Effect Of Intestinal Flora On The Pharmacokinetic Of Oral Anticoagulants And Its Mechanism Based On "Transport-metabolism Overlap"

Warfarin as the most widely used oral anticoagulant in clinical practice,the related research is very thorough,but the known influencing factors can only explain the individualized difference of 40～60% of warfarin.Related studies have shown that intestinal flora can affect host metabolism,so intestinal flora may affect warfarin in vivo disposal process.In this paper,the relationship between warfarin and intestinal flora was investigated through clinical study,and animal experiments were designed to study its pharmacokinetic effects and mechanism.Because the new oral anticoagulants are used more frequently in clinic,dabigatran and rivaroxaban are also included in the study.ObjectiveThe effect of intestinal flora on the in vivo disposition of oral anticoagulants and its mechanism were studied,based on "transport metabolism overlap”.Methods1、Clinical study: Design inclusion exclusion criteria included patients,collected information and fecal samples,followed up patients,and according to the various indicators of the anticoagulant treatment of patients were grouped,using 16 S expander sequencing technology to analyze the difference of intestinal flora between different groups of patients.2、Animal experiment:(1)The rat model of intestinal flora disorder was established by antibiotic exposure.(2)The effects antibiotic exposure on intestinal flora were analyzed by 16 amplicon sequencing.(3)Different groups of rats were given warfarin,dabigatran and rivaroxaban by gavage.Blood collection points were designed.Drug concentration was detected by HPLC and GC-MS.The concentration time curve was established,and pharmacokinetic parameters were analyzed.(4)Western Blot and RT-PCR techniques were used to detect the expression of drug transporter P-gp,hepatic CYP1A2、CYP2C9、CYP3A4 and nuclear receptor PXR in rats with intestinal dysbacteriosis.Results1 、 Clinical study: A total of 41 patients were collected fecal samples and information.After follow-up,11 patients(including missing patients and those who met the exclusion criteria)were excluded.The remaining 30 patients were included in the formal trial,including 15 patients in the stable group and 15 patients in the unstable group.Intestinal flora analysis results show that there are significant differences between the two groups of community ratio and dominant flora,in addition,warfarin anticoagulant efficacy unstable patients with less functional enrichment than the stable patients.2、Animal experiment:(1)Amoxicillin was used for 3 days and 7 days to successfully construct the model rats with intestinal flora imbalance.(2)The abundance and diversity of intestinal flora decreased after antibiotic modeling,and the community composition and the function of flora were significantly changed.(3)The bioavailability of warfarin in rats treated with antibiotics for 7 days increased significantly(115% AUC compared with control group and 74% Cmax peak concentration).The bioavailability of warfarin in rats treated with antibiotics for 3days also increased(17% AUC compared with control group and 6% Cmax).The bioavailability of Darbiga group increased significantly(53% higher than that of control group AUC and 130% higher peak concentration Cmax)in rats with 3 days of antibiotic treatment.The bioavailability of rivaroxaban in rats treated with antibiotics for 3 days and 7 days Cmax increased by 10% compared with the control group,the peak concentration Cmax increased by 21%,the AUC of rats in7 days increased by 5% and the peak concentration Cmax increased by 59%.(4)The expression of intestinal P-gp and liver hepatic enzymes decreased compared with CYP1A2 、 CYP2C9 、 CYP3A4 control group after 3 days of antibiotic treatment.the expression of intestinal P-gp was significantly increased compared with the control group after 7 days of treatment,while the expression CYP1A2、CYP2C9、CYP3A4 liver enzymes was significantly decreased.ConclusionChanges in the community composition and abundance of intestinal flora can affect the expression of liver enzyme and drug transporter P-gp,which in turn affects the transport and metabolism of oral anticoagulants(warfarin,dabigatran,rivaroxaban),leading to differences in oral anticoagulants in vivo disposal and changes in bioavailability,thus affecting the efficacy of oral anticoagulants and even inducing adverse reactions.