Treatment-resistant schizophrenia (TRS) poses a substantial clinical challenge for which clozapine (CLOZ) is the only effective treatment. However, clinicians may hesitate to prescribe CLOZ for TRS patients with epilepsy due to its presence because CLOZ is contraindicated in patients with uncontrolled epilepsy in some countries including Japan. A 36-year-old male with TRS and focal epilepsy underwent CLOZ therapy. CLOZ was initiated at 6.25 mg and gradually titrated to 250 mg over 3 months, achieving a plasma concentration of 449 ng/mL. Psychotic symptoms improved without worsening of seizure frequency or severity. Mild side effects such as drowsiness and drooling were manageable. Long-term video electroencephalography monitoring detected spike-and-wave activity centered in the frontal region, but did not detect any overt epileptic seizures. We report a case of TRS with comorbid epilepsy in which CLOZ therapy was both effective and safe.

Clozapine levels in inpatients with schizophrenia are not a predictor of rehospitalization: a naturalistic retrospective study.

A retrospective comparative analysis of the clinical response to clozapine between patients with early and late treatment-resistant schizophrenia.

Abstract The accurate determination of clozapine (CLZ), an essential antipsychotic drug, remains challenging due to complex pharmaceutical matrices and the need for high sensitivity. Conventional methods like HPLC and UV-Vis are limited by time-consuming procedures and unsuitability for on-site analysis. To address this, a novel hybrid nanocomposite based on a chitosan Schiff base functionalized with 2-(2-acetamidophenyl)-2-oxoacetic acid (Chs-KA), β-cyclodextrin (β-CD), and zinc oxide nanoparticles (ZnO NPs) was successfully synthesized. Structural characterization confirmed the formation of the Chs-KA Schiff base via imine linkage (FTIR band at ~ 1640 cm⁻¹), while XRD analysis revealed the incorporation of crystalline ZnO with a wurtzite structure and an average crystallite size of 8–20 nm. This composite was employed to fabricate a modified carbon paste electrode (Chs-KA/β-CD/ZnO NPs/CPE) for the sensitive detection CLZ. The sensor demonstrated a proton-coupled electron transfer mechanism (slope of − 47 mV/pH) with adsorption-controlled kinetics (heterogeneous electron transfer rate constant, ks = 2.67 s⁻¹). Under optimized conditions, the electrode exhibited a wide linear response range from 0.5 nM to 7.0 µM, a very low detection limit of 0.13 nM, and high selectivity against common interferents (signal change < 5%). The sensor showed excellent reproducibility (RSD = 2.05%) and retained 95.64% of its initial response after one month. Practical applicability was validated by determining CLZ in commercial tablet formulations, with recovery rates ranging from 97.0% to 102.0%. This work provides a powerful platform for pharmaceutical quality control and establishes a design strategy for advanced biopolymer-based electrochemical sensors. Future work includes application to other psychoactive drugs, biological fluid monitoring, and commercial scalability.

Oxidative stress is intrinsically linked to mental disorders, involving an imbalance between reactive species and antioxidant defenses, where catalase is an essential, ubiquitous antioxidant enzyme. The pleiotropic effects of antipsychotic drugs, used for schizophrenia and mood disorders, are not fully elucidated at the molecular level. This study characterized the binding of a highly effective but potentially dangerous antipsychotic, clozapine (CLZ), to commercial bovine liver catalase (BLC). Using various spectroscopic methods under simulated physiological conditions, we found a moderate binding affinity of CLZ for BLC (Ka = 1.4 × 10-5 M-1), subtly influencing the protein's secondary and tertiary structures and slightly increasing its thermal stability. CLZ efficiently protected BLC against free-radical-induced oxidation and preserved its catalytic activity for decomposing toxic hydrogen peroxide. The effect of CLZ on BLC antioxidant activity was dual: no significant effect at lower, physiologically relevant concentrations, but significant inhibition at saturating, toxic drug concentrations. Molecular docking and molecular dynamics results indicated the presence of two specific binding sites within BLC monomers, one located near its active site. In conclusion, our in vitro results indicate that CLZ's specific binding to BLC can be both beneficial and potentially harmful, and that this effect is dose-dependent.

Microstructural and diffusion tensor imaging of clozapine for treatment-resistant schizophrenia.

Clozapine is widely used to treat schizophrenia, but it has low oral bioavailability (<27%), degradation in GI environment, and high hepatic first-pass metabolism. The present research work aimed to overcome these issues by designing tailored clozapine loaded nanoparticles (NPs) and to deliver them to the brain via the intranasal pathway, to enhance brain targeting of antipsychotic drugs. Chitosan–sodium tri polyphosphate and chitosan–sodium alginate NPs loaded with Clozapine (CZ) were prepared by using ionotropic gelation and ionotropic pre-gelation methods. SEM showed spherical-shaped NPs, while the average dynamic size was measured by dynamic light scattering. Optimized clozapine loaded nanoparticles (C2,C7) showed 56% and 55% encapsulation efficiency, respectively, with particle sizes of (453 - 425)nm. The in vitro drug release from formulas exhibited sustained release of clozapine, were 69.2% and 98.7% respectively, within 24h. The release profile was characterized by an initial fast release in phosphate buffer, followed by a continuous release phase; the drug release mechanism from polymers was in correspondence with Fickian diffusion. CZ-loaded NPs are a promising strategy for managing schizophrenia. Nose-to-brain pathway enhances safety and patient compliance of CZ.

Clozapine (CLZ) is an atypical antipsychotic mainly prescribed for treatment-resistant schizophrenia. Beyond psychotic symptoms, patients often exhibit persistent cognitive impairments across domains such as attention, learning, and memory. The mechanisms by which CLZ may influence cognition and provide neuroprotection are not fully elucidated. Accordingly, this study examined how CLZ modulates lipopolysaccharide (LPS)-induced neurotoxicity in rats. Rats were administered LPS to induce cognitive impairment and subsequently treated with CLZ. Behavioral assessments were performed using maze tests (elevated plus-maze (EPM), novel object recognition (NOR), and Y-maze). Biochemical analyses included cholinergic function (acetylcholine (ACh)), neurodegeneration-associated enzymes (glycogen synthase kinase-3 beta (GSK-3β), β-site amyloid precursor protein cleaving enzyme-1 (BACE-1), and dipeptidyl peptidase-4 (DPP-4)), oxidative stress markers (lipid Peroxidation (LPO), catalase, and reduced glutathione (GSH)), and apoptotic proteins (B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), and cleaved Caspase-3 (c-Caspase-3)). CLZ treatment markedly improved performance in EPM, NOR, and Y-maze tasks, indicating recovery of cognitive function in LPS-exposed rats. At the molecular level, CLZ enhanced ACh levels, upregulated the anti-apoptotic protein Bcl-2, and restored antioxidant defenses (catalase and GSH). Conversely, CLZ reduced LPS-induced neurotoxicity by lowering GSK-3β activity, LPO, and pro-apoptotic markers (Bax and c-Caspase-3). The findings demonstrate that CLZ exerts neuroprotective effects in an LPS-induced rat model, improving cognition through modulation of cholinergic transmission, oxidative stress, and apoptosis pathways. These results clarify key mechanistic pathways through which CLZ may exert cognitive benefits and highlight its potential relevance for improving schizophrenia-related cognitive dysfunction. Further molecular studies are warranted to confirm and extend these observations toward clinical translation.

Construction of multi-component covalent organic framework coated stainless steel fiber for efficient microextraction of psychiatric drugs: experiments and clinical applications.

BackgroundTreatment‐resistant schizophrenia (TRS) is often accompanied by poor insight and impaired decision‐making capacity (DMC), complicating the initiation of clozapine (CLZ) despite clear clinical indications. In Japan, CLZ utilization is markedly lower than in many foreign countries, partly due to concerns about adverse effects, mandatory blood monitoring, and difficulties in obtaining informed consent from patients with impaired DMC. Although CLZ can legally be initiated only with family consent, sustained treatment generally requires the patient's own agreement. Electroconvulsive therapy (ECT) may alleviate acute psychotic symptoms and temporarily improve DMC, thereby enabling informed participation in treatment decisions.Case PresentationA woman in her late thirties with TRS persistently refused CLZ. On readmission, her Clinical Global Impression‐Severity (CGI‐S) score was 7. Pseudo‐TRS was partly ruled out using a long‐acting injectable antipsychotic (LAI). Following a multidisciplinary case conference in the psychiatric department and family proxy consent, ECT was initiated during involuntary hospitalization for medical care and protection in accordance with clinical practice recommendations for ECT endorsed by the Japanese Society of Psychiatry and Neurology (JSPN). After three ECT sessions, the patient's DMC improved sufficiently to allow voluntary consent to CLZ initiation. Twelve ECT sessions were administered while CLZ was titrated to 225 mg/day. The CGI‐S score improved to 3, no serious adverse events occurred, and she was discharged approximately 120 days after admission with stable outpatient follow‐up.ConclusionThis case demonstrates that ECT can function as a capacity‐restoring intervention enabling voluntary CLZ initiation in selected TRS patients who initially lack DMC. In the context of low CLZ utilization in Japan, a staged approach combining acute stabilization and pharmacological consolidation may provide a clinically and ethically acceptable pathway when supported by multidisciplinary review and procedural safeguards.

Population pharmacokinetic analysis for simultaneous fit of clozapine and norclozapine concentrations in adult psychiatric patients.

Clozapine (CLZ), as an efficient atypical antipsychotic drug, faces significant challenges in clinical application, such as a narrow treatment window and huge pharmacokinetic differences between individuals. Therefore, developing sensitive and reliable detection methods for CLZ is crucial for ensuring therapeutic efficacy and reducing medication risks. Strongly blue-fluorescent g-C3N4 nanosheets (g-C3N4Ns) were successfully prepared from bulk g-C3N4 through a two-step process involving protonation and subsequent ultrasonic exfoliation. Leveraging the charge-transfer-induced dynamic quenching of their fluorescence by CLZ, we developed a sensitive analytical method using these nanosheets as a fluorescent probe. The experimental results indicate that this method exhibited a wide linear range (0.02-500 µM) and a low detection limit of 3 nM. When applied to the analysis of bovine serum and clozapine tablets, the spiked recovery rate of CLZ was within the range of 92.0-104.7%, with a relative standard deviation (RSD) of less than 5%, and the results were satisfactory. Moreover, the probe demonstrated excellent selectivity and anti-interference capability against multiple potential interferents. In summary, the g-C3N4Ns fluorescence sensing platform developed herein integrates rapid response, high sensitivity, and excellent reliability and has been successfully applied to CLZ detection in complex matrices. This work establishes a solid foundation for future applications in real human plasma samples and ultimately serves therapeutic drug monitoring (TDM).

Clozapine (CLZ) is the gold standard for treatment-resistant schizophrenia (TRS), but its use in Japan is limited by strict monitoring and titration-phase adverse events. This prospective 12-week study evaluated longitudinal trends in CLZ/norclozapine (NCLZ) levels, inflammatory markers, metabolic indices, and psychiatric symptoms. Twenty-one inpatients with TRS were analyzed. To focus on standard titration, patients with inflammatory symptoms (e.g., fever) requiring discontinuation were excluded. Serum CLZ and NCLZ were measured weekly via LC-MS/MS. Clinicians were blinded to these levels; dosing was guided solely by clinical observation. IL-6, HOMA-IR, TG/HDL-C ratios, and PANSS scores were assessed at baseline and designated intervals. CLZ and NCLZ concentrations increased throughout the 12-week period. Significant sex differences emerged in CLZ concentration-to-dose (C/D) ratios, with females exhibiting significantly higher levels than males starting at week 2 (p < 0.05). While positive symptoms significantly improved (p < 0.05), no specific longitudinal correlations were found between CLZ/NCLZ levels and changes in IL-6, metabolic indices, or total PANSS scores. A "start low, go slow" titration approach can effectively achieve therapeutic concentrations even without real-time therapeutic drug monitoring. However, the significantly higher concentrations observed in female subjects suggest that more cautious dose titration is necessary for female patients, likely due to hormonal influences on metabolic enzymes. Further research is needed on the relationship between clozapine dosage, plasma concentrations and inflammatory side effects.

Schizophrenia affects a significant proportion of individuals, wherein a subset of patients is described as treatment-resistant. Clozapine (CLOZ) is an atypical antipsychotic, which is reserved for these patients and is superior in its anti-suicidal activity. However, it carries numerous serious warnings and is well-known for its risk of drug-induced agranulocytosis. The mechanism of toxicity is unclear and could be due to CLOZ's protein covalent binding and off-target effects through its reactive metabolites produced from neutrophil myeloperoxidase (MPO) activity. We hypothesize that identifying and analyzing the protein-CLOZ adducts will contribute to our understanding of toxicity pathways. We have developed a novel clickable CLOZ (Click-CLOZ) derivative and have designed click chemistry protocols for protein identification. The HL-60 (human promyelocytic leukemia) cell line and isolated human neutrophils express MPO significantly and were used to identify the protein covalent targets of Click-CLOZ. In HL-60 cells, LC/MS analysis revealed many Click-CLOZ-bound proteins (compared to the vehicle control). Some captured proteins were known for their roles in DNA replication, immune responses and oxidative stress, such as cathepsin G, MPO, ribophorin I and P1-MCM3. In neutrophils, Click-CLOZ-bound proteins included MPO, S100, and DEFA1B, which are also associated with neutrophil-mediated oxidative stress and immune responses. In conclusion, the application of click chemistry proteomics has facilitated a novel approach to identify multiple CLOZ-bound protein targets that will be used to advance our understanding of the toxicity of CLOZ.

L-DOPA-induced dyskinesia (LID) is a common complication in the treatment of Parkinson's disease (PD), characterized by involuntary excessive movements. The traditional Abnormal Involuntary Movement Scale (AIMs), used for quantifying abnormal involuntary movements, relies heavily on manual observation and is highly subjective. Unsupervised behavior classification typically requires joint modeling on the entire dataset, making it inflexible when dealing with new samples. Here, we propose an automated behavioral recognition framework integrating multi-view 3D motion reconstruction with a hypergraph self-attention neural network to precisely delineate LID behavioral phenotypes and evaluate pharmacological interventions. Using a synchronized four-camera setup, we collected large-scale motion data from WT, PD, and LID mice, tracking 16 key body points to reconstruct accurate 3D trajectories. By combining unsupervised clustering with manual annotation, we established a standardized behavioral database. We introduced a spatiotemporal hypergraph neural network model incorporating a self-attention mechanism, which demonstrated excellent recognition accuracy across all behaviors and effectively distinguished the behavioral profiles of WT, PD, and LID mice. Based on this, we compared the behavioral differences in treatment effects between amantadine (AMAN) and clozapine (CLZ). Overall, our automated 3D behavioral analysis framework offers a high-throughput, objective, and precise approach to behavioral quantification, presenting a powerful tool for unraveling the mechanisms underlying LID and other movement disorders, as well as for advancing pharmacological research.

Clozapine (CLZ); an atypical antipsychotic drug, is well known to have a significant role in managing schizophrenic patients with substance use disorder (SUD). Unfortunately, many patients are deprived of CLZ benefits due to its limited prescription. This is based upon concerns regarding the critical side effects of CLZ in case of overdosing especially, with the lack of accessible therapeutic drug monitoring (TDM) tools. In this contribution, a simple, accurate and sensitive electrochemical method is proposed for CLZ assay in human saliva. Unlike previously reported methods for TDM of CLZ that depends on invasive matrices as plasma and urine, this method employs electrochemical approaches in exploring human saliva as a patient-friendly alternative for assessing CLZ. The proposed method employs differential pulse voltammetry (DPV) with a sensitive and selective Ag-doped ZnO nanoparticles based carbon paste electrode (CPE). The adopted electrochemical sensor has not been previously reported for CLZ determination, despite it offers enhanced sensitivity together with simple synthesis. The synthesized nanoparticles were characterized through Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX). The developed sensor was optimized and validated as per FDA guidelines of bioanalytical methods. The linear range in saliva was 0.31–3.67 µmol/L and the lower limit of quantitation (LLOQ) was 0.31 µmol/L. The high reliability and applicability of the suggested method has strong potential to be integrated in a point-of-care testing (POCT) device to introduce more accessible TDM that enables smooth TDM of CLZ. Therefore, it opens pathways for broader and safe use of CLZ.Graphical abstract

Development and validation of simultaneous quantification of total and free clozapine and its two metabolites in human plasma using ultra high-performance liquid chromatography coupled to tandem mass spectrometry.

Myotube-specific vulnerability to clozapine.

Clozapine (CZP) is not commonly prescribed mainly because of the risk of serious adverse effects, particularly neutropenia. It can also cause blood disorders, ranging from mild to severe, affecting different blood cell types. The aim of this study was to investigate the incidence of hematological abnormalities in a Pakistani cohort of patients treated with antipsychotics using a comprehensive hematological monitoring program. The study included 288 patients categorized into three clinical groups based on antipsychotic treatment: Group TA (typical antipsychotics), Group AA (atypical antipsychotics), and Group CZP. During the initial 20-week treatment period, patients were regularly monitored hematological to assess changes in their blood profiles. The occurrence of various hematological abnormalities was investigated and analyzed using statistical models. Blood dyscrasias were observed more frequently in patients in the CZP group than in the other groups. Specifically, neutropenia was seen in a few patients in group CZP. Notably, a positive clinical response to CZP treatment was significantly correlated with transient leukocytosis, transient neutrophilia, and persistent anemia. In contrast, poor clinical response to CZP medication was associated with transient leukopenia. CZP treatment resulted in more blood disorders than TA or AA treatments. While blood abnormalities were common with CZP, severe neutropenia was rare. Certain blood changes were linked to better treatment outcomes. Therefore, regular blood monitoring is recommended to optimize treatment effectiveness and manage side effects.

A sensitive colorimetric immunosensor for clozapine, norclozapine, and clozapine-N-oxide simultaneous detection based on monoclonal antibodies preparation with homogeneous cross-reactivity.