Mechanism of long noncoding RNA LINC00987 promoting AML cell apop-tosis through cytochrome P450 pathway / 中国病理生理杂志

AIM:To investigate the role and molecular mechanism of long noncoding RNA LINC00987 in the apoptosis of acute myeloid leukemia(AML)cells induced by antitumor drugs.METHODS:The LINC00987 expression in AML was detected by RT-qPCR.The Molm13 cells with stable knockdown of LINC00987 gene(shLINC00987)were constructed,and the effect of low LINC00987 expression on the apoptosis of AML cells induced by cytarabine was detected by annexin V/PI staining.Signaling pathway enrichment of LINC00987-coexpressed genes was performed to analyze the ef-fect of LINC00987 expression on cytochrome family genes.RESULTS:Compared with healthy individual group,the ex-pression of LINC00987 was significantly down-regulated in AML cell lines and patients,but highly up-regulated in the complete remission group after anti-AML treatment.In addition,low LINC00987 expression was associated with poor prog-nosis among the patients with AML.The LINC00987 expression in AML cell lines Molm13 and MV411 was significantly induced by antitumor drugs such as cytarabine,doxorubicin,arsenic trioxide,and venetoclax.Meanwhile,LINC00987 down-regulation could inhibit the apoptosis of Molm13 cells induced by cytarabine.The LINC00987-coexpressed genes were enriched in cytochrome P450(CYP450)-mediated oxidative stress pathways,and the LINC00987 expression was positively correlated with the expression of CYP450 family genes CYP11B1,CYP2U1 and CYP2C9.Down-regulation of LINC00987 could inhibit the mRNA expression of CYP11B1,CYP2U1 and CYP2C9 induced by cytarabine.CONCLU-SION:Long noncoding RNA LINC00987 can be used as a prognostic marker for AML and may promote cytarabine-in-duced AML cell apoptosis through CYP450-mediated oxidative stress pathways.

Research progress on the role of cytochrome P450 in plant sesquiterpene biosynthesis / 药学学报

Sesquiterpenes are natural terpenoids with 15 carbon atoms in the basic skeleton, which mainly exist in plant volatile oil and have important physiological and medicinal value. Cytochrome P450 (CYP450) is a kind of monooxygenase encoded by supergene family, which is one of the largest gene families in plants. It is involved in the synthesis and metabolism of terpenoids, alkaloids and other secondary metabolites. In the process of terpene biosynthesis, CYP450 participates in the post-modification stage of terpenes by introducing functional groups such as hydroxyl, carboxyl and carbonyl, which plays an important role in enriching the diversity of terpenes. The CYP450 enzymes involved in sesquiterpene synthesis and their substrate catalytic specificity mechanisms have been partially investigated. In this paper, the biosynthetic pathway of plant sesquiterpenes, the structure and classification of CYP450 enzymes were briefly introduced, and the CYP450 enzymes involved in sesquiterpene biosynthesis were summarized, in order to provide a reference for intensive study of the role of CYP450 enzymes in the synthesis of sesquiterpenoids.

Evaluation of the predictive ability of individualized drug administration adjuvant decision-making system JPKD for tacrolimus blood concentration in kidney transplant recipients / 器官移植

Objective To evaluate the predictive ability and influencing factors of individualized drug administration adjuvant decision-making system Java PK® for Desktop (JPKD) for tacrolimus blood concentration in kidney transplant recipients. Methods The monitoring data of tacrolimus blood concentration from 149 recipients early after kidney transplantation were collected. The trough blood concentration of tacrolimus was predicted by JPKD. The absolute weighted deviation and relative prediction deviation between the actual and predicted concentration were calculated. The influencing factors of the absolute weighted deviation were analyzed by univariate and multivariate logistic regression analyses, and the predictive values of these influencing factors on the accuracy of software prediction were assessed by delineating the receiver operating characteristic (ROC) curve. Results Two hundred and sixty-six samples of tacrolimus blood concentration data were collected from 149 patients. The measured blood concentration of tacrolimus was (6.5±3.0) ng/mL (1.1-16.6 ng/mL), and the predicted value calculated by JPKD was (5.6±2.5) ng/mL (1.4-14.4 ng/mL). The absolute weighted deviation of the calculated data was 28.38%, and the relative prediction deviation was −13.55%. Univariate analysis showed that gender, albumin, changes in hematocrit, cytochrome P450 (CYP)3A5*3 genotype and C3435T genotype were associated with the inaccurate prediction results. Multivariate logistic regression analysis found that CYP3A5*3 genotype of AA and the changes in hematocrit were the independent risk factors affecting the accuracy of tacrolimus blood concentration predicted by JPKD. ROC curve analysis showed that when the changes in hematocrit exceeded 2.25%, the risk of inaccurate software prediction was increased. Conclusions JPKD possesses certain accuracy in predicting the blood concentration of tacrolimus in kidney transplant recipients, which may improve the qualified rate of tacrolimus blood concentration. Nevertheless, CYP3A5*3 genotype and the changes of hematocrit may affect the accuracy of predictions.

A rare optineurin mutation in an Indian family with coexistence of JOAG and PCG

Purpose: This study focused on the genetic screening of Myocilin (MYOC), Cytochrome P450 family 1 subfamily B member 1 (CYP1B1), Optineurin (OPTN), and SIX homeobox 6 (SIX6) genes in a family with coexistence of primary congenital glaucoma (PCG) and juvenile open?angle glaucoma (JOAG). Methods: Sanger sequencing was used to examine the coding region of all four genes. Six different online available algorithms were used for the pathogenicity prediction of missense variant. Structural analysis was done using Garnier–Osguthorpe–Robson (GOR), PyMol, ChimeraX, and Molecular Dynamic (MD) Simulations (using Graphics Processing Unit (GPU)?enabled Desmond module of Schrödinger). Results: There were a total of three sequence variants within the family. All seven algorithms determined that a single mutation, G538E, in the OPTN gene is pathogenic. The loops connecting the strands became more flexible, as predicted structurally and functionally by pathogenic mutations. Mutations create perturbations and conformational rearrangements in proteins, hence impairing their functioning. Conclusion: In this study, we describe a North Indian family in which members were having JOAG and PCG due to a rare homozygous/ heterozygous mutation in OPTN. The coexistence of two types of glaucoma within a single pedigree suggests that certain OPTN mutations may be responsible for the onset of different glaucoma phenotypes.

Avaliação in silico e in vivo do perfil de metabolismo e toxicidade hepática de alcaloides ß-carbolínicos sintéticos com prévia atividade antimalárica / In silico and in vivo evaluation of the metabolism profile and hepatic toxicity of synthetic ß-carboline alkaloids with previous antimalarial activity

Introdução: A malária continua sendo um grave problema de saúde pública mundial, dado os elevados índices anuais de morbimortalidade. O tratamento baseia-se no uso de medicamentos, entretanto, a resistência dos parasitos aos medicamentos disponíveis tem se tornado uma realidade alarmante, o que torna urgente o desenvolvimento de novos fármacos com atividade antimalárica. Em um estudo prévio, selecionou-se três alcaloides ß-carbolínicos que apresentaram atividade antimalárica. Dessa forma, o presente trabalho se propôs a dar continuidade ao estudo dessas moléculas avaliando o perfil de metabolismo e toxicidade hepática. Objetivo: Avaliar o perfil de metabolismo e toxicidade hepática de três alcaloides ß-carbolínicos (1, 2 e 3) selecionados em estudo prévio, que apresentaram atividade antimalárica in vitro e in vivo.Material e Métodos: Trata-se de um estudo de abordagem tanto qualitativa quanto quantitativa com caráter experimental e analítico. Foi realizada análise in silico das propriedades de metabolismo e toxicidade dos alcaloides empregando a notação SMILES por meio do programa AdmetSAR 2.0. A toxicidade hepática foi avaliada por meio da análise bioquímica da aspartato aminotrasferase (AST) e alanina aminotransferase (ALT) no soro de camundongos da linhagem C57BL/6, tratados com os alcaloides ou com cloroquina. Resultados: Na análise in silico foi observada a predição de baixo potencial hepatotóxico para os alcaloides 1 e 2, sendo este resultado corroborado pela dosagem de ALT, que apresentou resultados semelhantes ao do grupo controle. O alcaloide 3, no entanto, apresentou dados contrastantes, indicando potencial hepatotóxico na predição in silico, porém, baixo potencial em análise in vivo, com valores de ALT também próximos do grupo controle. Todos os alcaloides em estudo apresentaram potencial para interações medicamentosas. Conclusão: Os alcaloides avaliados nesse estudo apresentaram parâmetros metabólicos e de toxicidade promissores, podendo ser bons adjuvantes à farmacoterapia da malária. Entretanto, esses resultados precisam ser confirmados para seguimento das moléculas nos estudos pré-clínicos.

Introduction: Malaria continues to be a serious global public health problem, given the high annual morbidity and mortality rates. It is caused by protozoa of the genus Plasmodium, with P. falciparum responsible for most serious cases and deaths. Treatment is based on the use of drugs, however, the resistance of parasites to available drugs has become an alarming reality, which makes the development of new drugs with antimalarial activity urgent. In a previous study, our research group selected three ß-carboline alkaloids that showed antimalarial activity. Therefore, the present work proposed to continue the study of these molecules by evaluating the metabolism profile and liver toxicity. Objective: To evaluate the metabolism and liver toxicity profile of three ß-carboline alkaloids (1, 2 and 3) selected in a previous study, which showed antimalarial activity in vitro and in vivo. Material and Methods: This is a study with both a qualitative and quantitative approach with an experimental and analytical nature. In silico analysis of the metabolism and toxicity properties of alkaloids was carried out using the SMILES notation through the AdmetSAR 2.0 program. Liver toxicity was evaluated through biochemical analysis of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in the serum of mice of the C57BL/6 lineage, treated with the alkaloids or chloroquine. Results: In the in silico analysis, the prediction of low hepatotoxic potential for alkaloids 1 and 2 was observed, and this result was corroborated by the ALT dosage, which presented results similar to those of the control group. Alkaloid 3, however, presented contrasting data, indicating hepatotoxic potential in in silico prediction, however, low potential in in vivo analysis, with ALT values also close to the control group. All alkaloids under study showed potential for drug interactions. Conclusion: The alkaloids evaluated in this study showed promising metabolic and toxicity parameters and could be good adjuvants for malaria pharmacotherapy. However, these results need to be confirmed to follow the molecules in preclinical studies.

Detoxification Mechanism of Combined Chebulae Fructus， Glycyrrhizae Radix et Rhizoma and Prepared Aconiti Kusnezoffii Radix Cocta in Regulating Cardiac Metabolic Enzyme CYP2J3 / 中国实验方剂学杂志

ObjectiveTo investigate the detoxification mechanism of Chebulae Fructus， Glycyrrhizae Radix et Rhizoma and Prepared Aconiti Kusnezoffii Radix Cocta， and their effective components ellagic acid， liquiritin and aconitine based on cardiac cytochrome P450 （CYP450） system. MethodIn in vivo experiments， rats were randomly divided into control group， prepared Aconiti Kusnezoffii Radix Cocta group （0.25 g·kg-1）， Chebulae Fructus group （0.252 g·kg-1）， Glycyrrhizae Radix et Rhizoma group （0.25 g·kg-1） and combination group （0.25 g·kg-1 Chebulae Fructus+0.25 g·kg-1 Glycyrrhizae Radix et Rhizoma+0.25 g·kg-1 prepared Aconiti Kusnezoffii Radix Cocta， with prepared Aconiti Kusnezoffii Radix Cocta as standard）. After 8 days of administration， creatine kinase （CK） and lactate dehydrogenase （LDH） in rats were detected to observe the pathological changes of heart tissue. Real-time PCR and Western blot were performed to detect the mRNA and protein expressions of CYP2J3， respectively. In in vitro experiments， control group， aconitine group， ellagic acid group， liquiritin group and combination group （aconitine+ellagic acid+liquiritin） were set， and their effects on cell number， DNA content， reactive oxygen species （ROS） and mitochondrial membrane potential （MMP） were detected by high content analysis. The changes in the mRNA and protein expressions of CYP2J3 were also observed. ResultIn vivo experiments， compared with the control group， the prepared Aconiti Kusnezoffii Radix Cocta group had increased CK and LDH in serum （P<0.05， P<0.01）， while the combination group had decreased activities of CK and LDH. Additionally， pathological staining results showed that Chebulae Fructus and Glycyrrhizae Radix et Rhizoma reduced the cardiac toxicity caused by prepared Aconiti Kusnezoffii Radix Cocta. Real-time PCR found that compared with the control group， prepared Aconiti Kusnezoffii Radix Cocta down-regulated the mRNA level of CYP2J3 （P<0.05）， while up-regulated that expression when used in combination with Chebulae Fructus and Glycyrrhizae Radix et Rhizoma （P<0.01）. The protein and mRNA translation levels were basically consistent. In vitro experiments， high content analysis revealed that there was a decrease in the cell number， DNA content and MMP fluorescence value of the aconitine group （P<0.01） and the combination group （P<0.05， P<0.01）， and the fluorescence value of the combination group was higher than that of the aconitine group. Moreover， aconitine down-regulated the mRNA level of CYP2J3 （P<0.05）， but the down-regulating ability of aconitine was reversed in the combination group （P<0.05）. ConclusionThe detoxification mechanism of combined Chebulae Fructus， Glycyrrhizae Radix et Rhizoma and prepared Aconiti Kusnezoffii Radix Cocta is mainly that the combination of ellagic acid， liquiritin and aconitine can up-regulate the expression of CYP2J3， and promote the metabolism of arachidonic acid （AA） to produce epoxyeicosatrienoic acids （EETs）， thus reducing the cardiac toxicity， and this effect may start from the transcriptional link.

Distribution of gene mutation sites affecting artemisinin metabolism in the Kazak population in Xinjiang / 中国热带医学

@#Abstract: Objective　To detect the distribution of CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 loci affecting the metabolism of artemisinins in Kazak population in Xinjiang. To explore the pharmacogenetic background of the Kazak population in Xinjiang for artemisinin drugs and provide clinical decision support for the treatment and prevention of malaria based on artemisinin drugs. Methods　Six SNPs including CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 were selected for the sequencing experiment. 330 whole blood samples were collected from the Kazak population in Xinjiang. After extracting the whole blood DNA genome, multiplex PCR and high-throughput sequencing were used for genotyping. The allele frequencies were analyzed using the Hardy-Weinberg equilibrium. Results　In this study all SNPs follow the Hardy-Weinberg equilibrium (P>0.05), there was no significant difference in the distribution of SNPs between different genders (P>0.05). The number of successfully sequenced samples of CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 were 326, 319, 328, 318, 322 and 328 respectively. The frequencies of variant alleles of CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 in Kazak population are: 0.61%, 0%, 0%, 30.97%, 22.98%, 0%. Conclusions　Mutation alleles affecting the metabolism of artemisinins exist in the Kazak population in Xinjiang. When using artemisinins, the relationship between the drug effect and individual pharmacogenetic background should be further explored.

Effects of Compound troxerutin and poreine cerebroside injection on cytochrome P450 enzyme in vivo and in vitro / 中国药房

OBJECTIVE To investigate the effects of Compound troxerutin and poreine cerebroside injection on the activity of cytochrome P450 （CYP450） enzyme in vivo and in vitro. METHODS Human liver microsomes were incubated with Compound troxerutin and poreine cerebroside injection （volume fraction 0.05%-10%） and the specific probe substrates of CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6 and CYP3A4 for 30 min. The production of corresponding metabolites was detected by ultra-high performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS）， and the half inhibitory concentration （IC50） was calculated. The relative mRNA expression （i.e. induction multiple） of CYP450 enzyme was determined by real-time fluorescence quantitative PCR after human primary hepatocytes were incubated with Compound troxerutin and poreine cerebroside injection （volume fraction 0.05%-10%） or 3 positive inducers of CYP1A2， CYP2B6， CYP3A4 for 48 hours. Male SD rats were randomly divided into control group （normal saline+probe substrates of CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6， CYP3A4 8， 2， 1， 1， 10， 10， 8 mg/kg） and experimental group （Compound troxerutin and poreine cerebroside injection 0.9 mL/kg+probe substrates of CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6， CYP3A4 8， 2， 1， 1， 10，10， 8 mg/kg）， with 6 rats in each group. The pharmacokinetic parameters of probe substrates were detected by UPLC-MS/MS and Cocktail probe drug method. RESULTS After the lzqpharm@126.com treatment of 0.05%-10% Compound troxerutin and poreine cerebroside injection， the activities of CYP2B6， CYP2C8 and CYP2C19 in human liver microsomes had no significant change， and IC50 could not be fitted； IC50 of CYP1A2， CYP2C9， CYP2D6 and CYP3A4 were 419.90%， 97.78%， 176.00%， 19.42%， respectively. After the treatment of 0.05%-10% Compound troxerutin and poreine cerebroside injection， the average induction multiple of CYP3A4 mRNA in human primary hepatocytes （No. MHK） was 4.88 （and the average induction multiples of 2 concentration points were higher than 2）. After the treatment of Compound troxerutin and poreine cerebroside injection， AUC0－t and AUC0－∞ of CYP2C8， CYP2C9 and CYP2C19 substrates were increased significantly， CL of CYP2C8 and CYP2C19 substrates were decreased significantly， while t1/2 of CYP2C9 substrate was prolonged significantly （P＜0.05）. CONCLUSIONS Compound troxerutin and poreine cerebroside injection has no obvious inhibitory effect on CYP1A2， CYP2B6， CYP2C8， CYP2C9， CYP2C19， CYP2D6 and CYP3A4 in human liver microsomes in vitro， but can induce the mRNA expression of CYP3A4 in human primary hepatocytes in vitro， and can inhibit the activities of CYP2C8， CYP2C9 and CYP2C19 in rats in vivo.

Modification of C20 oxidase in tanshinone biosynthesis pathway / 中国中药杂志

Tanshinones are one of the main effective components of Salvia miltiorrhiza, which play important roles in the treatment of cardiovascular diseases. Microbial heterogony production of tanshinones can provide a large number of raw materials for the production of traditional Chinese medicine(TCM) preparations containing S. miltiorrhiza, reduce the extraction cost, and relieve the pressure of clinical medication. The biosynthetic pathway of tanshinones contains multiple P450 enzymes, and the catalytic element with high efficiency is the basis of microbial production of tanshinones. In this study, the protein modification of CYP76AK1, a key P450-C20 hydroxylase in tanshinone pathway, was researched. The protein modeling methods SWISS-MODEL, Robetta, and AlphaFold2 were used, and the protein model was analyzed to obtain the reliable protein structure. The semi-rational design of mutant protein was carried out by molecular docking and homologous alignment. The key amino acid sites affecting the oxidation activity of CYP76AK1 were identified by molecular docking. The function of the obtained mutations was studied with yeast expression system, and the CYP76AK1 mutations with continuous oxidation function to 11-hydroxysugiol were obtained. Four key amino acid sites that affected the oxidation acti-vity were analyzed, and the reliability of three protein modeling methods was analyzed according to the mutation results. The effective protein modification sites of CYP76AK1 were reported for the first time in this study, which provides a catalytic element for different oxidation activities at C20 site for the study of the synthetic biology of tanshinones and lays a foundation for the analysis of the conti-nuous oxidation mechanism of P450-C20 modification.

Late-stage cascade of oxidation reactions during the biosynthesis of oxalicine B in Penicillium oxalicum

Oxalicine B ( 1) is an α-pyrone meroterpenoid with a unique bispirocyclic ring system derived from Penicillium oxalicum. The biosynthetic pathway of 15-deoxyoxalicine B ( 4) was preliminarily reported in Penicillium canescens, however, the genetic base and biochemical characterization of tailoring reactions for oxalicine B ( 1) has remained enigmatic. In this study, we characterized three oxygenases from the metabolic pathway of oxalicine B ( 1), including a cytochrome P450 hydroxylase OxaL, a hydroxylating Fe(II)/α-KG-dependent dioxygenase OxaK, and a multifunctional cytochrome P450 OxaB. Intriguingly, OxaK can catalyze various multicyclic intermediates or shunt products of oxalicines with impressive substrate promiscuity. OxaB was further proven via biochemical assays to have the ability to convert 15-hydroxdecaturin A ( 3) to 1 with a spiro-lactone core skeleton through oxidative rearrangement. We also solved the mystery of OxaL that controls C-15 hydroxylation. Chemical investigation of the wild-type strain and deletants enabled us to identify 10 metabolites including three new compounds, and the isolated compounds displayed potent anti-influenza A virus bioactivities exhibiting IC50 values in the range of 4.0-19.9 μmol/L. Our studies have allowed us to propose a late-stage biosynthetic pathway for oxalicine B ( 1) and create downstream derivatizations of oxalicines by employing enzymatic strategies.

Activity prediction of human cytochrome P450 inhibitors based on multiple deep learning and machine learning methods / 中国药科大学学报

@#Inhibition of human cytochrome P450 (CYP) can lead to drug-drug interactions, resulting in serious adverse reactions.It is therefore crucial to accurately predict the inhibitory power of a given compound against a particular CYP isoform.This study compared 11 machine learning methods and 2 deep learning models based on different molecular representations.The experimental results showed that the CatBoost machine learning model based on RDKit_2d+Morgan outperformed other models in terms of accuracy and Mathews coefficient, and even outperformed previously published models.Moreover, the experimental results also showed that the CatBoost model not only had superior performance, but also consumed less computational resources.Finally, this study combined the top 3 performing models as co_model, which slightly outperformed the CatBoost model alone in terms of performance.

The evaluation value of cytochrome P450 2C19 *2 gene polymorphism, Helicobacter pylori infection for clopidogrel effect in patients with coronary heart disease / 中国医师进修杂志

Objective:To explore the evaluation of cytochrome P450 2C19 *2 (CYP2C19 *2) gene polymorphism and Helicobacter pylori (Hp) infection for clopidogrel efficacy in patients with coronary heart disease. Methods:The clinical data of 113 patients with coronary heart disease from February 2016 to March 2020 in Suzhou High-tech Zone People′s Hospital were retrospectively analyzed. The CYP2C19 *2 gene polymorphism was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method, the Hp infection was detected by 13C urea breath test. The patients were treated with clopidogrel, the effect after 4 weeks was evaluated. The receiver operating characteristic (ROC) curve was used to evaluate the CYP2C19 *2 gene polymorphism and Hp infection for evaluating clopidogrel effect in patients with coronary heart disease. Results:The CYP2C19 *2 genotype in patients with coronary heart disease conformed to Hardy-Weinberg balance ( χ2 = 0.33, P>0.05). Among 113 patients with coronary heart disease, Hp infection was in 27 cases, and Hp non-infection in 86 cases. Among Hp infection patients, CYP2C19 *2 gene GG was in 2 cases, GA in 6 cases, AA in 19 cases; among Hp non-infection patients, CYP2C19 *2 gene GG was in 23 cases, GA in 46 cases, AA in 17 cases, there was statistical difference in CYP2C19 *2 gene polymorphism between the two ( χ2 = 24.35, P<0.01). After clopidogrel treatment, effectiveness was in 79 cases, inefficiency in 34 cases. Among effectiveness patients, YP2C19 *2 gene GG was in 20 cases, GA in 43 cases, AA in 16 cases; among inefficiency patients, CYP2C19 *2 gene GG was in 5 cases, GA in 9 cases, AA in 20 cases, there was statistical difference in CYP2C19 *2 gene polymorphism between the two groups ( χ2 = 16.35, P<0.01). The rate of Hp infection in effectiveness patients was significantly lower than that in inefficiency patients: 12.66% (10/79) vs. 50.00% (17/34), and there was statistical difference ( χ2 = 18.23, P<0.05). ROC curve analysis result showed that the area under the curve of CYP2C19 *2 gene polymorphism combined with Hp infection for evaluating clopidogrel effect in patients with coronary heart disease was larger than CYP2C19 *2 gent GG, GA, AA and Hp infection alone evaluating (0.973 vs. 0.869, 0.679, 0.884 and 0.728) . Conclusions:The CYP2C19 *2 gene polymorphism is associated with Hp infection in patients with coronary heart disease, and the CYP2C19 *2 gene polymorphism combined with Hp infection has the evaluation value for the efficacy of clopidogrel.

Effect and mechanism of cytochrome P450 1A1 on regulating phagocytosis of macrophage / 中华危重病急救医学

Objective:To explore the effect and mechanism of cytochrome P450 1A1 (CYP1A1) on regulating phagocytosis of macrophage treated with Escherichia coli ( E.coli). Methods:① The mouse leukemia cells lines of monocyte macrophage RAW264.7 (RAW) were cultured in vitro and treated with 30 multiplicity of infection (MOI) dosages of E.coli for 40 minutes, glycerin control group was set up to observe the change of CYP1A1 during infection. ② The RAW cells with CYP1A1 overexpression (CYP1A1/RAW) and knock out (CYP1A1 KO/RAW) were cultured in vitro and treated with 30 MOI E. coli for 40 minutes, while the negative controlled RAW cells (NC/RAW) were established as control to observe the relationship between cell phagocytosis and CYP1A1 expression, and the effect of CYP1A1 on phagocytic receptor [scavenger receptor-A (SR-A)] and its signal pathway [mitogen-activated protein kinase (MAPK) pathway]. ③ NC/RAW and CYP1A1 KO/RAW cells were cultured in vitro and pretreated with 1 μmol/L extracellular signal-regulated kinase (ERK) inhibitor (U0126) for 2 hours, and then treated with 30 MOI E.coli for 40 minutes, phosphate buffered solution (PBS) control group was set up to observe whether the effect of CYP1A1 on phagocytosis through controlled the MAPK pathway. ④ The RAW cells were cultured in vitro and pretreated with 100 nmol/L CYP1A1 hydroxylase active product 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE] for 2 hours, and then treated with 30 MOI E.coli for 40 minutes, and PBS control group was set up to observe whether the effect of CYP1A1 on phagocytosis was related to CYP1A1 hydroxylating metabolite. ⑤ The RAW cells with overexpression CYP1A1 hydroxylase-activity mutation (CYP1A1m/RAW) were cultured in vitro and treated with 30 MOI E.coli for 40 minutes, the CYP1A1/RAW cells were set up as control group to observe whether the effect of CYP1A1 on phagocytosis was related to CYP1A1 hydroxylase-activity. Results:① Compared with glycerin control group, CYP1A1 mRNA expression was significantly increased by E.coli stimulation (2 -ΔΔCt: 7.79±0.71 vs. 1.00±0.00, P < 0.05), indicating that CYP1A1 might participate in regulating infection progress. ② Compared with NC/RAW cells, the number of E.coli colonies phagocytized by CYP1A1/RAW cells was significantly decreased after 40 minutes of E.coli stimulation (×10 3 CFU/mL: 4.67±3.06 vs. 15.67±5.03, P < 0.05), while CYP1A1 KO/RAW cells had a significant increase in the number of E.coli colonies phagocytized (×10 3 CFU/mL: 46.00±5.29 vs. 15.67±5.03, P < 0.05), suggesting that CYP1A1 might negatively control macrophage phagocytosis function. Meanwhile, compared with NC/RAW cells, the expression of SR-A mRNA in CYP1A1/RAW cells was significantly down-regulated (2 -ΔΔCt: 0.31±0.03 vs. 1.00±0.00, P < 0.05), and the activation level of ERK was significantly reduced. However, the expression of SR-A mRNA in CYP1A1 KO/RAW cells was significantly up-regulated (2 -ΔΔCt: 3.74±0.25 vs. 1.00±0.00, P < 0.05), and the activation of ERK was enhanced, indicating that CYP1A1 could negatively regulate phagocytic receptors and their signaling pathways.③ Compared with PBS, U0126 pretreatment significantly inhibited the CYP1A1 knockout induced upregulation of SR-A mRNA expression (2 -ΔΔCt: 0.62±0.05 vs. 4.38±0.39, P < 0.05) and ERK activation, and inhibited the enhancement of phagocytosis in macrophages induced by CYP1A1 knock out [ E.coli colonies phagocytized by cells (×10 3 CFU/mL): 12.67±1.15 vs. 45.33±4.16, P < 0.05], suggesting that CYP1A1 inhibited macrophage phagocytosis function by regulating ERK activation. ④ Compared with PBS, the phagocytosis of RAW cells pretreated with 12(S)-HETE did not change significantly [ E.coli colonies phagocytized by cells (×10 3 CFU/mL): 17.00±1.00 vs. 16.33±2.52, P > 0.05], suggesting that CYP1A1 might not control phagocytosis function by its hydroxylase-activity metabolism 12(S)-HETE. ⑤ Compared with CYP1A1/RAW cells, there was no significant change in the phagocytic function of CYP1A1m/RAW cells [ E.coli colonies phagocytized by cells (×10 3 CFU/mL): 3.67±1.15 vs. 3.33±0.58, P > 0.05], suggesting that CYP1A1 might not control phagocytosis function by its hydroxylase-activity. Conclusion:CYP1A1 can negatively regulate the phagocytosis of macrophages by inhibiting the activation of ERK and reducing the expression of SR-A, but this regulatory effect is not related to the activity of CYP1A1 hydroxylase and its pro-inflammatory metabolism 12(S)-HETE.

Efficacy analysis of anti-platelet in the treatment of high-risk non-disabling ischemic cerebrovascular events guided by point-of-care testing of CYP2C19 gene / 中华神经科杂志

Objective:To explore the efficacy and safety of different anti-platelet regimens in the treatment of high-risk non-disabling ischemic cerebrovascular events (HR-NICE) guided by point-of-care testing of CYP2C19 gene. Methods:A single-centre, prospective, randomised, open-label, and blinded endpoint design was uesd in the study. From July 2020 to January 2022, HR-NICE patients were enrolled in the Stroke Green Channel and Department of Neurology of Xuzhou Central Hospital, and all patients were scraped the buccal mucosa for screening for CYP2C19 loss-of-function allele carriers by point-of-care testing . Patients with intermediate metabolism were defined as those who carried 1 loss-of-function allele and patients with poor metabolism were those who carried 2 loss-of-function alleles. This study reduced the test turnaround time to 1 hour by using a fully automated medical polymerase chain reaction analyzer for a point-of-care test of CYP2C19 genotype. CYP2C19 loss-of-function allele carriers were divided according to the random number table method into the conventional treatment group (clopidogrel 75 mg, once a day), the ticagrelor group (ticagrelor 90 mg, twice a day) and the intensive dose group (clopidogrel 150 mg, once a day) separately combined with aspirin (100 mg, once a day) dual antiplatelet for 21 days. Baseline information, Acute Stroke Org 10172 Treatment Trial staging, 90-day modified Rankin Scale score, occurrence of adverse events and severe adverse events were collected for all the 3 groups. The primary efficacy outcome was new stroke within 90 days, and the primary safety outcome was severe or moderate bleeding within 90 days. Results:A total of 716 patients were included: 240 in the conventional treatment group, 240 in the ticagrelor group and 236 in the intensive dose group. There was no statistically significant difference between the 3 groups at baseline (all P>0.05). There were 26 cases (10.8%) with new stroke events in the conventional treatment group, 11 cases (4.6%) in the ticagrelor group and 4 cases (1.7%) in the intensive dose group, with statistically significant differences among the 3 groups (χ 2=19.28, P<0.05), and the differences between the conventional treatment group and the ticagrelor group (χ 2=6.59, P=0.010) and between the conventional treatment group and the intensive dose group (χ 2=16.83, P<0.001) were statistically significant, whereas the difference between the ticagrelor group and the intensive dose group was not statistically significant ( P>0.05). In the 3 groups, there was 1 case (0.4%) of severe bleeding in the conventional treatment group, 6 cases (2.5%) in the ticagrelor group and none in the intensive dose group, which showed statistically significant differences (χ 2=7.23, P<0.05), and there was statistically significant difference between the ticagrelor group and the intensive dose group ( P=0.030). Among the patients with intermediate CYP2C19 metabolism, there were 13 cases (13/158, 8.2%) with 90-day recurrent stroke in the conventional treatment group, 4 cases (4/153, 2.6%) in the ticagrelor group, and 0 case (0/159) in the intensive dose group, with statistically significant difference (χ 2=16.04, P<0.001), and the differences between the intensive dose group and the conventional treatment group were statistically significant (χ 2=13.64, P<0.001), whereas there was no statistically significant difference between the intensive dose group and the ticagrelor group ( P>0.05). In the patients with 90-day recurrent stroke in the intensive dose group, there was 0 case (0/159) with intermediate metabolism and 4 cases (4/77,5.2%) with poor metabolism, with statistically significant differences ( P=0.011), whereas there were no statistically significant differences in the conventional treatment group and the ticagrelor group ( P>0.05). Conclusions:Screening carriers of CYP2C19 loss-of-function alleles by point-of-care testing can quickly and precisely guide the treatment of patients with non-cardiogenic HR-NICE. An intensive clopidogrel dose of 150 mg, once a day combined with aspirin was effective in reducing stroke recurrence with less occurrence of any bleeding and adverse events, and patients with intermediate CYP2C19 metabolism may be the best population to benefit.

Dual antiplatelet therapy in patients with minor stroke or high-risk transient ischemic attack carrying CYP2C19 loss-of-function alleles / 国际脑血管病杂志

Dual antiplatelet therapy has been widely used for the secondary prevention in patients with minor stroke and high-risk transient ischemic attack (TIA). Currently, the commonly used antiplatelet drugs are aspirin and clopidogrel. The therapeutic effect of antiplatelet drugs varies among individuals, namely platelet resistance. Among them, aspirin resistance is often caused by poor drug compliance, while clopidogrel resistance is often associated with CYP2C19 allele mutations. Patients with minor stroke and high-risk TIA carrying CYP2C19 loss-of-function alleles have poor preventive effects when using clopidogrel. Early screening of the CYP2C19 loss-of-function alleles and targeted measures can benefit such patients. This article reviews the research progress on the selection of antiplatelet therapy for minor stroke or high-risk TIA patients carrying the CYP2C19 loss-of-function alleles.

Research progress regarding CYP3A gene family in gastric cancer / 中南大学学报(医学版)

Cytochrome P450 family 3 subfamily A(CYP3A),a major member of cytochrome P450(CYP)family,is one of the most important drug metabolizing enzymes in human.CYP3A includes 4 gene subtypes(CYP3A4,CYP3A5,CYP3A7,and CYP3A43),which is involved in 60%of drug metabolism in the human.It is not only widely distributed in normal tissues,but also significantly overexpressed in various tumor tissues.Recently,CYP3A has attracted great attention due to its involvement in the progression from chronic atrophic gastritis to gastric cancer,as well as the differential metabolism and resistance of chemotherapeutic drugs.Targeting CYP3A gene mediated-prodrug provides new ideas for the treatment of gastric cancer and is expected to become a new target for the diagnosis and treatment of gastric cancer.

Cloning and expression analysis of cytochrome P450 AsCYP71D1 from Aquilaria sinensis (Lour.) Gilg / 药学学报

Cytochrome P450 (CYP450) is a kind of superfamily oxidase containing heme, which is distributed in various aerobic organisms. They are widely involved in the biosynthesis of terpenoids, alkaloids, flavonoids, fatty acids, etc. In this study, the full-length cDNA sequence of a P450 was cloned by reverse transcription-PCR (RT-PCR) and rapid amplification of cDNA ends (RACE) technology, with the specific primers that designed according to the sequence of a transcript annotated as P450 from the Aquilaria sinensis (Lour.) Gilg transcriptome database. The tissue expression and subcellular localization were also studied. The full-length cDNA of the cloned P450 gene is 1 920 bp, with 88 bp 5′-untranslated region (UTR), 344 bp 3′-UTR and a 21 bp polyA tail, and 1 488 bp open reading frame (ORF), encoding 495 amino acids. Sequence alignment revealed that the protein belonged to CYP71D family of cytochrome P450 family, and named AsCYP71D1. Tissue expression analysis indicated that AsCYP71D1 was mainly expressed in stem. Further subcellular localization of onion epidermis showed that AsCYP71D1 was expressed in cytoplasm, nucleus and cell membrane. This study will provide a foundation for further research on its function in agarwood sesquiterpene biosynthesis.

Correlation of maternal drug use and cytochrome P450 geneticpolymorphisms with congenital heart disease in offspring / 西安交通大学学报(医学版)

【Objective】 To investigate the association of maternal medication during early pregnancy and cytochrome P450 (CYP450) genetic polymorphisms with the risk of congenital heart disease (CHD) in offspring. 【Methods】 We selected 127 pregnant women with CHD fetuses as the observation group and 132 pregnant women with non-CHD fetuses as the control group. Their characteristics and medication history were investigated, and CYP450 polymorphisms were detected. Logistic regression analysis was used to assess the association between maternal medication, CYP450 gene variations, and offspring CHD risk. 【Results】 The risk of CHD in offspring was higher in the observation group with maternal use of ovulation induction drugs, antihypertensive drugs, antibiotics, antidepressants, miscarriage prevention drugs, and traditional Chinese medicine (P<0.05). The A/T and T/T genotypes in rs1065852 and the C/G and G/G genotypes in rs16947 increased the risk of CHD in offspring compared to their respective genotypes. The risk of CHDs in offspring increased with the presence of risk genotypes (A/T or T/T) at the rs1065852 locus of the maternal CYP450 gene and early pregnancy medication use (P<0.05); the same was observed for risk genotypes (C/G or G/G) at the rs16947 locus (P<0.05). 【Conclusion】 Maternal medication during early pregnancy may be associated with offspring CHD, and the rs1065852 and rs16947 loci of CYP450 are significantly related to the risk of CHD in offspring.

Research progress on metabolic activation of indirect carcinogens by CYP1A1 / 中国职业医学

Cytochrome P450 enzyme 1A1 (CYP 1A1) is one of the main members of CYP1A subfamily, which participates in metabolizing and activating a variety of indirect carcinogens. CYP1A1 can induce carcinogenesis by participating in activating exogenous compounds to produce intermediates or active metabolites that bind to specific biomolecules. CYP1A1 plays a critical role in the metabolic activation of benzo(a)pyrene e ［B(a)P］, and plays a key role in activating the toxic and carcinogenic effects of B(a)P. CYP1A1 involves in the metabolic activation of 7,12-dimethyl benzanthracene and 2-amino-1-methyl-6-phenylimidazo［4,5-b］pyridine (PhIP), and plays an important role in PhIP-induced genotoxicity. CYP1A1 is the main enzyme to metabolize and activate 7H-dibenzo［c,g］carbazole (DBC), a key factor in the carcinogenic effect of DBC. CYP1A1 is also associated with metabolic activation of indirect carcinogens such as aflatoxin B1, 3-nitrobenzene, and naphthalene. Inhibition of the catalytic activity of CYP1A1 can decrease the CYP1A1-mediated activity of carcinogens, thus playing a role in the prevention and treatment of malignant tumors.

Effects of single nucleotide polymorphism of drug metabolizing enzyme cytochrome P450 on the efficacy of inhaled cortisol hormone in asthmatic children / 中国临床药理学与治疗学

AIM: To elucidate the relationship between childhood asthma susceptibility and clinical efficacy of inhaled glucocorticoids (ICS) in children with different genotypes of asthma by exploring rs776746 and rs15524 single nucleotide polymorphisms (SNPs) of cytochrome P450 enzyme 3A5 (CYP3A5) gene in asthmatic children and healthy children. METHODS: The CYP3A5 gene rs776746 and rs15524 polymorphic sites were detected in 79 children (Case group) with asthma of Han nationality and 100 healthy children (Control group) who met the inclusion criteria admitted to the Northern Theater General Hospital in Northeast China from October 2016 to October 2020, and genotype, allele and linkage analysis were performed. The case group was given inhaled glucocorticoids by nebulised inhalation for 3 months, and lung function and exhaled breath nitric oxide (FeNO) were measured at entry and after treatment, and asthma control score C-ACT/ACT was done after treatment, so as to compare the prevalence of different genotypes and the differences in the above test index scores. RESULTS: There was complete linkage disequilibrium at rs776746 and rs15524 loci. There were three genotypes of T/T, T/C and C/C at rs776746 locus of CYP3A5 gene. There were significant differences in the frequency of different genotypes between the case group and the control group (χ