Ontogeny of <scp>CYP</scp>3<scp>A</scp> and <scp>P</scp>‐Glycoprotein in the Liver and the Small Intestine of the Göttingen Minipig: An Immunohistochemical Evaluation

Peer, Els Van; Verbueken, Evy; Saad, Moayad A.; Casteleyn, Christophe; Ginneken, Chris Van; Cruchten, Steven Van

doi:10.1111/bcpt.12173

Cited by 26 publications

(16 citation statements)

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“…Moreover, the expression of hepatic genes for drug metabolism and excretion - phase I and phase II enzymes and drug transporters - reaches adult levels one week after birth (data not shown). This finding is consistent with limited public data on the developmental expression of some minipig and human CYP450 enzymes and drug transporters suggesting that the major switch-on appears to occur in minipig like in humans shortly after birth [ 39 , 40 ]. Compared to most pathways, maturation of the immune system is delayed and occurs between four and eight weeks in spleen and bone-marrow based on human B- and T-cell signatures (Fig.…”

Section: Resultssupporting

confidence: 88%

“…We have shown based on global transcript profiling that the developmental gene expression program is concluded about 4 weeks after birth in virtually all organs. The only study addressing juvenile toxicity in minipigs has investigated expression and activity of CYP3A4 during post-natal development [ 39 ]. Both CYP3A4 protein and enzymatic activity are detectable 28 days after birth which is in agreement with the kinetics in our dataset.…”

Section: Discussionmentioning

confidence: 99%

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Functional analysis and transcriptional output of the Göttingen minipig genome

et al. 2015

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BackgroundIn the past decade the Göttingen minipig has gained increasing recognition as animal model in pharmaceutical and safety research because it recapitulates many aspects of human physiology and metabolism. Genome-based comparison of drug targets together with quantitative tissue expression analysis allows rational prediction of pharmacology and cross-reactivity of human drugs in animal models thereby improving drug attrition which is an important challenge in the process of drug development.ResultsHere we present a new chromosome level based version of the Göttingen minipig genome together with a comparative transcriptional analysis of tissues with pharmaceutical relevance as basis for translational research. We relied on mapping and assembly of WGS (whole-genome-shotgun sequencing) derived reads to the reference genome of the Duroc pig and predict 19,228 human orthologous protein-coding genes. Genome-based prediction of the sequence of human drug targets enables the prediction of drug cross-reactivity based on conservation of binding sites. We further support the finding that the genome of Sus scrofa contains about ten-times less pseudogenized genes compared to other vertebrates. Among the functional human orthologs of these minipig pseudogenes we found HEPN1, a putative tumor suppressor gene. The genomes of Sus scrofa, the Tibetan boar, the African Bushpig, and the Warthog show sequence conservation of all inactivating HEPN1 mutations suggesting disruption before the evolutionary split of these pig species. We identify 133 Sus scrofa specific, conserved long non-coding RNAs (lncRNAs) in the minipig genome and show that these transcripts are highly conserved in the African pigs and the Tibetan boar suggesting functional significance. Using a new minipig specific microarray we show high conservation of gene expression signatures in 13 tissues with biomedical relevance between humans and adult minipigs. We underline this relationship for minipig and human liver where we could demonstrate similar expression levels for most phase I drug-metabolizing enzymes. Higher expression levels and metabolic activities were found for FMO1, AKR/CRs and for phase II drug metabolizing enzymes in minipig as compared to human. The variability of gene expression in equivalent human and minipig tissues is considerably higher in minipig organs, which is important for study design in case a human target belongs to this variable category in the minipig. The first analysis of gene expression in multiple tissues during development from young to adult shows that the majority of transcriptional programs are concluded four weeks after birth. This finding is in line with the advanced state of human postnatal organ development at comparative age categories and further supports the minipig as model for pediatric drug safety studies.ConclusionsGenome based assessment of sequence conservation combined with gene expression data in several tissues improves the translational value of the minipig for human drug development. The genome and gene ...

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Section: Resultssupporting

confidence: 88%

Section: Discussionmentioning

confidence: 99%

Functional analysis and transcriptional output of the Göttingen minipig genome

et al. 2015

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“…From day 7 onwards, a gradual increase in CYP3A abundance was noted with a significantly higher CYP3A expression in the adult animals compared to the younger animals. This post‐natal increase in CYP3A expression in Göttingen minipig livers is in agreement with data from a previous immunohistochemical study performed by our group and with data on CYP3A29 mRNA expression in the liver from Bama miniature pigs, showing very low expression in neonatal animals and increasing expression with age . Additionally, it corresponds to human data for CYP3A4.…”

Section: Discussionsupporting

confidence: 92%

Age-related Differences in CYP3A Abundance and Activity in the Liver of the Göttingen Minipig

Peer

Bock

Boussery

et al. 2015

Basic Clin Pharmacol Toxicol

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In view of paediatric drug development, regulatory authorities often request safety studies in juvenile animals, including minipigs. Unfortunately, knowledge on the ontogeny of the biotransformation processes in animal models remains scarce and impedes a correct interpretation of the toxicity findings. CYP3A4 is one of the most important drug-metabolizing enzymes in human beings and shows important similarities with CYP3A in the minipig. Therefore, the aim of this study was to assess the abundance and activity of CYP3A in liver microsomes from foetal, juvenile (days 1, 3, 7 and 28) and adult male and female G€ ottingen minipigs. CYP3A abundance was studied by an indirect enzyme-linked immunosorbent assay (ELISA), whereas CYP3A activity was assessed by a biotransformation assay with Luciferin-IPA. CYP3A abundance could not be detected until day 3. From day 7 onwards, a gradual increase in expression was noted, leading to the highest abundance in adult animals. CYP3A activity was not detectable in foetuses and 1-day-old animals. The CYP3A activity was detectable, but below the LLOQ in day 3 animals and increased gradually with age to reach the highest level in adults. The CYP3cide and ketoconazole inhibition, and testosterone and midazolam reduction of Luciferin-IPA metabolism in minipig liver microsomes substantiate that Luciferin-IPA is metabolized by CYP3A in minipigs. A positive correlation was found between CYP3A abundance and biotransformation of Luciferin-IPA (Pearson r = 0.863; p < 0.0001). In conclusion, both abundance and activity of CYP3A increased gradually in juvenile minipigs, but remained below the levels observed in adult animals.

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“…However, CYP3A29 mRNA expression has recently been found to be low at the age of 1 day and then significantly increases with age from 40 days, 4 months and 6 months to 2 years in Bama miniature pig liver (Shang et al ., ). This phenomenon in CYP3A expression has also been observed in the liver of Göttingen miniature pig from the ages of 1 day to about 2 years (Van Peer et al ., ). Therefore, impact of age on hepatic CYP450 may be different from different breeds of pigs including miniature pigs.…”

Section: Discussionmentioning

confidence: 97%

Impact of age on hepatic cytochrome P450 of domestic male Camborough‐29 pigs

2014

Vet Pharm & Therapeutics

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Swine is not only an important species in veterinary medicine research but also a popular animal model for human drug discovery. It is valuable to understand the impact of pig age on abundance and activity of porcine hepatic cytochrome P450 (CYP450). Liver microsomes were prepared from Camborough-29 intact male pigs at the age of 1 day and 2 weeks and the castrated male pigs at the age of 5, 10, and 20 weeks. Hepatic CYP450 content in the liver microsomes was measured using a UV/visible spectroscopic method. The activities of CYP450s were evaluated by metabolism of phenacetin, coumarin, tolbutamide, bufuralol, chlorzoxazone, and midazolam. The porcine hepatic CYP450 content increased with age with a plateau between age 2 and 5 weeks. Activities of all CYP450 enzymes increased with age of pigs too. The bufuralol 1'-hydroxylase showed the highest hepatic activities compared with other CYP enzymes at all ages of pigs. The average activities at the age of 20 weeks were about five times higher than those at the age of 5 weeks for most of the CYP enzymes. With compensation of the ratio of liver to body weights, the overall CYP450 metabolism capability of the pigs may be peaked around ages of 10 to 20 weeks. Those findings suggest that metabolism can be significantly different in growing phase of pigs and that the age may be an important factor in porcine medicine evaluation and pig model development.

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Ontogeny of CYP3A and P‐Glycoprotein in the Liver and the Small Intestine of the Göttingen Minipig: An Immunohistochemical Evaluation

Cited by 26 publications

References 38 publications

Functional analysis and transcriptional output of the Göttingen minipig genome

Functional analysis and transcriptional output of the Göttingen minipig genome

Age-related Differences in CYP3A Abundance and Activity in the Liver of the Göttingen Minipig

Impact of age on hepatic cytochrome P450 of domestic male Camborough‐29 pigs

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