 6% of that measured in the plasma and did not differ across drug preparation, indicating that CSF concentrations may be approximated from plasma samples. In conclusion, CNO-HCl demonstrated improved bioavailability compared with CNO-DMSO with less conversion to clozapine. Further investigation is needed to determine if brain concentrations of clozapine following CNO-HCl administration are pharmacologically active at off-target monoaminergic receptor systems in the primate brain.

Quetiapine affects neuropeptide Y and corticotropin-releasing hormone in cerebrospinal fluid from schizophrenia patients: relationship to depression and anxiety symptoms and to treatment response.

Cumulative evidence indicates that neuropeptides play a role in the pathophysiology of schizophrenia. Early data showed increased neuropeptide Y (NPY) in cerebrospinal fluid (CSF) from schizophrenia patients and data from rodents show that antipsychotic drugs modulate NPY levels in and release from selected rat brain regions. In view of these findings we investigated whether the atypical antipsychotic quetiapine, originally used as an antipsychotic but subsequently shown to be efficient also in major depressive disorder and in both poles of bipolar disorder, would affect NPY-like immunoreactivity (-LI), and corticotropin-releasing hormone (CRH)-LI levels in CSF of schizophrenia patients. NPY-LI and CRH-LI in CSF were determined in 22 patients with schizophrenia. Lumbar puncture was performed at baseline and again after 4 wk of quetiapine treatment (600 mg/d). Patients were assessed with the Positive and Negative Syndrome Scale (PANSS) at baseline and at weekly intervals. Quetiapine treatment was associated with a significant increase in NPY-LI (p<0.001) and decrease in CRH-LI (p<0.01). Stepwise multiple regression analysis revealed that ΔNPY-LI and ΔCRH-LI levels predicted 63% (p<0.001) of the variability of the ΔPANSS total score, ΔNPY-LI 42% (p<0.05) of the ΔPANSS anxiety items (G2) and ΔCRH-LI 40% (p=0.05) of the ΔPANSS depression items (G6). These results suggest that while quetiapine's effects on monoamines are probably related to its antipsychotic properties, the modulation of NPY and CRH accounts for its antidepressant and anxiolytic effects and can be markers of response.

High correlation between serum and cerebrospinal fluid olanzapine concentrations in patients with schizophrenia or schizoaffective disorder medicating with oral olanzapine as the only antipsychotic drug.

The primary aim of the present study was to investigate the relationship between steady state serum and cerebrospinal fluid (CSF) concentrations of olanzapine (OLA) and its metabolite 4'-N-desmethylolanzapine (DMO) in patients with schizophrenia or schizoaffective disorder treated with oral OLA as the only antipsychotic drug. The influence of smoking, gender, age, as well as polymorphisms in cytochrome P450 CYP2D6, CYP1A2, and ABCB1 genes on the serum and CSF drug levels was also analyzed. Thirty-seven white outpatients (10 smokers and 27 nonsmokers) were included. From 29 of them, CSF was collected successfully. A strong correlation (Spearman rank correlation [rs] = 0.93; P < 0.05) was found between serum and CSF concentrations of OLA and a somewhat weaker correlation (rs = 0.5; P < 0.05) between those of DMO. The CSF concentrations of OLA and DMO were on average 12% and 16% of those in serum. Extensive metabolizers of CYP2D6 had higher (P < 0.05) daily doses than poor metabolizers when the influence of smoking was taken into account. Smokers had lower (P < 0.01) concentration-to-dose ratios of OLA in serum (mean, 2.23 ng/mL per mg vs 3.32 ng/mL per mg) and CSF (0.27 ng/mL per mg vs 0.41 ng/mL per mg) than nonsmokers. The concentration-to-dose ratio for serum DMO decreased with increasing age (rs = -0.41; P < 0.05). Carriers of ABCB1 1236T/2677T/3435T haplotype had higher serum (mean, 37.7 ng/mL vs 22.5 ng/mL; P = 0.035) and CSF (4.7 ng/mL vs 2.6 ng/mL; P = 0.018) OLA concentrations than patients without this haplotype. The present study shows a strong correlation between serum and CSF concentrations of OLA, indicating that concentrations of OLA in serum reflect those in CSF.

Metabolism and Distribution of Clozapine-N-oxide: Implications for Nonhuman Primate Chemogenetics.

The use of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) in neuroscience has rapidly expanded in rodent studies but has lagged behind in nonhuman primate (NHP) experiments, slowing the development of this method for therapeutic use in humans. One reason for the slow adoption of DREADD technology in primates is that the pharmacokinetic properties and bioavailability of clozapine-n-oxide (CNO), the most commonly used ligand for human muscarinic (hM) DREADDs, are not fully described in primates. We report an extensive pharmacokinetic study using subcutaneous (SC) administration of CNO in five adult rhesus monkeys. CNO reached maximal plasma and cerebrospinal fluid (CSF) concentrations within 2 h after injection, with an observed dose-dependent increase in levels following a 3 and 10 mg/kg SC dose. Since CSF concentrations were below values predicted from unbound plasma concentrations, we investigated whether CNO was restricted from the CNS through active transport at the blood-brain barrier. In vitro assessment demonstrated that CNO is a substrate for P-glycoprotein (Pgp; efflux ratio, 20), thus providing a likely mechanism limiting CNO levels in the CNS. Furthermore, CNO is metabolized to the psychoactive compounds clozapine and n-desmethylclozapine in monkeys. The concentrations of clozapine detected in the CSF are sufficient to activate several types of receptor (including the hM-DREADDs). Our results suggest that CNO metabolism and distribution may interfere with reproducibility and interpretation of DREADD-related experiments in NHPs and calls for a re-evaluation of the use of CNO in DREADD-related experiments in NHPs along with the need to test alternative compounds.

CSF and serum concentrations of clozapine and its demethyl metabolite: a pilot study.

With the aim of exploring putative correlations between serum and CSF levels of clozapine and its demethyl metabolite, lumbar puncture was performed on four male and five female schizophrenic patients during long-term treatment with clozapine. Three consecutive 6-ml fractions were collected after at least 8 h of bedrest and fasting. On comparing serum and CSF levels, a correlation was found for norclozapine in the third (13-18 ml) CSF fraction. Norclozapine in the first (0-6 ml) CSF correlated significantly with height. The CSF/serum ratio of clozapine in the first fraction was correlated significantly with body weight. No correlations were found between serum levels of clozapine and norclozapine, or between the serum and CSF levels of clozapine. The study suffers from a small number of patients (for ethical reasons), but the present results might be explicable if the first (0-6 ml) CSF fraction represents a cul-du-sac of the CSF, mirroring the previous day's drug levels. The second fraction, then, will represent the CSF level in the steady state during the night.

Determination of olanzapine in a postmortem case.

Olanzapine, a new antipsychotic agent, was approved by the U.S. Food and Drug Administration in 1996 for use in the treatment of schizophrenia. It is structurally similar to clozapine, has a low incidence of extrapyramidal effects, and is effective in treating both the positive and negative symptoms of schizophrenia. This paper describes the determination of olanzapine in biological specimens obtained from the autopsy of a 35-year-old white male found dead in bed at a psychiatric facility. In the months prior to his death, the deceased was prescribed multiple medications, including olanzapine. Olanzapine was identified qualitatively by full scan gas chromatography-mass spectrometry, with quantitative analysis performed by liquid-liquid extraction followed by dual-column gas chromatography. The following concentrations were determined in the specimens analyzed: heart blood, 550 ng/mL; bile, 6346 ng/mL; and gastric contents, 157 ng/mL. Vitreous humor, cerebrospinal fluid, and urine specimens were negative. Although steady-state plasma concentrations of 10-25 ng/mL olanzapine have been reported, effective levels are known to be highly variable and a plasma concentration of 300 ng/mL has been tolerated without adverse effects. Based upon the autopsy, toxicological findings, and case investigation, the cause of death was determined to be intramyocardial arteriosclerosis with severe stenosis of the nodal artery due to hypertensive cardiovascular disease, and the manner was natural.

A validated UHPLC-diode array detector method for the bioanalysis of atypical antipsychotics in whole blood, urine and cerebrospinal fluid following SPE.

BACKGROUND: Risperidone, quetiapine, amisulpride and aripiprazole are atypical antipsychotics increasingly prescribed in Greece. RESULTS: We have developed and validated a UHPLC-diode array detector method for the determination of these drugs in whole blood, urine and cerebrospinal fluid, following SPE. The separation was achieved within 7 min. Validation was performed in terms of linearity, selectivity, accuracy, precision and stability. Following validation, the method was applied to post-mortem blood, urine and cerebrospinal fluid samples. CONCLUSION: The method proved fast and simple for the determination of four atypical antipsychotic drugs in whole blood, urine and cerebrospinal fluid. To the best of our knowledge, this is the only available method for the simultaneous determination of these drugs in cerebrospinal fluid.

Cerebroventricular clozapine would be a viable treatment modality for clozapine-dependent schizophrenia patients with neutropenia.

The atypical antipsychotic clozapine is very effective for treatment of schizophrenia, but it causes agranulocytosis requiring drug cessation in up to 2% of cases. There has been some success rechallenging with clozapine at a later date or giving granulocyte colony stimulating factor or lithium while continuing clozapine. However, there are still some patients for whom these strategies do not work yet who cannot be controlled on other medications. This paper proposes that for such individuals, cerebroventricular administration of clozapine via Ommaya catheters could allow continued use of clozapine therapy. Direct infusion into cerebrospinal fluid means far smaller amounts of drug would be needed for efficacy, and clozapine concentrates in the central nervous system where it would not be exposed to bone marrow stem cells to cause agranulocytosis. This treatment paradigm would also provide a means for court-ordered clozapine therapy and a possible delivery system for future therapeutics based on trophic factors such as brain-derived neurotrophic factor.

Cytochrome P450 and ABCB1 genetics: association with quetiapine and norquetiapine plasma and cerebrospinal fluid concentrations and with clinical response in patients suffering from schizophrenia. A pilot study.

Variability in response to atypical antipsychotic drugs is due to genetic and environmental factors. Cytochrome P450 (CYP) isoforms are implicated in the metabolism of drugs, while the P-glycoprotein transporter (P-gp), encoded by the ABCB1 gene, may influence both the blood and brain drug concentrations. This study aimed to identify the possible associations of CYP and ABCB1 genetic polymorphisms with quetiapine and norquetiapine plasma and cerebrospinal fluid (CSF) concentrations and with response to treatment. Twenty-two patients with schizophrenia receiving 600 mg of quetiapine daily were genotyped for four CYP isoforms and ABCB1 polymorphisms. Quetiapine and norquetiapine peak plasma and CSF concentrations were measured after 4 weeks of treatment. Stepwise multiple regression analysis revealed that ABCB1 3435C > T (rs1045642), 2677G > T (rs2032582) and 1236C > T (rs1128503) polymorphisms predicted plasma quetiapine concentrations, explaining 41% of the variability (p = 0.001). Furthermore, the ABCB1 polymorphisms predicted 48% (p = 0.024) of the variability of the Δ PANSS total score, with the non-carriers of the 3435TT showing higher changes in the score. These results suggest that ABCB1 genetic polymorphisms may be a predictive marker of quetiapine treatment in schizophrenia.

Relationship between dopamine D2 receptor occupancy, clinical response, and drug and monoamine metabolites levels in plasma and cerebrospinal fluid. A pilot study in patients suffering from first-episode schizophrenia treated with quetiapine.

Combining measurements of the monoamine metabolites in the cerebrospinal fluid (CSF) and neuroimaging can increase efficiency of drug discovery for treatment of brain disorders. To address this question, we examined five drug-naïve patients suffering from schizophrenic disorder. Patients were assessed clinically, using the Positive and Negative Syndrome Scale (PANSS): at baseline and then at weekly intervals. Plasma and CSF levels of quetiapine and norquetiapine as well CSF 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxyindole-acetic acid (5-HIAA) and 3-methoxy-4-hydroxyphenylglycol (MHPG) were obtained at baseline and again after at least a 4 week medication trail with 600 mg/day quetiapine. CSF monoamine metabolites levels were compared with dopamine D(2) receptor occupancy (DA-D(2)) using [(18)F]fallypride and positron emission tomography (PET). Quetiapine produced preferential occupancy of parietal cortex vs. putamenal DA-D(2), 41.4% (p<0.05, corrected for multiple comparisons). DA-D(2) receptor occupancies in the occipital and parietal cortex were correlated with CSF quetiapine and norquetiapine levels (p<0.01 and p<0.05, respectively). CSF monoamine metabolites were significantly increased after treatment and correlated with regional receptor occupancies in the putamen [DOPAC: (p<0.01) and HVA: (p<0.05)], caudate nucleus [HVA: (p<0.01)], thalamus [MHPG: (p<0.05)] and in the temporal cortex [HVA: (p<0.05) and 5-HIAA: (p<0.05)]. This suggests that CSF monoamine metabolites levels reflect the effects of quetiapine treatment on neurotransmitters in vivo and indicates that monitoring plasma and CSF quetiapine and norquetiapine levels may be of clinical relevance.

CSF concentrations and clinical effects following intravenous dixyrazine premedication.

Single concurrent plasma and lumbar spinal fluid samples were collected from twenty-two patients given IV dixyrazine 0.15 mg/kg, 10-106 min earlier in an open study. Dixyrazine penetrated rapidly into the spinal fluid; the CSF/P ratio was 0.3 by 10 min after drug administration and persisted at that level during the study period. The uncontrolled sedative and anxiolytic effects assessed by the patients were slight, and were correlated with the CSF drug concentrations only during the first 30 min after medication.

Neurotensin-like immunoreactivity in cerebrospinal fluid of patients with schizophrenia, depression, anorexia nervosa-bulimia, and premenstrual syndrome.

Neurotensin (NT) concentrations in cerebrospinal fluid (CSF) were measured by a sensitive and specific radioimmunoassay in psychiatric patients and age- and sex-matched normal controls. No increase in CSF NT concentrations was observed after antipsychotic drug treatment. CSF NT concentrations were significantly lower in one group of schizophrenic subjects. NT concentrations were unaltered in patients with depression, anorexia/bulimia, or premenstrual syndrome, and no rostral-caudal gradient for NT in CSF was evident. NT concentrations were not related to age or sex, and probenecid treatment did not alter CSF NT concentrations. Finally CSF NT concentrations were unaltered in paranoid schizophrenic subjects. These findings confirm and extend previous studies of CSF NT that showed certain patients with schizophrenia, nonparanoid type, have reduced CSF concentrations of this tridecapeptide.