title: Atypical Antipsychotic (AAP)-induced Metabolic Toxicity: Study of Mechanisms, Biomarkers and Reversal Strategies
Creators: name: Sadiq, S
Publisher: name: University of Liverpool Repository
Year: 2018

description: Atypical antipsychotics are commonly prescribed for the treatment of psychiatric disorders such as schizophrenia. However, long term use of atypical antipsychotics lead to metabolic adverse effects such as weight gain, insulin resistance, and dyslipidemia, all of which are independent risk factors for cardiometabolic disease. Adipose tissue plays an important role in glucose and lipid homeostasis and has been linked to atypical antipsychotic-induced metabolic toxicity. The main aim of this thesis was to characterise the effect of atypical antipsychotics in vitro using murine 3T3-F442A preadipocyte cell lines and primary human adipocytes. Cells were incubated with clozapine (1-20μM), olanzapine (0.2-20μM) and aripiprazole (0.2-20μM) every 48 hours for 10 (3T3-F442A) or 13 (primary human adipocytes) days. Lipid accumulation was measured by Oil Red O assay. Protein and gene expression of peroxisome proliferator-activated receptor gamma and lipin 1 were measured by Western Blot and RT-PCR respectively. Adipokine secretion was measured by ELISA. Global lipidomic profiling of drug-treated adipocytes was undertaken using LC-MS and selected lipid species were validated by RT-PCR of enzymes that mediate the ceramide metabolism pathway. In differentiating 3T3-F442A cells clozapine (20μM:1.56 absorbance units±0.097; p=0.001) and olanzapine (20μM:1.57±0.14; p=0.07) but not aripiprazole showed an increase in lipid accumulation as compared to the vehicle. Clozapine but not olanzapine or aripiprazole upregulated the protein expression of peroxisome proliferator-activated receptor gamma (Mean fold change±SD; 307.34±26.30; p=0.0001) and lipin1 (213.46±26.43; p=0.02), and increased the secretion of adiponectin (736.77ng/ml±66.06; p=0.001; Vehicle: 323.26 ng/ml±53.07) and tumour necrosis factor alpha (45.24pg/ml±0.16; p=0.0001; Vehicle: 32.89pg/ml±0.34). In primary human adipocytes clozapine significantly increased lipid accumulation (1μM: 1.10±0.08; p=0.04) and leptin secretion (1μM: 1.06±0.10; p=0.04) but both clozapine and olanzapine led to a reduction in adiponectin secretion (Clozapine: 0.31±0.18; p=0.03, olanzapine: 0.39±0.21; p=0.01). Aripiprazole showed opposite effects to that of clozapine and olanzapine on lipid accumulation, gene expression and adipokine release. Drug uptake experiments showed that there was a 20-fold increase in clozapine uptake by differentiating primary human adipocytes (1293.4 pmol/million cells±170.56; p=0.006) in comparison to the murine adipocytes (64.25pmol/million cells±26.96). Co-incubation of clozapine with aripiprazole showed that aripiprazole significantly reversed clozapine-mediated reduction in secretion of adiponectin (1.23±0.09; p=0.04). Higher doses of both clozapine and olanzapine but not aripiprazole resulted in significant changes in the global lipidome profile of primary human adipocytes. Clozapine and olanzapine but not aripiprazole significantly decreased the ceramide 18 species and resulted in significant downregulation in the gene expression of ceramide biosynthesis enzymes, serine palmitoyltransferase long chain base subunit 2 (Clozapine: 0.60±0.13; p=0.003; Olanzapine: 0.62±0.16; p=0.007) and Delta-4-Desaturase Sphingolipid 1 (Clozapine: 0.67±0.18; p=0.01; Olanzapine: 0.83±0.11; p=0.02). In conclusion, we have shown that atypical antipsychotics may cause metabolic toxicity by directly affecting adipocyte function and metabolism; however, this could be potentially reversed in vitro by various therapeutic strategies. Atypical antipsychotics also significantly alter the adipocyte lipidome which may contribute to metabolic adverse effects in schizophrenia patients. The mechanisms and pathways identified in this study now need to be validated in in vivo and clinical models which will aid the identification of toxicity biomarkers in atypical antipsychotic-treated individuals.