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Genes and Environment : the Official... 2017Genetic and environmental risk factors play an important role for the susceptibility to sporadic Parkinson's disease (PD). It was hypothesized that a splice variant of...
BACKGROUND
Genetic and environmental risk factors play an important role for the susceptibility to sporadic Parkinson's disease (PD). It was hypothesized that a splice variant of the gene ( allele) is associated with PD because it alters the ability to metabolize toxins and in particular neurotoxins. codes for the drug metabolizing enzyme debrisoquine 4-hydroxylase. The CYP2D6*4 variant results in an undetectable enzyme activity and consequently in a reduction in metabolism of some toxins.
METHODS
Some of agricultural chemicals have neurotoxic potential and CYP2D6 is involved in their detoxification. Thus, we conducted a case control study to investigate the association of the CYP2D6*4 with PD in a Pakistani subpopulation that is known to be exposed to high levels of some agricultural pesticides, insecticides and herbicides.
RESULTS
We found a significantly higher allele and genotype frequency of the variant in 174 sporadic PD patients when compared to 200 controls. In addition, there was a trend to an earlier age of PD onset and a tremor dominant phenotype in variant carriers.
CONCLUSION
Our data provide further evidence that a poor metabolizer status may increase the risk to develop PD especially in populations that are exposed to environmental toxins.
PubMed: 28680508
DOI: 10.1186/s41021-017-0078-8 -
Stem Cells Translational Medicine Jan 2017Liver disease affects large numbers of patients, yet there are limited treatments available to replace absent or ineffective cellular function of this crucial organ....
Liver disease affects large numbers of patients, yet there are limited treatments available to replace absent or ineffective cellular function of this crucial organ. Donor scarcity and the necessity for immunosuppression limit one effective therapy, orthotopic liver transplantation. But in some conditions such as inborn errors of metabolism or transient states of liver insufficiency, patients may be salvaged by providing partial quantities of functional liver tissue. After transplanting multicellular liver organoid units composed of a heterogeneous cellular population that includes adult stem and progenitor cells, both mouse and human tissue-engineered liver (TELi) form in vivo. TELi contains normal liver components such as hepatocytes with albumin expression, CK19-expressing bile ducts and vascular structures with α-smooth muscle actin expression, desmin-expressing stellate cells, and CD31-expressing endothelial cells. At 4 weeks, TELi contains proliferating albumin-expressing cells and identification of β2-microglobulin-expressing cells demonstrates that the majority of human TELi is composed of transplanted human cells. Human albumin is detected in the host mouse serum, indicating in vivo secretory function. Liquid chromatography/mass spectrometric analysis of mouse serum after debrisoquine administration is followed by a significant increase in the level of the human metabolite, 4-OH-debrisoquine, which supports the metabolic and xenobiotic capability of human TELi in vivo. Implanted TELi grew in a mouse model of inducible liver failure. Stem Cells Translational Medicine 2017;6:238-248.
Topics: Adult Stem Cells; Animals; Arginase; Cell Proliferation; Disease Models, Animal; Female; Hepatocytes; Humans; Liver; Mice, SCID; Organoids; Tissue Engineering
PubMed: 28170183
DOI: 10.5966/sctm.2016-0205 -
American Journal of Veterinary Research Sep 2016OBJECTIVE To characterize polymorphisms of the gene for cytochrome P450 isozyme 2D50 (CYP2D50) and the disposition of 2 CYP2D50 probe drugs, dextromethorphan and...
OBJECTIVE To characterize polymorphisms of the gene for cytochrome P450 isozyme 2D50 (CYP2D50) and the disposition of 2 CYP2D50 probe drugs, dextromethorphan and debrisoquine, in horses. ANIMALS 23 healthy horses (22 Thoroughbreds and 1 Standardbred). PROCEDURES Single-nucleotide polymorphisms (SNPs) in CYP2D50 were identified. Disposition of dextromethorphan (2 mg/kg) and debrisoquine (0.2 mg/kg) were determined after oral (dextromethorphan) or nasogastric (debrisoquine) administration to the horses. Metabolic ratios of plasma dextromethorphan and total dextrorphan (dextrorphan plus dextrorphan-O-β-glucuronide) and 4-hydroxydebrisoquine concentrations were calculated on the basis of the area under the plasma concentration-versus-time curve extrapolated to infinity for the parent drug divided by that for the corresponding metabolite. Pharmacokinetic data were used to categorize horses into the phenotypic drug-metabolism categories poor, extensive, and ultrarapid. Disposition patterns were compared among categories, and relationships between SNPs and metabolism categories were explored. RESULTS Gene sequencing identified 51 SNPs, including 27 nonsynonymous SNPs. Debrisoquine was minimally detected after oral administration. Disposition of dextromethorphan varied markedly among horses. Metabolic ratios for dextromethorphan ranged from 0.03 to 0.46 (mean, 0.12). On the basis of these data, 1 horse was characterized as a poor metabolizer, 18 were characterized as extensive metabolizers, and 3 were characterized as ultrarapid metabolizers. CONCLUSIONS AND CLINICAL RELEVANCE Findings suggested that CYP2D50 is polymorphic and that the disposition of the probe drug varies markedly in horses. The polymorphisms may be related to rates of drug metabolism. Additional research involving more horses of various breeds is needed to fully explore the functional implication of polymorphisms in CYP2D50.
Topics: Animals; Cytochrome P-450 Enzyme System; Debrisoquin; Dextromethorphan; Female; Horses; Isoenzymes; Male; Polymorphism, Single Nucleotide
PubMed: 27580115
DOI: 10.2460/ajvr.77.9.1029 -
Biochimica Et Biophysica Acta. General... Jan 2017Cytochrome P450 2U1 (CYP2U1) has been identified from the human genome and is highly conserved in the living kingdom. It is considered as an "orphan" protein as few data...
BACKGROUND
Cytochrome P450 2U1 (CYP2U1) has been identified from the human genome and is highly conserved in the living kingdom. It is considered as an "orphan" protein as few data are available on its physiological function(s) and spectral characteristics. Its only known substrates reported so far are unsaturated fatty acids such as arachidonic acid (AA), and, more recently, N-arachidonoylserotonin (AS) and some xenobiotics related to debrisoquine (Deb) and terfenadine.
METHODS
We have expressed CYP2U1 in E. coli and performed UV-vis and EPR spectroscopy experiments with purified CYP2U1 alone and in the presence of substrates and imidazole and pyridine derivatives. Docking experiments using a 3D homology model of CYP2U1 were done to explain the observed spectroscopic data and the different regioselectivities of the oxidations of AA and AS.
RESULTS
The UV-vis and EPR spectra of native recombinant human CYP2U1 revealed a predominant low-spin hexacoordinate Fe state. Imidazole (Im) derivatives, such as miconazole, acted as Fe ligands, contrary to ketoconazole, whereas the previously described substrates AS and Deb led to "reverse type I" difference UV-vis spectra. These data, as well as the different regioselectivities of AA and AS oxidations, were supported by docking experiments performed on our previously reported CYP2U1 3D model.
MAJOR CONCLUSION AND GENERAL SIGNIFICANCE
Our study describes for the first time the mode of interaction of several Fe-heme ligands and substrates with the active site of CYP2U1 on the basis of spectroscopic and molecular docking data. The good agreement between these data validates the used CYP2U1 3D model which should help the design of new substrates or inhibitors of this orphan CYP.
Topics: Arachidonic Acid; Arachidonic Acids; Biocatalysis; Cytochrome P450 Family 2; Debrisoquin; Electron Spin Resonance Spectroscopy; Escherichia coli; Humans; Imidazoles; Lauric Acids; Ligands; Models, Molecular; Molecular Docking Simulation; Oxidation-Reduction; Protein Binding; Pyridines; Recombinant Proteins; Serotonin; Spectrophotometry, Ultraviolet; Substrate Specificity
PubMed: 27456766
DOI: 10.1016/j.bbagen.2016.07.018 -
Bulletin of Experimental Biology and... Mar 2016Polymorphic variants of CYP1A2 and CYP2D6 genes of the cytochrome P450 system were studied in patients with schizophrenia with drug-induced motor disorders and...
Polymorphic variants of CYP1A2 and CYP2D6 genes of the cytochrome P450 system were studied in patients with schizophrenia with drug-induced motor disorders and hyperprolactinemia against the background of long-term neuroleptic therapy. We revealed an association of polymorphic variant C-163A CYP1A2*1F of CYP1A2 gene with tardive dyskinesia and association of polymorphic variant 1846G>A CY2D6*4 and genotype A/A of CYP2D6 gene (responsible for debrisoquin-4-hydroxylase synthesis) with limbotruncal tardive dyskinesia in patients with schizophrenia receiving neuroleptics for a long time.
Topics: Antipsychotic Agents; Cytochrome P-450 CYP1A2; Cytochrome P-450 CYP2D6; Drug-Related Side Effects and Adverse Reactions; Female; Genetic Predisposition to Disease; Humans; Hyperprolactinemia; Male; Motor Disorders; Polymorphism, Single Nucleotide; Schizophrenia; Tardive Dyskinesia
PubMed: 27021090
DOI: 10.1007/s10517-016-3250-4 -
Current Drug Metabolism 2016Non-human primates are valuable animal models in drug discovery and biomedical research. Human CYP2D6 accounts for 1.3-4.3% of total hepatic CYP content in the liver,... (Review)
Review
BACKGROUND
Non-human primates are valuable animal models in drug discovery and biomedical research. Human CYP2D6 accounts for 1.3-4.3% of total hepatic CYP content in the liver, but is involved in the metabolism of more than 150 drugs. With the advancement of genomic sequencing and annotation, a panel of CYP2D genes have been cloned from non-human primates. This review highlights the similarities and differences of these CYP2D genes non-human primates.
METHODS
We conducted a structured PubMed search using a focused review question and proper inclusion/exclusion criteria. The quality of retrieved papers was assessed and briefed using standard tools and expert knowledge.
RESULTS
Most studies on CYP expression in non-human primates have been carried out in the cynomolgus and Rhesus monkeys. Deduced amino acid sequences of primate CYP2D cDNAs share high sequence identity (93-96%) with human CYP2D6. The chimpanzee genome has CYP2D6 and 2D7 but bonobos only contain CYP2D6. The CYP2D6 gene is located on chromosome 22 in the chimpanzee genome (human CYP2D6 maps to chromosome 22q13.1), and on chromosome 10 in the genome of the Rhesus monkey. Cynomolgus monkey CYP2D17 and Japanese monkey 2D29 metabolize bufuralol and dextromethorphan. CYP2D17 metabolizes bufuralol and dextromethorphan, whereas CYP2D29 metabolizes bufuralol and debrisoquine. In addition, quinidine inhibits both cynomolgus monkey CYP2D17 and Japanese monkey 2D29.
CONCLUSION
The CYP2D members from non-human primates show differential genomic contexts, catalytic activities toward substrates and inhibitory profiles. Further studies are warranted to elucidate the structural and functional features of CYP2D members in non-human primates and thus offer a solid base for the application of these animals in drug discovery.
Topics: Animals; Cytochrome P450 Family 2; Drug Discovery; Isoenzymes; Liver; Models, Animal; Primates; Protein Conformation; Species Specificity; Structure-Activity Relationship; Substrate Specificity; Xenobiotics
PubMed: 26892731
DOI: 10.2174/1389200217666160219114241 -
Pharmacogenomics 2016This study was aimed to functionally characterize four novel CYP2D6 alleles identified in Chinese Han population.
AIM
This study was aimed to functionally characterize four novel CYP2D6 alleles identified in Chinese Han population.
MATERIALS & METHODS
CYP2D6 proteins of wild-type and the four novel variants along with CYP2D6.2 and CYP2D6.10 were heterologously expressed in yeast cells and the kinetic parameters were determined.
RESULTS
Compared with CYP2D6.1 (frequency in Chinese 24.65%), CYP2D6.X (1.63%), CYP2D6.Y (1.50%), CYP2D6.Z (0.81%), CYP2D6.10 (52.53%) and CYP2D6.75 (0.13%) exhibited low activity at different degrees, whereas the kinetic parameters of CYP2D6.2 (11.06%) were much the same with CYP2D6.1. The novel allele CYP2D6.75 showed decreased enzyme activity.
CONCLUSION
This is the first study to conduct functional analysis of CYP2D6 four novel alleles in Chinese Han population, which might be helpful for optimizing pharmacotherapy and the design of personalized medicine.
Topics: Alleles; Asian People; Cytochrome P-450 CYP2D6; Debrisoquin; Genetic Variation; Humans; Recombinant Proteins; Saccharomyces cerevisiae
PubMed: 26652007
DOI: 10.2217/pgs.15.148 -
Pharmacological Research Nov 2015We tested the influence of four polymorphisms and gene duplication in CYP2D6 on in vivo enzyme activity in a Chilean mestizo population in order to identify the most...
We tested the influence of four polymorphisms and gene duplication in CYP2D6 on in vivo enzyme activity in a Chilean mestizo population in order to identify the most relevant genetic profiles that account for observed phenotypes in this ethnic group. CYP2D6*2 (2850C>T), *3 (2549A>del), *4 (1846G>A), *17 (1023C>T) and gene duplication were determined by PCR-RFLP or PCRL in a group of 321 healthy volunteers. Individuals with different variant alleles were phenotyped by determining debrisoquine 4-hydroxylase activity as a metabolic ratio (MR) using a validated HPLC assay. Minor allele frequencies were 0.41, 0.01, 0.12 and 0.00 for CYP2D6*2, *3, *4 and *17 variants, respectively, and the duplication frequency was 0.003. Genotype analysis correlated with phenotypes in 18 of 23 subjects (78.3%). 11 subjects were extensive metabolizers (EM), 8 were intermediate metabolizers (IM), 2 were poor metabolizers (PM) and 2 were ultra-rapid metabolizers (UM) which is fairly coincident with expected phenotypes metabolic ratios ranged from 0.11 to 126.41. The influence of CYP2D6*3 was particularly notable, although only heterozygote carriers were present in our population. Individuals homozygous for *4 were always PM. As expected, the only subject with gene duplication was UM. In conclusion, there was a clear effect of genotype on observed CYP2D6 activity. Classification of EM, PM and UM through genotyping was useful to characterize CYP2D6 phenotype in the Chilean mestizo population.
Topics: Adolescent; Adult; Chile; Cytochrome P-450 CYP2D6; Debrisoquin; Female; Gene Duplication; Gene Frequency; Genetic Association Studies; Humans; Indians, South American; Kinetics; Male; Polymorphism, Genetic; White People; Young Adult
PubMed: 26211952
DOI: 10.1016/j.phrs.2015.07.020 -
Stem Cell Research & Therapy Mar 2015Chimeric mice with humanized livers were recently established by transplanting human hepatocytes. This mouse model that is repopulated with functional human hepatocytes...
INTRODUCTION
Chimeric mice with humanized livers were recently established by transplanting human hepatocytes. This mouse model that is repopulated with functional human hepatocytes could be a useful tool for investigating human hepatic cell biology and drug metabolism and for other preclinical applications. Successfully transplanting human hepatocytes into mice requires that recipient mice with liver failure do not reject these human cells and provide a suitable microenvironment (supportive niche) to promote human donor cell expansion and differentiation. To overcome the limitations of current mouse models, we used Alb-TRECK/SCID mice for in vivo human immature hepatocyte differentiation and humanized liver generation.
METHODS
1.5 μg/kg diphtheria toxin was administrated into 8-week-old Alb-TRECK/SCID mice, and the degree of liver damage was assessed by serum aspartate aminotransferase activity levels. Forty-eight hours later, mice livers were sampled for histological analyses, and the human donor cells were then transplanted into mice livers on the same day. Chimeric rate and survival rate after cell transplantation was evaluated. Expressions of human hepatic-related genes were detected. A human albumin enzyme-linked immunosorbent assay was performed after 50 days of transplantation. On day 60 after transplantation, drug metabolism was examined in mice.
RESULTS
Both human primary fetal liver cells and hepatic stem cells were successfully repopulated in the livers of Alb-TRECK/SCID mice that developed lethal fulminant hepatic failure after administering diphtheria toxin; the repopulation rate in some mice was nearly 100%. Compared with human primary fetal liver cells, human hepatic stem cell transplantation rescued Alb-TRECK/SCID mice with lethal fulminant hepatic failure, and human hepatic stem cell-derived humanized livers secreted more human albumin into mouse sera and also functioned as a "human liver" that could metabolize the drugs ketoprofen and debrisoquine.
CONCLUSION
Our model of a humanized liver in Alb-TRECK/SCID mice may provide for functional applications such as drug metabolism, drug to drug interactions, and promote other in vivo and in vitro studies.
Topics: Animals; Cell Differentiation; Cell Proliferation; Cells, Cultured; Chimera; Debrisoquin; Diphtheria Toxin; Disease Models, Animal; Hepatocytes; Humans; Inactivation, Metabolic; Ketoprofen; Liver; Liver Failure, Acute; Liver Regeneration; Mice; Mice, Knockout; Mice, SCID; Stem Cell Transplantation; Stem Cells; Transplantation, Heterologous
PubMed: 25889844
DOI: 10.1186/s13287-015-0038-9 -
Biochimica Et Biophysica Acta Jul 2015Cytochrome P450 2U1 (CYP2U1) has been identified from the human genome and is highly conserved in the living kingdom. In humans, it has been found to be predominantly...
Expression in yeast, new substrates, and construction of a first 3D model of human orphan cytochrome P450 2U1: Interpretation of substrate hydroxylation regioselectivity from docking studies.
BACKGROUND
Cytochrome P450 2U1 (CYP2U1) has been identified from the human genome and is highly conserved in the living kingdom. In humans, it has been found to be predominantly expressed in the thymus and in the brain. CYP2U1 is considered as an "orphan" enzyme as few data are available on its physiological function(s) and active site topology. Its only substrates reported so far were unsaturated fatty acids such as arachidonic acid, and, much more recently, N-arachidonoylserotonin.
METHODS
We expressed CYP2U1 in yeast Saccharomyces cerevisiae, built a 3D homology model of CYP2U1, screened a library of compounds known to be substrates of CYP2 family with metabolite detection by high performance liquid chromatography-mass spectrometry, and performed docking experiments to explain the observed regioselectivity of the reactions.
RESULTS
We show that drug-related compounds, debrisoquine and terfenadine derivatives, subtrates of CYP2D6 and CYP2J2, are hydroxylated by recombinant CYP2U1 with regioselectivities different from those reported for CYP2D6 and 2J2. Docking experiments of those compounds and of arachidonic acid allow us to explain the regioselectivity of the hydroxylations on the basis of their interactions with key residues of CYP2U1 active site.
MAJOR CONCLUSION
Our results show for the first time that human orphan CYP2U1 can oxidize several exogenous molecules including drugs, and describe a first CYP2U1 3D model.
GENERAL SIGNIFICANCE
These results could have consequences for the metabolism of drugs particularly in the brain. The described 3D model should be useful to identify other substrates of CYP2U1 and help in understanding its physiologic roles.
Topics: Blotting, Western; Catalytic Domain; Chromatography, High Pressure Liquid; Computer Simulation; Cytochrome P-450 Enzyme System; Cytochrome P450 Family 2; Debrisoquin; Kinetics; Mass Spectrometry; Models, Molecular; Molecular Structure; Oxidation-Reduction; Protein Binding; Protein Structure, Tertiary; Recombinant Proteins; Saccharomyces cerevisiae; Substrate Specificity
PubMed: 25857771
DOI: 10.1016/j.bbagen.2015.03.014