Publication: Kronik böbrek yetersiziği olan hastalarda CYP3A4 enzim aktivitesinin epigenetik düzenlenmesi
Abstract
Amaç: Sitokrom P450 (CYP) 3A4, insan karaciğerinde en yaygın bulunan CYP enzimidir ve gen anlatımı bireyler arası büyük farklılık göstermektedir. Genetik faktörler kadar epigenenik faktörler de CYP3A4 gen anlatımını etkiler ve DNA metilasyonunun gen anlatımını değiştirmede önemli rolü olduğu ileri sürülmektedir. Kronik böbrek hastalığının (KBH), karaciğer ve böbreklerde sitokrom P450-aracılı ilaç metabolizması üzerindeki etkileri daha önce bildirilmiştir. Bu çalışmanın amacı, CYP3A4 enzim aktivitesinin düzenlenmesini etkileyen genetik ve epigenetik faktörlerinin araştırılmasıdır.Gereç ve yöntem: Çalışmamızda, kronik böbrek yetersizliği (KBY) olan hastalarda (n=67) ve sağlıklı kontrollerde (n=29), CYP3A4 ve nükleer reseptör PXR genlerinin anlatım düzeyleri RT-PZR ile değerlendirildi. CYP3A4 promotöründe farklı metillenen CpG bölgeleri bisülfit dizileme yöntemi ile analiz edildi. CYP3A4 gen aktivitesi üzerine önemli rolü olduğu bildirilen tek nokta polimorfizmi (TNP) CYP3A4*22’in (rs35599367) gen anlatımı üzerine etkisi TNP Genotipleme Analizi ile belirlendi.Bulgular: CYP3A4 ve PXR genlerinin anlatım düzeyleri KBY olan hastalarda (1.920.22 ve 0.870.07, sırasıyla) kontrol grubuna kıyasla (2.541.08 ve 1.420.25, sırasıyla) düşük bulundu ve bu fark PXR geni için istatistiksel olarak anlamlıdır (p<0.05). CYP3A4 promotör bölgesinin demetilasyon oranının glomerüler filtrasyon hızları ile ilişkili olduğu gösterildi (p<0.05). TNP CYP3A4*22 ve KBH arasında istatistiksel olarak anlamlı bir korelasyon bulunmadı. Sonuç: Belirli hastalık süreçleri dahilinde CYP3A4 geninin düzenlenmesinde genetik ve epigenetik faktörler görev almaktadır ve bu alanda yapılan çalışmalar, ilaç metabolizasyon enzimlerinin gen anlatımlarının düzenlenmesi ile ilgili yeni bakış açıları kazandıracaktır.
Objective: Cytochrome P450 (CYP) 3A4, is the most abundant CYP enzyme in human liver and has a large inter-individual difference. As well as genetic factors, epigenetic factors also affect CYP3A4 gene expression and it has been suggested that DNA methylation is important in altering gene expression. The effects of chronic kidney disease (CKD) on cytochrome P450-mediated drug metabolism in the liver and kidney have been reported previously. The aim of this study is to investigate the genetic and epigenetic factors altering the regulation of CYP3A4 enzyme activity.Material and methods: We evaluated gene expression profiles of CYP3A4 and the nuclear receptor PXR genes in patients with CKD (n=67) and healthy controls (n=29) by RT-PCR analysis. The differentially methylated CpG promoter region of the CYP3A4 gene were analyzed by bisulfite sequencing. The effect of single‐nucleotide polymorphism (SNP) CYP3A4*22 (rs35599367) on expression, which has recently been reported to have a critical role in CYP3A4 gene activity, has been determined by SNP Genotyping Assay. Results: The expression levels of CYP3A4 and PXR genes were low in CKD patients (1.920.22 and 0.870.07, respectively), when compared to healthy controls (2.541.08 and 1.420.25, respectively) and the association was found significant for PXR (p<0.05). The demethylation rates of CYP3A4 promoter were associated with glomerular filtration rates (p<0.05). The correlation between the SNP CYP3A4*22 and CKD was not significant. Conclusion: The genetic and epigenetic changes involved in gene regulation of CYP3A4 could be observed in certain disease processes and further investigations will provide new insights into the regulation of drug-metabolizing enzymes.
Objective: Cytochrome P450 (CYP) 3A4, is the most abundant CYP enzyme in human liver and has a large inter-individual difference. As well as genetic factors, epigenetic factors also affect CYP3A4 gene expression and it has been suggested that DNA methylation is important in altering gene expression. The effects of chronic kidney disease (CKD) on cytochrome P450-mediated drug metabolism in the liver and kidney have been reported previously. The aim of this study is to investigate the genetic and epigenetic factors altering the regulation of CYP3A4 enzyme activity.Material and methods: We evaluated gene expression profiles of CYP3A4 and the nuclear receptor PXR genes in patients with CKD (n=67) and healthy controls (n=29) by RT-PCR analysis. The differentially methylated CpG promoter region of the CYP3A4 gene were analyzed by bisulfite sequencing. The effect of single‐nucleotide polymorphism (SNP) CYP3A4*22 (rs35599367) on expression, which has recently been reported to have a critical role in CYP3A4 gene activity, has been determined by SNP Genotyping Assay. Results: The expression levels of CYP3A4 and PXR genes were low in CKD patients (1.920.22 and 0.870.07, respectively), when compared to healthy controls (2.541.08 and 1.420.25, respectively) and the association was found significant for PXR (p<0.05). The demethylation rates of CYP3A4 promoter were associated with glomerular filtration rates (p<0.05). The correlation between the SNP CYP3A4*22 and CKD was not significant. Conclusion: The genetic and epigenetic changes involved in gene regulation of CYP3A4 could be observed in certain disease processes and further investigations will provide new insights into the regulation of drug-metabolizing enzymes.