Identification of EP300 as a Key Gene Involved in Antipsychotic-Induced Metabolic Dysregulation Based on Integrative Bioinformatics Analysis of Multi-Tissue Gene Expression Data.

Antipsychotics (APs) are associated with weight gain and other metabolic abnormalities such as hyperglycemia, dyslipidemia and metabolic syndrome. This translational study aimed to uncover the underlying molecular mechanisms and identify the key genes involved in AP-induced metabolic effects. An integrative gene expression analysis was performed in four different mouse tissues (striatum, liver, pancreas and adipose) after risperidone or olanzapine treatment. The analytical approach combined the identification of the gene co-expression modules related to AP treatment, gene set enrichment analysis and protein-protein interaction network construction. We found several co-expression modules of genes involved in glucose and lipid homeostasis, hormone regulation and other processes related to metabolic impairment. Among these genes, EP300, which encodes an acetyltransferase involved in transcriptional regulation, was identified as the most important hub gene overlapping the networks of both APs. Then, we explored the genetically predicted EP300 expression levels in a cohort of 226 patients with first-episode psychosis who were being treated with APs to further assess the association of this gene with metabolic alterations. The EP300 expression levels were significantly associated with increases in body weight, body mass index, total cholesterol levels, low-density lipoprotein cholesterol levels and triglyceride concentrations after 6 months of AP treatment. Taken together, our analysis identified EP300 as a key gene in AP-induced metabolic abnormalities, indicating that the dysregulation of EP300 function could be important in the development of these side effects. However, more studies are needed to disentangle the role of this gene in the mechanism of action of APs.

Integrative DNA Methylation and Gene Expression Analysis of Cognitive Behavioral Therapy Response in Children and Adolescents with Obsessive-Compulsive Disorder; a Pilot Study.

PURPOSE: Here, we propose an integrative analysis of genome-wide methylation and gene expression to provide new insight into the biological mechanisms of Cognitive behavioral therapy (CBT) in pediatric obsessive-compulsive disorder (OCD). PATIENTS AND METHODS: Twelve children and adolescents with OCD receiving CBT for the first time were classified as responders or non-responders after eight weeks of CBT. Differentially methylated positions (DMPs) and gene co-expression modules were identified using specific R software packages. Correlations between the DMPs and gene co-expression modules were investigated. RESULTS: Two genes were enriched with significant DMPs (Δß > ± 0.2, FDR-adjusted p-value < 0.05): PIWIL1 and MIR886. The yellowgreen module of co-expressed genes was associated with CBT response (FDR-adjusted p-value = 0.0003). Significant correlations were observed between the yellowgreen module and the CpGs in PIWIL1 and MIR886 (p < 0.008). Patients showing hypermethylation in these CpGs presented an upregulation in the genes in the yellowgreen module. CONCLUSION: Taken together, the preliminary results of this systems-level approach, despite the study limitations, provide evidence that the epigenetic regulation of ncRNAs could be a predictor of CBT response. LIMITATIONS: The sample size limited the statistical power, and given that the study was hypothesis-driven, our results should be seen as preliminary.

DNA Methylation of Fluoxetine Response in Child and Adolescence: Preliminary Results.

PURPOSE: The search for predictors of antidepressant response is gaining increasing attention, with epigenetic markers attracting a great deal of interest. We performed a genome-wide study assessing baseline differences in DNA methylation between Responders and Non-Responders. PATIENTS AND METHODS: Twenty-two children and adolescents, receiving fluoxetine treatment for the first time, were classified as Responders or Non-Responders according to CGI-I score after 8 weeks of fluoxetine treatment. Genome-wide DNA methylation was profiled using the Illumina Infinium MethylationEPIC BeadChip Kit and analyzed using the Chip Analysis Methylation Pipeline (ChAMP). RESULTS: We identified 21 CpG sites significantly (FDR<0.05) associated with fluoxetine response that showed meaningful differences (Δß> ±0.2) in methylation level between Responders and Non-Responders. Two genes, RHOJ (Ras Homolog Family Member J) and OR2L13 (Olfactory Receptor family 2 subfamily L member 13), presented more than one significant CpG sites. CONCLUSION: Our findings provide new insights into the molecular mechanisms underlying the complex phenotype of antidepressant response, indicating that methylation at specific genes could be a promising biomarker that needs further replication in large cohorts.

Identifying key transcription factors for pharmacogenetic studies of antipsychotics induced extrapyramidal symptoms.

INTRODUCTION: We explore the transcription factors involved in the molecular mechanism of antipsychotic (AP)-induced acute extrapyramidalsymptoms (EPS) in order to identify new candidate genes for pharmacogenetic studies. METHODS: Protein-protein interaction (PPI) networks previously created from three pharmacogenomic models (in vitro, animal, and peripheral blood inhumans) were used to, by means of several bioinformatic tools; identify key transcription factors (TFs) that regulate each network. Once the TFs wereidentified, SNPs disrupting the binding sites (TFBS) of these TFs in the genes of each network were selected for genotyping. Finally, SNP-basedassociations with EPS were analyzed in a sample of 356 psychiatric patients receiving AP. RESULTS: Our analysis identified 33 TFs expressed in the striatum, and 125 SNPs disrupting TFBS in 50 genes of our initial networks. Two SNPs (rs938112,rs2987902) in two genes (LSMAP and ABL1) were significantly associated with AP induced EPS (p < 0.001). These SNPs disrupt TFBS regulated byPOU2F1. CONCLUSION: Our results highlight the possible role of the disruption of TFBS by SNPs in the pharmacological response to AP.

The positive allosteric modulator of the mGlu2 receptor JNJ-46356479 partially improves neuropathological deficits and schizophrenia-like behaviors in a postnatal ketamine mice model.

Current antipsychotics have limited efficacy in controlling cognitive and negative symptoms of schizophrenia (SZ). Glutamatergic dysregulation has been implicated in the pathophysiology of SZ, based on the capacity of N-methyl-D-aspartate receptor (NMDAR) antagonists such as ketamine (KET) to induce SZ-like behaviors. This could be related to their putative neuropathological effect on gamma-aminobutyric (GABAergic) interneurons expressing parvalbumin (PV), which would lead to a hyperglutamatergic condition. Metabotropic glutamate receptor 2 (mGluR2) negatively modulates glutamate release and has been considered a potential clinical target for novel antipsychotics drugs. Our aim was to evaluate the efficacy of JNJ-46356479 (JNJ), a positive allosteric modulator (PAM) of the mGluR2, in reversing neuropathological and behavioral deficits induced in a postnatal KET mice model of SZ. These animals presented impaired spontaneous alternation in the Y-maze test, suggesting deficits in spatial working memory, and a decrease in social motivation and memory, assessed in both the Three-Chamber and the Five Trial Social Memory tests. Interestingly, JNJ treatment of adult mice partially reversed these deficits. Mice treated with KET also showed a reduction in PV+ in the mPFC and dentate gyrus together with an increase in c-Fos expression in this hippocampal area. Compared to the control group, mice treated with KET + JNJ showed a similar PV density and c-Fos activity pattern. Our results suggest that pharmacological treatment with a PAM of the mGluR2 such as JNJ could help improve cognitive and negative symptoms related to SZ.

Association study of candidate genes with obesity and metabolic traits in antipsychotic-treated patients with first-episode psychosis over a 2-year period.

AIMS: Patients with a first episode of psychosis (FEP) often display different metabolic disturbances even independently of drug therapy. However, antipsychotic (AP) treatment, especially with second-generation APs, is strongly linked to weight gain, which increases patients' risk of developing obesity and other metabolic diseases. There is an important genetic risk component that can contribute to the appearance of these disturbances. The aim of the present study was to evaluate the effect of polymorphisms in selected candidate genes on obesity and other anthropometric and metabolic traits in 320 AP-treated FEP patients over the course of a 2-year follow-up. METHODS: These patients were recruited in the multicentre PEPs study (Phenotype-genotype and environmental interaction; Application of a predictive model in first psychotic episodes). A total of 127 validated single nucleotide polymorphisms (SNPs) in 18 candidate genes were included in the genetic analysis. RESULTS: After Bonferroni correction, SNPs in ADRA2A, FTO, CNR1, DRD2, DRD3, LEPR and BDNF were associated with obesity, abdominal circumference, triglycerides, HDL cholesterol, and/or percentage of glycated haemoglobin. CONCLUSIONS: Although our results should be interpreted as exploratory, they support previous evidence of the impact of these candidate genes on obesity and metabolic status. Further research is required to gain a better knowledge of the genetic variants that can be considered relevant metabolic risk factors. The ability to identify FEP patients at higher risk for these metabolic disturbances would enable clinicians to better select and control their AP treatment.

Altered frequencies of Th17 and Treg cells in children and adolescents with obsessive-compulsive disorder.

OBJECTIVE: Obsessive-compulsive disorder (OCD) is a debilitating neuropsychiatric disorder with an etiopathophysiology that seems to include immune alterations. Previous studies have suggested that variations in the levels of circulating T cell subpopulations may be involved in psychiatric diseases. However, the role of these cells in OCD remains unexplored. Hence, the present study aimed to examine the levels of T helper 1 (Th1), Th2, Th17 and regulatory T (Treg) cells in patients with early-onset OCD and healthy controls. METHODS: The assessment was performed in 99 children and adolescents with OCD and 46 control subjects. The percentages of circulating Th1, Th2, Th17 and Treg cells were evaluated using flow cytometry. RESULTS: OCD patients had significantly higher levels of Th17 cells and lower percentages of Treg cells than healthy controls (p = 0.001 and p = 0.005, respectively). Furthermore, levels of Th17 cells progressively increased with the duration (p = 0.005) and severity of OCD (p = 0.008), whereas the percentages of Treg cells significantly declined with the duration of the disorder (p = 1.8 × 10-5). CONCLUSIONS: These results provide more evidence of the involvement of immune dysregulation, specifically an imbalance in the levels of circulating T helper and regulatory T cells, in the pathophysiology of early-onset OCD.

Author Correction: Improving pharmacogenetic prediction of extrapyramidal symptoms induced by antipsychotics.

One of the funding sources (FEDER-Unión Europea) was not previously acknowledged in this Article. This study was supported by the Spanish Ministry of Health, Instituto de Salud Carlos III (FIS, Fondo de Investigacion Sanitaria PI13/00812, PI16/0122) and FEDER-Unión Europea.

Gene-environment interaction between an endocannabinoid system genetic polymorphism and cannabis use in first episode of psychosis.

Alterations of the endocannabinoid system (ECS) may play an important role in the development of schizophrenia and other psychotic disorders. Cannabis use is one of the environmental factors more repeatedly related to an increase the risk of developing a psychotic episode, while its use modifies the ECS normal function. In the present study we purposed to examine the gene by environment (GxE) interaction between 15 selected single nucleotide polymorphisms (SNPs) related to the ECS and cannabis use in a cohort of 321 patients with a first episode of psychosis (FEP) and 241 matched healthy controls. We found the fatty-acid amide hydrolase (FAAH) rs2295633 SNP genetic polymorphism was associated with a greater risk of presenting a FEP in subjects with relevant cannabis use, but not in subjects without a history of cannabis use. The probability of presenting a FEP was tenfold higher (OR: 10.69) in cannabis users who were homozygote carriers of the T allele of the FAAH rs2295633 SNP, compared to users of cannabis without this genotype. We also found that a higher a proportion of TT carriers of the FAAH rs2295633 SNP with a positive history of cannabis use was treated with high potency antipsychotic. This study has identified a GxE-environment interaction between a genetic polymorphism from the ECS and cannabis use involved in the risk of presenting a FEP. Although this preliminary data should be replicated with independent samples, our results highlight the importance of the pro-psychotic effects of exogenous cannabis use over the ECS in certain subjects.

Human-leukocyte antigen class II genes in early-onset obsessive-compulsive disorder.

Objective: The exact aetiology of obsessive-compulsive disorder (OCD) is unknown, although there is evidence to suggest a gene-environment interaction model. Several lines of evidence support a possible role of the immune system in this model. Methods: The present study explores the allele variability in HLA genes of class II (HLA-DRB1, HLA-DQB1) in a sample of 144 early-onset OCD compared with reference samples of general population in the same geographical area. Results: None of the 39 alleles identified (allele frequency >1%) showed significant differences between OCD and reference populations. Pooling the different alleles that comprised HLA-DR4 (including DRB1*04:01, DRB1*04:04 and DRB1*04:05 alleles) we observed a significantly higher frequency (X21 = 5.53, P = 0.018; OR = 1.64, 95% CI 1.08-2.48) of these alleles in the early-onset OCD sample (10.8%) than in the reference population (6.8%). Conclusions: Taking into account the role of HLA class II genes in the central nervous system, the results presented here support a role of the immune system in the pathophysiological model of OCD.

Genetic Associations of Serotoninergic and GABAergic Genes in an Extended Collection of Early-Onset Obsessive-Compulsive Disorder Trios.

OBJECTIVE: Obsessive-compulsive disorder (OCD) is a debilitating neuropsychiatric disorder whose etiology includes important genetic contributions. In a previous transmission disequilibrium study in which 75 complete trios were included, single-nucleotide polymorphisms (SNPs) in serotoninergic and GABAergic genes were associated with early-onset OCD. Our aim was to assess those findings in an extended collection of early-onset OCD trios. METHODS: A transmission disequilibrium test for SNPs in HTR1B (rs2000292), SLC18A1 (rs6586896), GAD1 (rs3791860), and GAD2 (rs8190748) was performed in a total of 101 early-onset OCD trios, from which 26 trios were newly recruited for the purpose of the present analysis. RESULTS: All the SNPs were overtransmitted from parents to OCD probands (p < 0.012, significant after Bonferroni correction). CONCLUSIONS: These results are consistent with the previous findings and constitute more evidence of the role of genetic factors related to serotoninergic and GABAergic pathways in the pathophysiology of early-onset OCD.

Genetic variability in the serotoninergic system and age of onset in anorexia nervosa and obsessive-compulsive disorder.

The age of onset of some psychiatric disorders may have etiopathogenic and clinical effects and may influence outcome. Following on from previous work by our group where we showed that early onset anorexia nervosa (AN) and obsessive-compulsive disorder (OCD) shared a common genetic background, the aim of the present study is to assess genetic pleiotropy related to the serotonergic system (SLC6A4, 5HTR2A, 5HTR2C, TPH2, SLC18A1), in a common phenotype such as very-early age of onset. One hundred and sixteen adolescents diagnosed with AN and 74 adolescents diagnosed with OCD participated in the present study. We confirmed the existence of a genetic overlap between OCD and AN. Specifically, we described genetic pleiotropy for age at onset across these disorders, associating two SNPs (rs6311, rs4942587) of the HTR2A with the very-early onset phenotype.

Improving pharmacogenetic prediction of extrapyramidal symptoms induced by antipsychotics.

In previous work we developed a pharmacogenetic predictor of antipsychotic (AP) induced extrapyramidal symptoms (EPS) based on four genes involved in mTOR regulation. The main objective is to improve this predictor by increasing its biological plausibility and replication. We re-sequence the four genes using next-generation sequencing. We predict functionality "in silico" of all identified SNPs and test it using gene reporter assays. Using functional SNPs, we develop a new predictor utilizing machine learning algorithms (Discovery Cohort, N = 131) and replicate it in two independent cohorts (Replication Cohort 1, N = 113; Replication Cohort 2, N = 113). After prioritization, four SNPs were used to develop the pharmacogenetic predictor of AP-induced EPS. The model constructed using the Naive Bayes algorithm achieved a 66% of accuracy in the Discovery Cohort, and similar performances in the replication cohorts. The result is an improved pharmacogenetic predictor of AP-induced EPS, which is more robust and generalizable than the original.

Further Support for the Involvement of Genetic Variants Related to the Serotonergic Pathway in the Antidepressant Response in Children and Adolescents After a 12-Month Follow-Up: Impact of the HTR2A rs7997012 Polymorphism.

Objective: Fluoxetine is an effective and well-tolerated pharmacological treatment for children and adolescents with major depressive disorder (MDD). However, a high percentage of patients do not respond. There is a substantial genetic contribution to this variable clinical outcome. Based on previous genetic results of our group and given the lack of pharmacogenetics studies of antidepressant response with a long follow-up period, we evaluated the influence of single nucleotide polymorphisms (SNPs) in genes related to the serotonergic pathway on remission and recovery in children and adolescents diagnosed with MDD after 12 months of initiating fluoxetine treatment. Methods: The assessment was performed in 46 patients. All of them were visited at least once a month during the 12-month follow-up. Psychiatrists interviewed patients and their parents to explore clinical improvement. A total of 75 genotyped SNPs in 10 candidate genes were included in the genetic association analysis with remission and recovery. Bonferroni correction for multiple testing was applied to avoid false positive results. Results: The HTR2A rs7997012 SNP was significantly associated after Bonferroni correction with clinical improvement. Particularly, the homozygotes for the major allele (GG) showed the highest percentage of remitters and the highest score reductions on the Clinical Global Impressions-Severity (CGI-S) scale. Moreover, although the results were on the border of statistical significance, the GG homozygotes also tended to experience fewer readmissions during the follow-up period Conclusions: These results provide more evidence of the involvement of genetic variants related to the serotonergic pathway in the antidepressant response. Studies with larger cohorts are needed to integrate all relevant variants into clinical predictors of antidepressant response.

Impact of NTRK2, DRD2 and ACE polymorphisms on prolactin levels in antipsychotic-treated patients with first-episode psychosis.

BACKGROUND: Hyperprolactinemia is a common side-effect of antipsychotics (APs), which may trigger serious secondary problems and compromise the adherence to treatment which is crucial for prognosis, especially in patients presenting with a first-episode of psychosis (FEP). AIMS: We evaluated, in some cases for the first time, the effect of polymorphisms in multiple candidate genes on serum prolactin (PRL) levels in an AP-treated FEP cohort recruited in the multicenter PEPs study (Phenotype - genotype and environmental interaction; Application of a predictive model in first psychotic episodes). METHODS: PRL concentration was measured in serum from 222 patients. A total of 167 polymorphisms were selected in 23 genes. Genetic association analysis was performed in the whole sample and also in homogenous subgroups of patients treated with APs with a high (N = 101) or low risk (N = 95) of increasing PRL release, which showed significant differences in their PRL levels. RESULTS: After Bonferroni correction, polymorphisms in NTRK2, DRD2 and ACE genes were associated with PRL concentration. CONCLUSION: Our results give more support to the impact of DRD2, but also of other genes related to dopamine availability such as ACE. Moreover, this study provides the first evidence for the involvement of NTRK2, which suggests that pathways other than the ones related to dopamine or serotonin may participate in the AP-related PRL levels.

SiNoPsis: Single Nucleotide Polymorphisms selection and promoter profiling.

MOTIVATION: The selection of a single nucleotide polymorphism (SNP) using bibliographic methods can be a very time-consuming task. Moreover, a SNP selected in this way may not be easily visualized in its genomic context by a standard user hoping to correlate it with other valuable information. Here we propose a web form built on top of Circos that can assist SNP-centered screening, based on their location in the genome and the regulatory modules they can disrupt. Its use may allow researchers to prioritize SNPs in genotyping and disease studies. RESULTS: SiNoPsis is bundled as a web portal. It focuses on the different structures involved in the genomic expression of a gene, especially those found in the core promoter upstream region. These structures include transcription factor binding sites (for promoter and enhancer signals), histones and promoter flanking regions. Additionally, the tool provides eQTL and linkage disequilibrium (LD) properties for a given SNP query, yielding further clues about other indirectly associated SNPs. Possible disruptions of the aforementioned structures affecting gene transcription are reported using multiple resource databases. SiNoPsis has a simple user-friendly interface, which allows single queries by gene symbol, genomic coordinates, Ensembl gene identifiers, RefSeq transcript identifiers and SNPs. It is the only portal providing useful SNP selection based on regulatory modules and LD with functional variants in both textual and graphic modes (by properly defining the arguments and parameters needed to run Circos). AVAILABILITY AND IMPLEMENTATION: SiNoPsis is freely available at https://compgen.bio.ub.edu/SiNoPsis/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Correction: Integrating Genetic, Neuropsychological and Neuroimaging Data to Model Early-Onset Obsessive Compulsive Disorder Severity.

[This corrects the article DOI: 10.1371/journal.pone.0153846.].

Microarray gene-expression study in fibroblast and lymphoblastoid cell lines from antipsychotic-naïve first-episode schizophrenia patients.

Schizophrenia (SZ) is a chronic psychiatric disorder whose onset of symptoms occurs in late adolescence and early adulthood. The etiology is complex and involves important gene-environment interactions. Microarray gene-expression studies on SZ have identified alterations in several biological processes. The heterogeneity in the results can be attributed to the use of different sample types and other important confounding factors including age, illness chronicity and antipsychotic exposure. The aim of the present microarray study was to analyze, for the first time to our knowledge, differences in gene expression profiles in 18 fibroblast (FCLs) and 14 lymphoblastoid cell lines (LCLs) from antipsychotic-naïve first-episode schizophrenia (FES) patients and healthy controls. We used an analytical approach based on protein-protein interaction network construction and functional annotation analysis to identify the biological processes that are altered in SZ. Significant differences in the expression of 32 genes were found when LCLs were assessed. The network and gene set enrichment approach revealed the involvement of similar biological processes in FCLs and LCLs, including apoptosis and related biological terms such as cell cycle, autophagy, cytoskeleton organization and response to stress and stimulus. Metabolism and other processes, including signal transduction, kinase activity and phosphorylation, were also identified. These results were replicated in two independent cohorts using the same analytical approach. This provides more evidence for altered apoptotic processes in antipsychotic-naïve FES patients and other important biological functions such as cytoskeleton organization and metabolism. The convergent results obtained in both peripheral cell models support their usefulness for transcriptome studies on SZ.

Association of regulatory TPH2 polymorphisms with higher reduction in depressive symptoms in children and adolescents treated with fluoxetine.

Genetic variability related to the brain serotonergic system has a significant impact on both the susceptibility to psychiatric disorders, such as major depressive disorder (MDD), and the response to antidepressant drugs, such as fluoxetine. TPH2 is one of the most important serotonergic candidate genes in selective serotonin reuptake inhibitors (SSRIs) pharmacogenetic studies. The aim of the present study was to evaluate the influence of regulatory polymorphisms that are specifically located in human TPH2 transcription factor binding sites (TFBSs), and therefore could be functional by altering gene expression, on clinical improvement in children and adolescents treated with fluoxetine. The selection of SNPs was also based on their linkage disequilibrium with TPH2 rs4570625, a genetic variant with questionable functionality, which was previously associated with clinical response in our pediatric population. A total of 83 children and adolescents were clinically evaluated 12weeks after initiating antidepressant treatment with fluoxetine for the first time. Clinical improvement was assessed by reductions in depressive symptoms measured using the Children's Depression Inventory (CDI) scale. The polymorphisms rs11179002, rs60032326 and rs34517220 were, for the first time in the literature, significantly associated with higher clinical improvement. The strongest association was found for rs34517220. In particular, minor allele homozygotes showed higher score reductions on the CDI scale compared with the major allele carriers. Interestingly, this polymorphism is located in a human TPH2 TFBS for two relevant transcription factors in the serotoninergic neurons, Foxa1 and Foxa2, which together with the high level of significance found for this SNP, could indicate that rs34517220 is in fact the crucial functional genetic variant related to the fluoxetine response. These results provide new evidence for the role of regulatory genetic variants that could modulate human TPH2 expression in the SSRI antidepressant response.