Metabolism Of Drugs And Xenobiotics By Human CYPs

With the development of the pharmaceutical industry and the continuous improvement of human health,the pharmaceutical research and development will not only focus on the drugs innovation any more,and their research focus point move to the drugs pharmacokinetic properties gradually.Nowadays,to judge whether a drug has application prospects,especially market prospects,efficacy and side effects are still the most important factors,but absolutely not the decisive factor.In the drug research and development department,the researchers want to pay more and more attention to test whether the new drugs have good pharmacokinetic properties,otherwise,the drugs will not work successfully in the human body even if they have curative effect,and sometimes it may accumulate toxins in the human body.Just give an example,the typical peptide drugs,many of them are bioactive peptides,such as endorphins.It has good characteristics of high efficiency and low toxicity,but due to instability in the human body,oral administration of such drugs cannot play any effect.Therefore,the study of pharmacokinetics is a very important part in the pharmaceutical research and development process.Pharmacokinetics is the study which mainly investigating the process after taking drug into the human body.And the basic pharmacokinetics processes including absorption,distribution,metabolism and excretion.This article is intended to study the process of drug metabolism,especially the metabolism of exogenous substances.Cytochrome P450 which abbreviation is CYP or P450,it is a group of iso-enzymes which encoded by a set of structurally and functionally related superfamily genes.Cytochrome P450 is mainly distributed in the liver,while some P450 enzymes are also distributed in the small intestine,kidney,lung and brain.Since about 90% drugs are metabolized by cytochrome P450,so most of the research on drug metabolism is focused on the study of cytochrome P450.Cytochrome P450 can metabolize about 250,000 exogenous substances,including drugs,chemicals,environmental pollutants,pesticides,halogenated hydrocarbons,polycyclic aromatic hydrocarbons,aromatic amines,burned ingredients,herbicides and so on.This research topic including two major parts: the metabolization of steroid doping drug;specific substrate screening and inhibition determination of cytochrome 3A family enzymes.First part was the metabolization of steroid doping drug.Anabolic androgenic steroids(AAS)are misused very frequently in sport competitions as performance enhancing agents.One of the doping compounds that has been detected with increased frequency in the last few years is dehydrochloromethyltestosterone(DHCMT,4-chloro-17?-hydroxy-17?-methylandrosta-1,4-dien-3-one;brand name Oral Turinabol).The long-term DHCMT metabolite 20?OH-NorDHCMT(4-chloro-17?-hydroxymethyl-17?-methyl-18-norandrosta-1,4,13-trien-3-one)was reported earlier to be detectable in urine samples for more than 22 days after DHCMT administration;however,purified reference material was not available so far.In this study we demonstrated a successful combination of Wagner-Meerwein rearrangement of DHCMT to NorDHCMT(4-chloro-17,17-dimethyl-18-norandrosta-1,4,13-trien-3-one)and subsequent whole-cell biotransformation with a recombinant fission yeast strain expressing the human cytochrome P450 enzyme CYP21A2 for the synthesis of mg amounts of this metabolite.It was then used as reference for the analysis of a post administration urine of DHCMT.The availability of this reference compound will provide an incontestable proof for DHCMT abuse.The second part was activity and inhibition determination of four iso-enzymes(CYP3A4,CYP3A5,CYP3A7,CYP3A43)in CYP3 A family.This section was intended to provide data of metabolic activity and inhibition for all 57 CYPs.Because cytochrome P450 can metabolize 90% of the drugs on the market,that information will be very helpful when pharmaceutical companies detecting whether a new drug is toxic due to CYP metabolism or screening specific inhibitor for a certain enzyme in a large number of P450 s.“Cocktail” which means mix several P450 s randomly,specific substrates or inhibitors could be screened out rapidly by this method,but has no reports about that so far.Our group intended to construct a library of recombinant fission yeast strains that can express these 57 human P450s;we have a mature method for rapid screening substrates and inhibitors rapidly,also known as “enzyme bags” method;and we have a large number of various luminogenic substrates and common inhibitors,so it is possible to conclude the information of all 57 human P450 s.This part focused on four CYP3 A enzymes with two luminescent substrates(Luciferin H and Luciferin ME)and three common inhibitors(ketoconazole,letrozole and 1-benzylimidazole).But for CYP3A43,which is an “orphan CYP”,CYP3A43 was tested for six luminescent substrates(Luciferin H,Luciferin ME,Luciferin IPA,Luciferin PFBE,Luciferin BE and Luciferin PPXE).The results showed that both CYP3A5 and CYP3A7 could metabolize Luciferin H and Luciferin ME,but CYP3A4 only can metabolize Luciferin H.And CYP3A7 had the highest metabolic activity on Luciferin H,CYP3A5 had higher metabolic activity on Luciferin ME,and CYP3A43 could metabolize Luciferin PPXE.To increase metabolic activity of CYP3A43 to Luciferin PPXE,recombinant strain that can express cytochrome B5 and cytochrome B5 reductase was made,and the metabolic activity of CYP3A43 to Luciferin PPXE can be increased about 46%.In summary,this article synthesized 20?OH-NorDHCMT successfully,a long-term metabolite of Oral Turinabol by a chemical-biological method,which provided a solid evidence for the World Anti-Doping Agency.Also,it enriched the research scope of cytochrome P450 on activity and inhibition,especially discovered a new luminogenic substrate of CYP3A43,which provided new material for CYP3A43 research in the future and laid a certain foundation for using “Cocktail” method in companies.