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    Home > Biochemistry News > Microbiology News > Cell . . . Personalized map of intestinal flora metabolites.

    Cell . . . Personalized map of intestinal flora metabolites.

    • Last Update: 2020-07-27
    • Source: Internet
    • Author: User
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    Oral administration is one of the most common routes of administration. Through this route of administration, drugs will be absorbed in the intestine and then enter the systemic circulation.however, in recent years, more and more studies have found that microorganisms in the gut interact directly or indirectly with drugs, including microbial derived enzymes that convert drugs into other active metabolites, MDM), microbial metabolites compete with drugs for the same host enzyme, and inactive metabolites secreted by drugs are reactivated by microorganisms [1-6].although people have realized the important impact of MDM on pharmacokinetics, due to the complexity of microbiome and the huge technical challenges of testing hundreds of drugs and thousands of metabolites under various conditions, systematic research on MDM is lacking.in addition, due to the obvious differences in gene expression between the strains cultured alone and co cultured with other microorganisms, it is necessary to develop an in vitro mixed culture system which can better reflect the status of the in vivo flora and is suitable for MDM research.on June 10, 2020, Mohamed s. Donia team of Preston University published a research paper entitled personalized mapping of drug metabolism by the human gut microbiology in cell magazine, which provided a solution to this problem.this study provides us with a quantitative experimental protocol, MDM screen, to evaluate the ability of intestinal flora to metabolize oral drug delivery.the team first studied the intestinal flora of a volunteer (PD) to establish a preliminary experimental protocol.the researchers cultured fecal samples of PD in vitro using 14 different media, and compared the culture results with fecal samples by 16S rRNA sequencing technology. The results showed that the sequencing results of mgam medium and fecal samples were the most similar, and the medium had good repeatability.furthermore, based on the mgam medium, the researchers co cultured the microbial samples with 575 oral drugs at 37 ℃, and then identified the metabolites by HPLC-MS, so as to screen and explore the drugs with MDM. At the same time, two control groups were set up: (1) co culture of intestinal flora with DMSO; (2) co culture of sterile mgam with drug.if the culture result of the drug meets the following conditions, it is considered as a positive result (MDM +): (1) new metabolites are obtained from the culture medium or the original drug cannot be identified in the culture medium (2) Consistent results were obtained at least two of the three independent repeated trials. Through the above experimental scheme, the researchers successfully analyzed the metabolism of 438 drugs, 57 of which were MDM +, involving 28 drug categories.among the 57 drugs, 45 were newly discovered MDM +, including 10 drugs that did not exist after culture and 35 drugs that produced new metabolites.figure 1.a-b: the process and results of determining the best culture medium in vitro culture; C-D: the process and results of MDM evaluation. In order to further expand the scope of MDM screening and study the metabolism of drugs by intestinal flora of more individuals, the researchers improved the experimental scheme.in order to determine the best medium for more individuals, the team has developed an indicator, expected number of detectable strips (ends), which considers the probability and biomass of microbial metabolites to be detected.with this indicator, the team obtained a new optimal medium: 70% BB medium + 30% mgam medium (called "BG" medium).at the same time, the team used 96 well plates instead of ordinary tubes for downstream metabolomic analysis and identification of MDM + drugs.through the above method, the team analyzed the samples of 20 volunteers and 23 drugs, and the results showed that the method could effectively identify the MDM differences among individuals.to further evaluate the reliability and practicability of this method, the team further explored microbial derived enzymes involved in drug metabolism. the researchers took capecitabine, a chemotherapeutic drug for the treatment of breast cancer, colorectal cancer and gastric cancer as an example. in the MDM screen study, the researchers found that capecitabine was deglycosylated. previous studies have reported that thymidine phosphorylase (TP) and uridine phosphorylase (up) are involved in this process, but whether there is a similar role in microbiome has not been studied. therefore, based on the homology, the researchers identified the related microbial homologous genes, and constructed TP (Δ deoa), up (Δ UDP) and double knockout E.coli bw25113 strains. the results showed that wild-type E.coli could effectively de glycosylate capecitabine, while the effect of Δ UDP and Δ deoa / Δ UDP strains was significantly decreased. similar results were found in doxifluridine and trifluridine, which are oral fluorouracils (FP) with capecitabine. the above results provide a new explanation for the different efficacy of FP in different patients. in addition, the researchers further verified that the deglycosylation of capecitabine can also occur in vivo by using mouse models. Figure 2. Identification of microbial derived enzymes based on orthogonal strategy. Due to the limitations of the identification method based on homology, the researchers also used the orthogonal strategy to identify microbial derived enzymes. through the construction of different E.coli Expression Vectors for functional screening, the microbial derived enzyme which can transform hydrocortisone into 20b dihydrocortisone was successfully identified. the above results indicate that the combination of MDM screen and functional metagenomic methods is an effective strategy to establish the relationship between MDM screen results and metabolic enzymes from different bacteria without bacterial isolation. to sum up, this study provides us with a quantitative experimental scheme that can effectively explore the metabolic drugs of intestinal flora, and provides a new method for the study of drug microbiology. however, there are still some limitations in this study: firstly, 24% of the tested drugs could not be analyzed in the initial 575 drugs; secondly, the study only focused on oral drugs, and some non intestinal drugs may also have MDM; in addition, the study still failed to restore the intestinal flora 100%, that is, to support the total microbial growth in the original sample. 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Commensal Bifidobacterium promotes antitumor immunity and facilitates anti-PD-L1 efficacy. Science 350, 1084–1089.4. Ve ́tizou, M., Pitt, J.M., Daille` re, R., Lepage, P., Waldschmitt, N., Flament, C., Rusakiewicz, S., Routy, B., Roberti, M.P., Duong, C.P., et al. (2015). Anticancer immunotherapy by CTLA-4 blockade relies on the gut microbiota. Science 350, 1079–1084.5. Wallace, B.D., Wang, H., Lane, K.T., Scott, J.E., Orans, J., Koo, J.S., Venka- tesh, M., Jobin, C., Yeh, L.A., Mani, S., and Redinbo, M.R. (2010). Alleviating cancer drug toxicity by inhibiting a bacterial enzyme. Science 330, 831–835.6. Meinl, W., Sczesny, S., Brigelius-Flohe ́, R., Blaut, M., and Glatt, H. (2009). Impact of gut microbiota on intestinal and hepatic levels of phase 2 xenobi- otic-metabolizing enzymes in the rat. Drug Metab. Dispos. 37, 1179–1186.
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