Psychiatric Level 1A evidence pharmacogenomics in a Brazilian admixed cohort and global populations.

Purpose: To compare minor allele frequencies (MAFs) of psychiatric drug response variants in a Brazilian admixed cohort with global populations and other Brazilian groups. Methods: PharmGKB MAFs were gathered from publicly available genetic datasets for Brazil and worldwide. Results: Among 146 variants in CYP2D6 and CYP2C19, 41 were present in Brazil, mostly rare (MAF <1%). 11 variants showed significant MAF differences with large effect sizes compared with global populations. CYP2C19*3 (rs4986893), CYP2C19*17 (rs12248560), CYP2D6*17 (rs28371706-A) and CYP2D6*29 (rs61736512) exhibited higher frequencies in Brazil, with the latter three also differing from other Brazilian groups. Conclusion: This study highlights significant pharmacogenomic diversity in Brazil and globally, underscoring the need for more research in personalized psychiatric drug therapy.

Combined therapy with cisplatin and 5-AZA-2CdR modifies methylation and expression of DNA repair genes in oral squamous cell carcinoma.

The methylation and expression of DNA repair system genes has been studied in several tumor types. These genes have been associated with resistance to chemotherapy treatments by epigenetic regulation. Studies have yet to show the effects of combined therapy using an epigenetic drug (5-aza-2CdR) and cisplatin (CDDP) on DNA repair genes in oral squamous cell carcinoma (OSCC). This study proposed to investigate the effects of CDDP in combination with 5-aza-2CdR on the methylation of MGMT and MLH1 genes in oral cancer cells. Oral squamous cell carcinoma cell lineages (SCC-9, SCC-15, and SCC-25) were submitted to 72 hours of treatment: 0.1 µM CDDP (or 4.44 µM SCC-9), 0.1 µM and 0.3 µM 5-aza-2CdR (or 1 µM and 3 µM SCC-9), and the drugs in combination. Cell viability was assessed by MTT, DNA methylation of MGMT and MLH1 genes by Methylation Sensitivity High-Resolution Melting (MS-HRM), and the relative expression of the genes by RT-qPCR. The results show that all treatments reduced cell viability; however, in SCC-15 and SCC-9 (IC50 value), 5-aza-2CdR promotes cell sensitization to cytotoxic effect of cisplatin. The MGMT promoter region was 100% demethylated in the SCC-15 and SCC-25 cells but partially (50%) methylated in SCC-9 before drug treatment. Treatment with IC50 CDDP value kept the methylation status and decreased MGMT expression in SCC-9; MGMT gene in SCC-15 and SCC-25 cells became downregulated after treatment with 5-aza-2CdR. MLH1 was demethylated, but the treatments with low-doses and combined drugs decreased the expression in SCC-9 and SCC-25; however high doses of 5-aza-2CdR and drug combination with IC50 value CDDP increased expression of MLH1 in SCC-9. The data presented suggest that epigenetic drugs associated with chemotherapy have clinical translational potential as a therapy strategy to avoid or reverse cancer resistance, requiring further investigation.

Perception and knowledge of pharmacogenetics among Brazilian psychiatrists.

Pharmacogenetics (PGx) can optimize drug therapy in psychiatry and is particularly important in admixed populations. Here we developed and successfully validated a questionnaire for assessing the perception and knowledge of PGx among Brazilian psychiatrists. Overall, the participants showed some familiarity with PGx. Most psychiatrists reported to have knowledge of PGx and recognized its usefulness in psychiatry; however, they declared concerns regarding PGx education, the request of tests and their interpretation, cost-effectiveness, and ethical issues. PGx testing is relatively prevalent in their clinical practice, but education on the topic is lacking. Bivariate analysis revealed significant associations. Psychiatrists > 40 years of age more frequently had a positive perception of other clinicians' familiarity with PGx. Psychiatrists in private health services showed less self-reported competency in the use of PGx testing. Furthermore, women had better perception of PGx education. The present study adds knowledge about PGx in psychiatry and encourages the development of educational and training resources for PGx to improve its clinical implementation.

Association of SOCS1 (- 820) (rs33977706) gene polymorphism with chronic periodontitis: A case-control study in Brazilians.

It is evident that the accumulation of periodontal pathogens over the teeth surface triggers periodontitis; however, its aggravation and severity depend on other elements such as environmental factors, systemic health and the host genetic and/or epigenetic background. To address this issue, we investigated the association of two genetic polymorphisms placed on promoter region of SOCS1 gene with chronic periodontal disease. SOCS1 regulates Jak/Kinase signaling pathway and changes in its mRNA expression have been related to different types of cancer and chronic inflammation, including chronic periodontitis. The frequency of alleles and genotypes of two polymorphisms in SOCS1 gene promoter (position - 820 (rs33977706) and position - 1478 (rs33989964)) were analyzed by performing RFLP and TaqMan system in a total of 257 non-smoking subjects. We found a low frequency of A allele and A/A genotype of SOCS1(- 820) polymorphism in the chronic periodontitis group, especially when severe periodontitis samples were separately analyzed (OR = 0.3933; p = 0.0084 (IC95% 0.2112 < µ < 0.7324)), suggesting that A allele plays protective effect against chronic periodontitis. We did not find association between SOCS1-1478 polymorphism and periodontitis. In addition, analysis of SOCS1 (- 820/- 1478) haplotype revealed that the frequency of A(- 820)/CA(- 1478) haplotype decreases in ChrP (p = 0.0089). In conclusion, our study found that SOCS1(- 820) polymorphism is associated with chronic periodontitis.

High-throughput DNA analysis shows the importance of methylation in the control of immune inflammatory gene transcription in chronic periodontitis.

BACKGROUND: Chronic periodontitis represents a complex disease that is hard to control and is not completely understood. Evidence from past studies suggests that there is a key role for DNA methylation in the pathogenesis of periodontitis. However, all reports have applied technologies that investigate genes in a low throughput. In order to advance in the knowledge of the disease, we analyzed DNA methylation variations associated with gene transcription using a high-throughput assay. Infinium® HumanMethylation450 (Illumina) was performed on gingival samples from 12 periodontitis cases and 11 age-matched healthy individuals. Methylation data of 1,284 immune-related genes and 1,038 cell cycle-related genes from Gene Ontology (GO) and 575 genes from a dataset of stably expressed genes (genes with consistent expression in different physiological states and tissues) were extracted from a microarray dataset and analyzed using bioinformatics tools. DNA methylation variations ranging from -2,000 to +2,000 bp from the transcription start site (TSS) were analyzed, and the results were tested against a differential expression microarray dataset between healthy and periodontitis gingival tissues. Differences were evaluated using tests from the R Statistical Project. RESULTS: The comparison of probes between periodontitis and normal gingival tissues showed that the mean methylation scores and the frequency of methylated probes were significantly lower in genes related to the immune process. In the immune group, these parameters were negatively correlated with gene expression (Mann-Whitney test, p < 2.2e - 16). CONCLUSIONS: Our results show that variations in DNA methylation between healthy and periodontitis cases are higher in genes related to the immune-inflammatory process. Thus, DNA methylation must be modulating chromatin regions and, consequently, modulating the mRNA transcription of immune-inflammatory genes related with periodontitis, impacting the prognosis of disease.

Porphyromonas gingivalis LPS stimulation downregulates DNMT1, DNMT3a, and JMJD3 gene expression levels in human HaCaT keratinocytes.

OBJECTIVE: The role of epigenetic regulation in inflammatory diseases such as periodontitis is poorly known. The aim of this study was to assess whether Porphyromonas gingivalis lipopolysaccharide (LPS) can modulate gene expression levels of the some enzymes that promote epigenetic events in cultures of the human keratinocytes and gingival fibroblasts. In addition, the same enzymes were evaluated in gingival samples from healthy and periodontitis-affected individuals. MATERIALS AND METHODS: Primary gingival fibroblast and keratinocyte (HaCaT) cultures were treated with medium containing P. gingivalis LPS or P. gingivalis LPS vehicle for 24 h. After this period, cell viability was assessed by MTT test and total RNA extracted to evaluate gene expression levels of the following enzymes by qRT-PCR: DNA methyltransferase 1 (DNMT1), DNA methyltransferase 3a (DNMT3a), histone demethylases Jumonji domain containing 3 (JMJD3) and ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX). To evaluate gene expression in healthy and periodontitis-affected individuals, total RNA was extracted from biopsies of gingival tissue from healthy and periodontitis sites, and gene expression of DNMT1, DNAMT3a, JMJD3, and UTX was evaluated by qRT-PCR. RESULTS: No significant differences were found in the gene expression analysis between healthy and periodontitis-affected gingival samples. The results showed that LPS downregulated DNMT1 (p < 0.05), DNMT3a (p < 0.05), and JMJD3 (p < 0.01) gene expression in HaCaT cells, but no modulation was observed in gingival fibroblasts. CONCLUSION: P. gingivalis LPS exposure to human HaCaT keratinocytes downregulates gene expression of the enzymes that promote epigenetic events. CLINICAL RELEVANCE: The advance knowledge about epigenetic modifications caused by periodontopathogens may to possibly led to the development of new periodontal therapies.

TLR2 and TLR4 gene promoter methylation status during chronic periodontitis.

AIM: The objective of this study was to analyse the status of DNA methylation in the promoter region of the toll-like receptor (TLR)2 and TLR4 genes in gingival tissue samples from healthy subjects, smokers and non-smokers affected by chronic periodontitis. MATERIAL AND METHODS: Genomic DNA and total RNA were purified from gingival tissue using the TRIZOL reagent protocol. Genomic DNA was then digested by methylation-sensitive restriction enzymes, amplified by polymerase chain reaction (PCR), electrophoresed on a 10% polyacrylamide gel and stained using SYBR Gold. Real-time PCR was also performed to verify the transcript levels. RESULTS: The CpG dinucleotides analysed were observed to be unmethylated in the majority of DNA samples of the three groups and statistical differences were not found among groups (p>0.05). However, a trend towards methylation was observed in the TLR2 HhaI site in the samples of the periodontitis non-smoker groups. In fact, the analysis of all CpG sites together shows which complete methylation is observed in the shortest level in the samples of periodontitis non-smoker group. The analysis of transcript levels demonstrated no difference among groups (p>0.05). CONCLUSION: The results demonstrated major unmethylation of the TLR4 gene promoter in all groups. However, the results for the TLR2 gene promoter are inconclusive; this gene was found as a mosaic of methylated and unmethylated DNA in the majority of samples of the three groups and we also observed a trend towards the DNA methylation of CpG sites recognized by the HhaI enzyme.