Effect Of Marine Compound Xyloketal B On The Cytochromes CYP3A2in Rats

Objective:Through computer-aided molecular docking method, in vitro liver microsomal incubationmethod and pharmacokinetic studies, aim to evaluating the ability of Xyloketal B to inhibit theP450enzyme CYP3A,predicting drug-drug interaction and providing experimental evidencefor clinical research.Methods:1. Using computer-aided molecular docking and the scoring function to judge space bindingcapacity between Xyloketal B and major CYP450isoforms.2. Midazolam as an "probe" substrates, liver microsomes in vitro incubation method was usedto investigate the potency and mechanism of Xyloketal B on rat hepatic CYP3A and and itsmechanism.3. Using midazolam as a probe drug,to study Xyloketal B on the pharmacokinetics ofmidazolam and its major metabolite1’-hydroxy midazolam in rats,meanwhile analyze theimpact Xyloketal B on on rat hepatic CYP3A with western blot method and P450-GloTMkitmethod.Results:1. Evaluation Xyloketal B and major CYP450isoforms binding capacity by the scoringfunction.Total-score results were as follows: CYP3A4-3NXU:6.0461; CYP11B2-4DVQ:5.0078;CYP2D6-3TBG:4.9342;CYP2C9-1R90:3.4475;CYP1A2-2HI4:-1.2051;CYP2E1-3T3Z:-8.3081;CYP2A6-2FDV:-16.2292; Binding capacity:CYP3A4>CYP11B2>CYP2D6>CYP2C9>CYP1A2>CYP2E1>CYP2A6. Prompting Xyloketal B with a maximumbinding capacity of CYP3A4.2. The results of enzyme kinetics show that Xyloketal B as rat liver microsomes weak CYP3Ainhibition, IC50was27.91μmol/L, Ki, Ki’, KIand Kinactrespectively41.85,31.67,0.063μmol/L and0.0037min-1. Xyloketal B is a mixed type of non-competitive-anti-competitiveinhibitor and with NADPH, time and concentration dependent manner.This resultsuggested that Xyloketal B may nihibit the activity of CYP3A through mechanism-basedmecheanism. 3. The results showed that compared with normal saline or soybean oil, ketoconazoleincreased both AUC0-tand Cmax, as well as reduced CL/F and AUC0-infratio of1’-OH MDZand MDZ, which were statistically significant. Compared with the physiological saline,14mg/kg Xyloketal B showed significantly elevated midazolam AUC0-tand Cmaxshowing1.68-fold and2.73-fold (p <0.05), respectively. Whereas, decreased CL/F and AUC0-infratio of1’-OH MDZ and MDZ by2.22-fold and1.63-fold, but7mg/kg Xyloketal B had noeffect on midazolam AUC0-t, Cmaxand CL/F. When compared to soybean oil, Xyloketal Bcaused dose-dependent increased of MDZ plasma concentration, with1.41-2.65times (p<0.05or p <0.01) in AUC0-t, and1.36-2.84times in Cmax(p<0.05or p<0.01) and1.35-2.71times reduction in CL/F (p<0.05). No matter compare to negative control group or vehicletreated group, Xyloketal B inhibited CYP3A2protein expression and CYP3A2enzymeactivity with dose-dependent manner.Conclusion:Through computer-aided molecular docking method screening the highest score betweenXyloketal B and CYP450s is CYP3A4,prompting Xyloketal B with a maximum bindingcapacity of CYP3A4. Liver microsomes experiment indicate that Xyloketal B as an mixed typeof non-competitive-anti-competitive inhibitor and with NADPH, time and concentrationdependent. All available PK data demonstrated that changes in the MDZ PK profiles in ratspossibly occur through inhibition of hepatic CYP3A activity by Xyloketal B.Xyloketal Binhibited CYP3A2protein expression and CYP3A2enzyme activity with dose–dependentmanner. Xyloketal B possesses a clinically beneficial property of altering the drug beingmetabolized by CYP3A.