Inhibitive Effect of Cremophor RH40 or Tween 80-based Self-microemulsiflying Drug Delivery System on Cytochrome P450 3A Enzymes in Murine Hepatocytes

This study examined the effect of self-microemulsiflying drug delivery system (SMEDDS) containing Cremophor RH40 or Tween 80 at various dilutions on cytochrome P450 3A (CYP3A) enzymes in rat hepatocytes, with midazolam serving as a CYP3A substrate.The particle size and zeta potential of microemulsions were evaluated upon dilution with aqueous medium.In vitro release was detected by a dialysis method in reverse.The effects of SMEDDS at different dilutions and surfactants at different concentrations on the metabolism of MDZ were investigated in murine hepatocytes.The cytotoxicity of SMEDDS at different dilutions was measured by LDH release and MTT technique.The effects of SMEDDS on the CYP3A enzymes activity were determined by Western blotting.Our results showed that dilution had less effect on the particle size and zeta potential in the range from 1:25 to 1:500.The MDZ was completely released in 10 h.A significant decrease in the formation of 1’-OH-MDZ in rat hepatocytes was observed after treatment with both SMEDDS at dilutions ranging from 1:50 to 1:250 and Cremophor RH 40 or Tween 80 at concentrations ranging from 0.1% to 1% (w/v), with no cytotoxicity observed.A significant decrease in CYP3A protein expression was observed in cells by Western blotting in the presence of either Cremophor RH40 or Tween 80-based SMEDDS at the dilutions ranging from 1:50 to 1:250.This study suggested that the excipient inhibitor-based formulation is a potential protective platform for decreasing metabolism of sensitive drugs that are CYP3A substrates.

Effect of Intestinal Cytochrome P450 3A on Phytochemical Presystemic Metabolism

Phytochemicals, orally administered substances, are found to undergo presystemic metabolism mainly in the intestine. Although early researches confirmed the role of intestinal bacteria in phytochemical presystemic metabolism, along with the development of molecular biology in investigating intestinal metabolism, a breakthrough has been won in research into metabolizing enzymes and transporters in intestine, which demands more attention and further studies. Recently, Cytochrome P450 3A has been found to be the most effective enzyme in mediating both oxidative （Phase Ⅰ ） and conjugative （Phase Ⅱ ） metabolism in the intestine. The present review summarizes the current findings correlated with the effect of intestinal cytochrome P450 3A on phytochemical presystemic metabolism, which provides a good basis for further research on phytochemical pharmacokinetics.

Cardioprotection of Shenfu preparata on cardiac myocytes through cytochrome P450 2J3

To evaluate whether Shenfu injection （SFI） protects against cardiac myocyte injury induced by Fupian injection （FPI） in vitro. METHODS： H9c2 cells were separately treated with FPI, Renshen injection （RSI） and SFI. Cell viability, lactate dehydrogenase （LDH） release, spontaneous beating rate of primative cardical cells, caspase-3/7 activity, cell apoptosis, and cytochrome P450 2J3 （CYP2J3） mRNA expression were analyzed. RESULTS： The viability of H9c2 cells treated with SFI （37 and 75 mg/mL） was significantly higher than that of H9c2 cells treated with FPI （25 and 50 mg/mL） （P〈0.05, P〈0.01, respectively）. LDH activity of H9c2 cells treated with SFI （75 mg/mL） was significantly decreased （P〈0.01） compared with that of H9c2 cells treated with FPI （50 mg/mL）. SFI （150 mg/mL） significantly attenuated FPI （100 mg/mL）-induced spontaneous beating rate decrease in primary myocardial cells after 4-hour treatment. Compared with FPI （12 and 25 mg/mL）, SFI （18 and 37 mg/mL） treatment could effectively reverse the change of caspase-3/7 activity （P〈0.01 and P〈0.01, respectively）. Compared with FPI （6 and 25 mg/mL）, apoptotic cells decreased significantly （P〈0.05, P〈0.01, respectively） when H9c2 cells were incubated with SFI （9 and 37 mg/mL）. The expression of CYP2J3 mRNA was down-regulated by FPI, while RSI and SFI could up-regulate the expression of CYP2J3 （P〈0.01）, which suggested the potential mechanism of protection of RSI against cardiac myocyte damage induced by FPI treatment. CONCLUSION： These observations indicate that SFI has the potential to exert cardioprotective effects against FPI toxicity. The effect was possibly correlated with the activation of CYP2J3.

Gene Variations of Sixth Complement Component Affecting Tacrolimus Metabolism in Patients with Liver Transplantation for Hepatocellular Carcinoma

Background：Orthotopic liver transplantation （OLT） improves the prognosis of patients with hepatocellular carcinoma （HCC）.Moreover,the complement system is a powerful immune effector that can affect liver function and process of liver cirrhosis.However,studies correlating the complement system with tacrolimus metabolism after OLT are scarce.In this study,the role of single nucleotide polymorphisms （SNPs） associated with the sixth complement component （C6） in tacrolimus metabolism was investigated during the early stages of liver transplantation.Methods：The study enrolled 135 adult patients treated with OLT for HCC between August 2011 and October 2013.Ten SNPs in C6 gene and rs776746 in cytochrome P450 3A5 （CYP3A5） gene were investigated.The tacrolilnus levels were monitored daily during 4 weeks after transplantation.Results：Both donor and recipient CYP3A5 rs776746 allele A were correlated with decreased concentration/dose （C/D） ratios.Recipient C6 rs9200 allele G and donor C6 rs10052999 homozygotes were correlated with lower C/D ratios.Recipient CYP3A5 rs776746 allele A （yielded median tacrolimus C/D ratios of 225.90 at week 1 and 123.61 at week 2）,C6 rs9200 allele G （exhibited median tacrolimus C/D ratios of 211.31 at week l,110.23 at week 2,and 99.88 at week 3）,and donor CYP3A5 rs776746 allele A （exhibited median C/D ratios of 210.82 at week l,111.06 at week 2,77.49 at week 3,and 85.60 at week 4） and C6 rs 10052999 homozygote （exhibited median C/D ratios of 167.59 at week 2,157.99 at week 3,and 155.36 at week 4） were associated with rapid tacrolimus metabolism.With increasing number of these alleles,patients were found to have lower tacrolimus C/D ratios at various time points during the 4 weeks after transplantation.In multiple linear regression analysis,recipient C6 rs9200 group （AA vs.GG/GA） was found to be related to tacrolimus metabolism at weeks 1,2,and 3 （P =0.005,P =0.045,and P =0.033,respectively）,whereas donor C6 rs10052999 group （CC/TT vs.TC） was demonstrated to be correlated with tacrolimus metabolism only at week 4 （P =0.001）.Conclusions：Recipient C6 gene rs9200 polymorphism and donor C6 gene rs10052999 polymorphism are new genetic loci that affect tacrolimus metabolism in patients with HCC after OLT.

Impact of gamma-glutamyl carboxylase gene genovariation in Chinese Han population on the response of warfarin initial anticoagulant therapy

Background In recent years, it is found that the polymorphisms of genes which are involved in the pharmacokinetic and pharmacodynamic pathways play important roles in the clinical anticoagulation treatment with warfarin. The aim of the study was to investigate the impact of the genetic polymorphism of r-glutamic acid carboxylase （gamma-glutamyl carboxylase gene, GGCX） on the response of warfarin initial anticoagulant therapy. Methods Seven hundred and ninety-eight Chinese Han patients who received valve replacement surgery and orally taken warfarin in long term for anticoagulant therapy in Guangdong General Hospital from 2000 to 2008 were enrolled in the study, a polymorphic SNPs point （rs699664） of GGCX was selected, and SnaPshot was adopted to perform single nucleotide polymorphism （SNP） test, and by grouping according to genotype, GGCX average daily dose of warfarin, time for PT-INR to reach the target value and differences in the incidence of excessive coagulation between different genotypes were compared respectively. Hardy-Weinberg genetic equilibrium test was applied for population representative test. Results Within the 20 days of warfarin initial therapy, male average daily dose of warfarin （2.92 ± 1.18 mg/d） was apparently higher than that of female （2.64 ± 0.98 mg/d）, while there were no significant differences in the average time required for PT-INR to reach the target value （ 1.8） and the excessive coagulation ratio at the initial therapy stage between male and female. And there were no significant differences in the average daily dose of warfarin, time to reach the target value and excessive coagulation ratio among different GGCX genotypes. Conclusions GGCX genovariation had no significant impact on the warfarin daily dose within the 20-day initial therapy of Chinese Han Population, and for the conventional dosage program, the risk of bleeding in the GGCX mutation individuals did not increase obviously at the initial administration period.