Creative Animodel has spent decades of efforts to develop and optimize UGT assessments, including UGT reaction phenotyping, induction and inhibition studies, to improve investigational drug development and approval. As a global company, we place high value on effectiveness and innovation to deliver comprehensive solutions with the highest levels of quality.
UDP-glucuronosyl transferases (UGTs) are a family of enzymes involved in conjugation reactions, such as glucuronic acid, sulfonates, glutathione, and amino acids conjugation in the Phase II drug metabolism. UGT isoforms expressed in liver (including UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A9, UGT2B7, UGT2B10, UGT2B15, etc.) as well as extrahepatic tissues (including UGT1A7, UGT1A8, UGT1A10, etc.) account for approximately 35% drug metabolism by phase II enzymes.There are three critical issues associated with UGT: UGT reaction phenotyping, UGT induction and UGT inhibition, indicating directions in investigating potential drug-drug interactions and an effective therapeutic range.
Figure 1. Schematic diagram showing primers designed for the specific UGT isoforms in liver and extrahepatic tissues.
UGT Reaction Phenotyping
UGT reaction phenotyping, also called UGT mapping, is a process to determine which enzymes may be responsible for the proper clearance of investigational compound. We provide methods including correlation analysis involving liver microsomes from at least 10 donors and UGT recombinant enzymes analysis to study polymorphisms and determine which isoforms account for metabolizing the test compound. Test compound is incubated with recombinant UGT enzymes at 37°C, and clearance of the test compound with time is monitored by LC-MS/MS. The profile of the compound clearance in the presence of UGT is corrected by comparison with control groups. In addition, the half life and standard error of half life are also analyzed and contained in our result report. UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A9, UGT2B7 and UGT2B15 are critical to test, and more UGT enzymes are available on request.
UGT Induction Studies
Drugs may increase the activity of drug metabolism by enhancing the expression of relevant genes. This process, called induction, may give rise to decreased plasma levels and impaired pharmacological response of co-administrated medication. FDA and EMA demonstrate that the interaction towards UGT should be under attention.
We provide UGT induction assays using UGT activity endpoint. For activity endpoint, the incubation of specific UGT enzyme with substrates is monitored by LC-MS/MS to determine the probe metabolite levels. A comprehensive portfolio of UGT induction assays are provided upon your request.
Table 1. The substrates and inducers involved in our UGT induction studies.
|UGT1A1||β-Estradiol||β-glucuronidation||Omeprazole (50 μM)|
|UGT1A9||Mefenamic acid||glucuronidation||Omeprazole (50 μM)|
|UGT2B7||Naloxone||3-glucuronidation||Phenobarbital (1 mM)|
UGT Inhibition Studies
Inhibition studies are performed to investigate potential drug-drug interactions and identify the ability of investigational article to inhibit the clearance of other compounds, since drug-drug interactions resulting from inhibition of UGT have been reported. The UGT isoforms recommended by FDA to be studied are UGT1A1, 1A3, 1A4, 1A6, 1A9, 2B7, and 2B15. Other UGT enzymes inhibition assays are also included on request.
Our inhibition studies are typically to incubate the known UGT substrate with recombinant UGT enzymes, alamethicin, UDPGA and a range of investigational compound concentrations at 37°C. The formation of the UGT-specific metabolites is analyzed by LC-MS/MS, delivering IC50and Kivalues towards specific UGT. Our comprehensive reports also include IC50, standard error of IC50, and Ki, all providing information on the potency of the inhibition and any potential in vivo interactions.
Figure 2. The positive control experiment of UGT1A1 and UGT1A3 inhibition studies. Data are shown as mean ± standard deviation.
Creative Animodel provides a portfolio of UGT assessments to predict potential drug interactions and study metabolic characteristics at an early stage. The FDA approved methods are performed by a versatile and experienced expert team to guarantee that the data we deliver are consistently high-quality and cost-efficient.
1. Tukey, R.H.; Strassburg, C.P. Genetic multiplicity of the human UDP-glucuronosyl transferases and regulation in the gastrointestinal tract. Molecular Pharmacology. 2001, 59(3): 405-414.