The Inhibition Of UGTs By D-thyroxine

Object:Most of the endogenous and exogenous substances in the human body are removed in vitro mainly through the metabolism of?metabolic enzymes and?phase metabolic enzymes.The main metabolic enzymes?phase is uridine diphosphate glucuronic acid transferase(UGTs),to participate in the aldehyde to acidification of glucose in the bodies.UGTs activity is inhibited,which accumulates metabolites in the body and causes corresponding toxic and side effects.Thyroid tablets are commonly used as endocrine therapy after thyroid cancer surgery.Thyroid tablets contain varying proportions of thyroxine.Clinical discovery thyroid gland tablet can affect the action of other medicine.This experiment mainly studies the interaction between D-thyroxine and UGTs to explore whether D-thyroxine is the composition and mechanism of drug-drug interaction produced by thyroid tablets.Method:To simulate the catalytic environment of human metabolic enzymes,the UGTs catalyzed gluconaldehyde acidification incubation system was constructed in vitro.Nine UGTs(UGT1A1?UGT1A3?UGT1A6?UGT1A7?UGT1A8?UGT1A9?UGT1A10?UGT2B7?UGT2B17)of human recombinant liver microsomal were selected as the main enzymes to catalyze the metabolic reaction.We selected4-methyl umbel ketone(4-MU)as a non-selective metabolic substrate and D-thyroxine(D-T4)was selected as the inhibitor.A blank control group was set up in the experiment.At the end of the in vitro incubation system,the product 4-methyl umbrella ketone-d-glucuronide was detected by UPLC.Therefore,we obtained the residual activity of UGT subtype to metabolize 4-methyl umbrella ketone in the presence of D-thyroxine.The inhibitory activity of over 80%of the enzymes was followed by IC50 and K_i.The double reciprocal plot and in vitro and in vivo internal extrapolation method were used to obtain the inhibition types of D-thyroxine on different UGT subtypes and the minimum concentration of D-thyroxine in human body to inhibit UGTs.Finally,the docking software was used to make the docking between D-T4 and the UGT subtype inhibited by D-T4.The site and binding energy of D-T4 on UGT subtype were observed from the perspective of spatial structure,and the mechanism of D-T4's inhibition on UGTs was further discussed.Result:The residual inhibitory activities of D-thyroxine on UGT1A1,UGT1A3,UGT1A6,UGT1A7,UGT1A8,UGT1A9,UGT1A10,UGT2B7,UGT2B17 were1.74%,14.26%,11.66%,17.79%,19.23%,38.73%,1.85%,17.95%,7.41%,respectively.D-thyroxine inhibits UGT1A1,UGT1A3,UGT1A6,UGT1A7,UGT1A8,UGT1A10,UGT2B7 and UGT2B17 more than 80%.Subsequent experiments showed that the inhibitory concentration(IC50)of D-thyroxine on UGT1A1,UGT1A3,UGT1A6,UGT1A7,UGT1A8,UGT1A10,UGT2B7 and UGT2B17 was respectively 1?mol/L-5?mol/L,10?mol/L-20?mol/L,10?mol/L-20?mol/L,10?mol/L-20?mol/L,5?mol/L-10?mol/L,20?mol/L-40?mol/L,10?mol/L-20?mol/L,1?mol/L-5?mol/L,1?mol/L-5?mol/L.The Ki values obtained in subsequent experiments were 61.4 mol/L,12.85 mol/L,14.8 mol/L,28.5 mol/L,8.7mol/L,12.9 mol/L,25.2 mol/L,23.6 mol/L.By the body in vitro extrapolation get that the catalytic activity of UGT1A1 was inhibited when the concentration of D-thyroxine reached 6.14?mol/L,that the catalytic activity of UGT1A3 was inhibited when the concentration of D-thyroxine reached 12.85?mol/L,that the catalytic activity of UGT1A6 was inhibited when the concentration of D-thyroxine reached 14.8?mol/L,that the catalytic activity of UGT1A7 was inhibited when the concentration of D-thyroxine reached 28.5?mol/L,that the catalytic activity of UGT1A8 was inhibited when the concentration of D-thyroxine reached 8.7?mol/L,that the catalytic activity of UGT1A10 was inhibited when the concentration of D-thyroxine reached 12.9?mol/L,that the catalytic activity of UGT2B7 was inhibited when the concentration of D-thyroxine reached 25.2?mol/L,that the catalytic activity of UGT2B17 was inhibited when the concentration of D-thyroxine reached 23.6?mol/L,leading to drug-drug interaction.The Binding energy of D-thyroxine and UGTs was obtained by using the 4.2version autodock docking software.The results showed that the free binding energy of D-thyroxine and UGT2B7 was-8.63 kcal/mol,the free binding energy of D-thyroxine and UGT1A9 was-8.72 kcal/mol,the free binding energy of D-thyroxine and UGT1A8 was-9.83 kcal/mol,the free binding energy of D-thyroxine and UGT1A7 was-9.57 kcal/mol,and the free binding energy of D-thyroxine and UGT1A6 was-9.57 kcal/mol.The free binding energy of D-thyroxine and UGT1A3 was-8.4 kcal/mol,and the free binding energy of D-thyroxine and UGT1A1 was-8.76 kcal/mol.Conclusion:This study indicated that D-thyroxine had a strong inhibitory effect on the catalytic activities of UGT1A1,UGT1A3,UGT1A6,UGT1A7,UGT1A8,UGT1A10,UGT2B7 and UGT2B17.D-thyroxine competitively inhibits the activity of UGT1A1,UGT1A3,UGT1A7,UGT1A10 and UGT2B7,while non-competitively inhibits the activity of UGT1A6,UGT1A8 and UGT2B17.It is suggested that the dosage of drugs containing D-thyroxine should be limited to avoid drug-drug interaction.