Repeated Clozapine Administration Causes Extensive Changes to the Expression of Coding and Non-coding RNAs, Including...

Clozapine is arguably the most effective antipsychotic drug for the treatment of schizophrenia, but the mechanisms underlying its efficacy are poorly understood. Therefore, we perform deep RNA sequencing to test for differential transcription and exon use resulting from clozapine's effects in the mouse frontal cortex, and integrate our findings with known schizophrenia risk genes. We used a dose (4 mg/kg/day, i.p.) and duration (21 days) to approximate clinical exposure, followed by a 24-h washout to determine persistent changes resulting from biological remodeling. We observed significant (FDR < 0.05) differential expression of both mRNAs and long noncoding RNAs (lncRNAs), which were enriched in RNA processing and splicing pathways. Among the most significant lncRNAs, showing 2.3-fold upregulation, was the microRNA 124 host gene (Mir124a-1hg), a major source of miR-124, one of the most abundant microRNAs in the brain. Quantitative PCR analysis of the mature microRNAs miR-124-3p and miR-124-5p revealed a significant dose-dependent upregulation of miR-124-3p following 21-day repeated clozapine administration. RNA splicing was also profoundly impacted by clozapine, as revealed by differential exon use analysis, with mouse orthologs of 50 schizophrenia risk genes from the Psychiatric Genomics Consortium among the genes affected. These genes were enriched in "apical dendrite" and "distal axon" ontologies, supporting prior evidence that clozapine may target cortical pyramidal neuron deficits implicated in schizophrenia. Overall, this study demonstrates the profound effect of clozapine on cortical gene expression, affecting abundance of splicing of coding and non-coding transcripts. Future studies are needed to fully characterize our findings as potential preclinical markers of clozapine response.