Synthesis and anti-HBV activity of novel 5-substituted pyridin-2(1H)-one derivatives

Four novel 5-substituted pyridine-2(1H)-one derivatives were designed and synthesized by using addition-elimination reactions.The structures of these novelly synthesized compounds were verified by ~1H NMR,ESI-MS and single crystal X-ray diffraction.Furthermore,all four compounds(most notably compound 7a) were found to be highly efficient against hepatitis B virus (HBV) in cultured HepG2 2.2.15 cell,making them promising drug candidates for potential bioactive molecule against hepatitis B.

The Progress of Anti-HBV Constituents from Medicinal Plants in China

Hepatitis B virus(HBV)infection causing acute and chronic hepatitis is a serious problem worldwide,whereas the current treatment methods are unsatisfactory.Traditional Chinese herbs that have long been used for medicinal purposes are fascinating sources for novel anti-HBV candidates.This paper summarizes the progress of anti-HBV constituents from diverse medicinal plants in China to provide information for searching new anti-HBV drugs from natural sources.

Cloning,Eukaryotic Expression of Human ISG20 and Preliminary Study on the Effect of Its Anti-HBV

Human ISG20 gene was cloned and the effect of its anti-HBV was primarily studied. The ISG20 gene was amplified from HeLa cells by RT-PCR and recombinant vector expressing ISG20 was constructed by genetic engineering. The overexpression of ISG20 in HepG2 cells was detected by Western blot and the levels of secretion of HBs antigen and HBe antigen tested by ELISA. The results showed that： （1） Sequence of ISG20 cloned was consistent to that published in Genebank; （2） Recombinant vector expressing ISG20 could be expressed in HepG2 cells by transfection; （3） The overexpression of ISG20 protein could reduce the levels of the secretion of HBs antigen and HBe antigen in transfected HepG2 cells. It was suggested that the overexpression of recombinant ISG20 in culture cells could reduce the synthesis of HBV proteins.

Design, synthesis and in vitro evaluation of L-amino acid esters prodrugs of acyclic nucleoside phosphonates as anti-HBV agent

A series of novel L-amino acid esters prodrugs of acyclic nucleoside phosphonates was synthesized and their anti-HBV activity was evaluated in HepG2 2.2.15 cells. Compound 1d exhibited more potent anti-HBV activity and lower cytotoxicity than those of adefovir dipivoxil with EC50 and CC50 values of 0.207 μmol/L and 2530 μmol/L, respectively.

Design,synthesis and anti-HBV activity of novel bis(trifluoroethyl)phosphonomethyl ether derivatives of acyclovir

A series of novel bis（trifluoroethyl）phosphonomethyl ether derivatives of acyclovir was synthesized and their in vitro anti-HBV activity was evaluated in HepG2 2.2.15 cells. In contrast to acyclovir, most of the described phosphonates emerged as potent inhibitors of HBV replication. Especially, the most active compound 11 with IC50 value of 2.92 μmol/L was 33 times more potent than acyclovir with ICso value of 100 μmol/L.

Synthesis and anti-HBV activity of S-substituted 7-mercapto-4-methylcoumarin analogs

Twelve S-substituted 7-mercapto-4-methylcoumarin analogs were synthesized and evaluated for the inhibition to HBV in HepG2 2.2.1.5 cell. Among them, ten compounds exhibited potent inhibition to HBsAg and/or HBeAg with the IC50 values of sub μmol/L level. The ICso of anti-HBsAg activities of 5c and 5j reached 0.01 μmol/L respectively which were 16 fold more potent than that of 3TC. Compounds 3, 5e, 5g, 5h and 5i showed admirable inhibitory activity to both HBsAg and HBeAg. The bioassay results indicated the S-substituted 7-mercapto-4-methylcoumarin analogs merit attention as novel anti-HBV agents.

基于血清抗-HBc定量建立慢性HBV感染者显著肝组织病理学改变的无创诊断模型

目的建立慢性HBV感染者显著肝组织病理学改变的诊断模型,并评估模型的诊断价值。方法选取2011年12月—2017年12月在上海市公共卫生临床中心肝胆内科住院并进行肝活检且未经抗病毒治疗的457例慢性HBV感染者,收集患者的肝活检病理结果及常规实验室指标,并进行抗-HBc定量检测。依据Scheuer方法进行炎症分级(G)及纤维化分期(S),分为显著与非显著肝坏死性炎症、肝纤维化和肝损伤组。根据单因素分析和多因素logistic回归分析构建预测显著肝坏死性炎症、显著肝纤维化和显著肝损伤的数学模型,并与FIB-4、GPR、APRI、RPR利用ROC曲线分析评估模型的预测性能,根据ROC曲线下面积(AUC)比较诊断价值。结果457例患者中显著肝坏死性炎症(G≥2)178例,非显著肝坏死性炎症(G<2)279例,显著肝纤维化(S≥2)248例,非显著肝纤维化(S<2)209例,显著肝损伤(G≥2或/和S≥2)264例,非显著肝坏死性炎症(G<2和S<2)193例。根据单因素分析和多因素logistic回归分析结果,分别建立由抗-HBc、AST、PLT、TTR等指标组成的预测显著肝坏死性炎症的数学模型M-SHN,由抗-HBc、PLT、ChE、TTR、性别等指标组成的预测显著肝纤维化的数学模型M-SHF,及由抗-HBc、PLT、TTR、性别等指标组成的预测显著肝损伤的数学模型M-SHI,并通过ROC曲线分析各模型的预测价值,M-SHN模型的AUC为0.826(95%CI:0.788~0.860),M-SHF模型的AUC为0.776(95%CI:0.735~0.814),M-SHI模型的AUC为0.789(95%CI:0.748~0.825)。结论基于患者常规实验室指标及血清抗-HBc定量,建立了M-SHN、M-SHF、M-SHI模型,对于显著肝坏死性炎症、显著肝纤维化、显著肝损伤有较为可靠的预测价值,可帮助临床评估患者是否需要进行抗病毒治疗。

Geranyl phenyl ethers from Illicium micranthum and their anti-HBV activity

Fourteen new geranyl phenyl ethers(1–14)along with three known compounds(15–17)were isolated from Illicium micranthum,and their structures were elucidated by comprehensive spectroscopic methods.Illimicranins A–H(1–8)were characterized as geranyl vanillin ethers,while 9 and 10 were dimethyl acetal derivatives.Illimicranins I and J(11 and 12)were rare geranyl isoeugenol ethers.Illimicranins K and L(13 and 14)represented the first example of geranyl guaiacylacetone ether and geranyl zingerone ether,respectively.Compounds 1,2 and 15 exhibited anti-HBV(hepatitis B virus)activity against HBsAg(hepatitis B surface antigen)and HBeAg(hepatitis B e antigen)secretion,and HBV DNA replication.

Synthesis and anti-HBV evaluation of mono L-amino acid ester, mono non-steroid anti-inflammation drug carboxylic ester derivatives of acyclonucleoside phosphonates

A series of mono L-amino acid ester, mono non-steroid anti-inflammation drug （NSAID）, carboxylic ester derivatives of acyclonucleoside phosphonates were prepared by using a ＂one pot synthesis＂ method and their in vitro anti-HBV activity were evaluated in HepG 2.2.15 cells. Compound 9a exhibited more potent anti-HBV activity and lower cytotoxicity than those of adefovir dipivoxil with IC50 and selective index （SI） values of 0.48μmol/L and 763.72, respectively.

规范抗乙肝病毒治疗与肝癌术后肿瘤复发的关系

目的通过肝癌多学科协作(multi-disciplinary team,MDT)诊治模式下实施规范抗乙肝病毒方案在肝癌病人中的应用,探讨抗乙肝治疗与肝癌术后生存率及肿瘤复发的关系。方法回顾性分析2015年1月至2017年12月经我院肝癌MDT讨论后实施规范抗乙肝治疗病人(治疗组,36例)和未进行规范抗乙肝治疗病人(对照组,34例)的临床资料,比较两组病人肝癌术后肿瘤复发情况及1、3、5年内生存率。结果两组病人一般情况、实验室指标、肿瘤大小及术前HBV-DNA差异无统计学意义(P>0.05),手术后为期5年的随访中治疗组病人1、3、5年肿瘤复发率分别为8.3%、33.3%、41.6%;对照组1、3、5年肿瘤复发率分别为11.7%、41.1%、64.7%,差异均具有统计学意义(P<0.05)。治疗组病人1、3、5年生存率分别为91.6%、66.7%、58.3%;对照组1、3、5年生存率分别为88.2%、58.8%、29.5%,差异均具有统计学意义(P<0.05)。结论HBV相关性肝癌病人在经过肝癌MDT制定方案,术前和术后实施规范抗乙肝病毒治疗,可有效降低肝癌复发,提高远期生存率。

茵陈蒿的化学成分及其药理活性研究

目的研究传统中药材茵陈蒿的化学成分,以期从中发现具有药理活性的先导化合物。方法利用大孔吸附树脂、硅胶、ODS等柱色谱与半制备型高效液相色谱相结合对茵陈蒿中的化合物进行分离纯化,并结合化合物的理化性质与核磁共振波谱数据对分离得到的化合物进行结构鉴定。此外,采用MTT法和ELISA法对分离得到的化合物进行了抗肝癌以及抗HBV活性测试。结果从采自山西的茵陈蒿的乙醇提取物中分离出10个单体化合物,活性测试结果表明上述化合物未显示抗肝癌以及抗HBV活性。结论化合物1和2为新化合物,化合物1,3,4为烯炔类化合物,化合物2和10为脂肪酸类化合物,化合物5-9为萜类化合物。

32例药物性肝损伤肝衰竭患者影响因素分析及预后评价

目的探讨药物性肝损伤(DILI)患者发生药物性肝损伤肝衰竭(DILF)的危险因素,并对预后情况进行分析。方法选取2015年1月-2022年12月期间南通市肿瘤医院收治的DILI和肝衰竭患者,其中DILI 221例,男性97例、女性124例,年龄46(37,60)岁;DILF 32例,男性17例、女性15例,年龄47(40,58)。比较DILI、DILF患者以及不同临床结局DILF患者临床资料,运用多因素分析探讨影响DILI患者肝衰竭发生以及DILF预后不良的危险因素。结果DILI患者用药情况具体为中药/中成药120例(54.3%)、抗结核药物37例(16.7%)、感冒药29例(13.1%)、抗肿瘤药物11例(5.0%)、降血脂药物7例(3.2%)、抗甲状腺药物5例(2.3%)、神经精神类药物4例(1.8%)及其他药物8例(3.6%);DILF患者用药情况具体为抗结核药物14例(43.7%)、中药/中成药12例(37.5%)、感冒药2例(6.2%)以及抗肿瘤药物、免疫调节剂、精神类药物、降血压药各1例(3.1%)。单因素分析得出DILI患者HBV感染、抗结核药物、WBC、AST、TBil、血氨、INR及MELD评分低于DILF患者,而DILI患者PLT、PTA高于DILF患者,两组比较差异有统计学意义(P<0.05)。将肝功能、凝血功能等反映肝损伤严重程度的指标除外,多因素分析提示HBV感染、WBC升高及PLT减少为DILI发生肝衰竭的独立危险因素(P<0.05)。DILF患者系统治疗后好转、恶化病例分别为14例、18例。单因素分析得出好转、恶化DILF患者ALT、TBil、Scr、PTA、INR、MELD评分等比较,差异具有统计学意义(P<0.05)。多因素分析提示低PTA是DILF患者预后不良的独立危险因素(P<0.05)。结论DILI患者发生肝衰竭存在着多项危险因素,包括WBC升高及PLT减少。HBV感染病例服用抗结核药、中药/中成药等高风险致肝损伤药物时,需要警惕AILF的发生。另外,AILF患者存在凝血功能异常时,往往预后较差。

红树木榄生境中可培养细菌物种多样性及其体外抗乙肝病毒活性研究

红树生境中微生物菌群丰富,其次生代谢产物结构新颖,是挖掘新型药物的重要来源。该研究利用纯培养技术和16S rRNA分子生物学技术确定细菌种属,并进行物种多样性分析;以HepG2.2.15细胞株为模型,通过MTT和PCR技术测试细菌代谢产物的抗乙肝病毒活性;使用LC-HRMS技术对活性菌株代谢产物进行初步分析,初步评价木榄沉积物、根、叶以及胚轴的可培养细菌多样性以及细菌代谢物生物活性,寻找抗乙肝病毒的药源菌株。结果表明:(1)共获得细菌59种,分属于4门5纲14目23科36属,其中芽孢杆菌为优势菌属;菌株GXIMD07402、GXIMD07665、GXIMD07384分别为Pseudooceanicola属、Thioclava属和Aestuariibaculum属的潜在新种。(2)抗乙肝病毒活性结果显示GXIMD07366、GXIMD07616、GXIMD07384、GXIMD07550、GXIMD07445X提取物能显著降低HepG2.2.15细胞上清液中HBV DNA水平(P<0.05),抑制率分别为51%、47%、63%、52%、47%。(3)初步鉴定强活性菌株GXIMD07384的4个主要代谢产物有adenosine、cyclo(L-Pro-L-OMet)、acremine G和7,8-dimethylbenzo[g]pteridine-2,4(1H,3H)-dione。综上认为,木榄生境中可培养细菌物种多样性丰富且含有能产生抗乙肝病毒活性化合物的菌株。该研究结果为后续海洋微生物资源的应用提供了基础。

药用红树木榄胚轴抗HBV化学成分的研究

红树木榄(Bruguiera gymnorhiza)胚轴为京族常用治疗乙肝中草药。为研究木榄胚轴中化学成分及其抗乙型肝炎病毒(hepatitis B virus, HBV)活性,该文采用MTT法和实时荧光定量PCR方法测定木榄胚轴不同萃取部位抗HBV活性,使用现代色谱和波谱方法对活性萃取部位的化学成分进行分离鉴定,并测试获得的化学成分抗HBV活性。结果表明:(1)红树木榄胚轴的正丁醇萃取部位具有抗HBV活性;(2)从中分离了11个化合物,分别鉴定为尿嘧啶(1)、胸腺嘧啶(2)、腺嘌呤核苷(3)、oryzalactam (4)、正丁基-O-D-吡喃果糖苷(5)、nortetillapyrone (6)、(4R,6S)-4-methoxyl-2,3-dihydroaquilegiolide (7)、(4R,6S)-2-dihydromenisdaurilide (8)、没食子儿茶素(9)、1-(4-hydroxy-3-methoxy)-phenyl-2-[4-(1,2,3-trihydroxypropyl)-2-methoxy]-phenoxy-1,3-propandiol (10)和(-)-南烛木树脂酚-9-O-β-D-木吡喃糖苷(11),其中化合物4、5、7和8为首次从药用红树木榄中获得,化合物4具有抗HBV活性,其抑制率为23.59%。该研究结果丰富了木榄胚轴抗HBV化学成分。

Immunization against Hepatitis B Virus: Serological Status of West African Populations Residing in Cotonou (Benin) and Dakar (Senegal)

Background: According to WHO estimates, by 2022 over 296 million people are living with chronic hepatitis B virus (HBV) infection, and over 820,000 have died from complications. In sub-Saharan African countries such as Benin and Senegal, few research studies have addressed the issue of HBV immunization. Objective: The main objective of this study was to evaluate immunization against the hepatitis B virus in populations residing in Cotonou and Dakar by titrating anti-HBs antibodies (Ab) and detecting total anti-HBc immunoglobulins (Ig). Materials and Methods: This was a prospective, descriptive, analytical study of two West African populations recruited in Dakar at the Laboratory of Medical Biology (LBM) of the General Hospital Idrissa Pouye (HOGIP) and in Cotonou at the LBMs of the health centres of the Cotonou archdiocese. HBsAg-negative patients constituted our study population. The study took place in November-December 2019 for Dakar and February-March 2020 for Cotonou. Anti-HBs antibodies were tested and titrated. In the event of anti-HBs positivity, total anti-HBc was determined. A microparticle chemiluminescence immunoassay was used for marker determination. The detection threshold was 2.50 IU/L for anti-HBs. Excel and IBM SPSS Statistics software were used for data analysis. Subjects’ sociodemographic characteristics were collected using a questionnaire, as was knowledge of their vaccination status. The study was approved by the ethics committees in Benin and Senegal. Results: A total of 394 HBs antigen-negative participants were recruited: 205 in Cotonou and 189 in Dakar. The population was predominantly female, with 65.36% (N = 134) and 57.14% (N = 108) women in Cotonou and Dakar respectively. The median age of participants was 29 years in Cotonou, with extremes of 10 and 65 years, versus 39 years in Dakar, with extremes of 6 and 93 years. Some participants claimed to be unaware of their vaccination status: 33.17% in Cotonou and 56.61% in Dakar. The total prevalence of anti-HBs-positive subjects was 88.78% (N = 182) in Cotonou and 98.41% (N = 186) in Dakar. In Cotonou (N = 205), 35.61% (N = 73) of subjects had protective anti-HBs levels between 11.60 IU/L and 10,000 IU/L. In Dakar, 61.38% (N = 116) of subjects had protective HBV immunity, with anti-HBs titres ranging from 10.30 IU/L to 11357 IU/L. In Cotonou, 80.82% (N = 59) of immunized subjects (N = 73) had anti-HBC antibodies, compared with 84.48% (N = 98) of immunized individuals (N = 116) in the population recruited in Dakar, indicating immunization following HBV infection. Conclusion: Our study involved a predominantly female population, many of whom were unaware of their serological status. Vaccination policies and knowledge of the viral hepatitis B epidemic need to be strengthened.

拉米夫定联合抗乙肝胎盘转移因子治疗慢性乙型肝炎疗效及对T淋巴细胞的影响

目的探讨拉米夫定联合抗乙肝胎盘转移因子治疗慢性乙型肝炎疗效及对T淋巴细胞的影响。方法选择2021年1月至2022年1月南阳市中心医院收治的90例慢性乙肝患者,按照随机数字表法分为研究组与对照组各45例,两组患者均给予补充血浆、人血白蛋白,应用护肝、抗感染药物等常规对症治疗。对照组接受拉米夫定治疗,研究组使用拉米夫定及抗乙肝胎盘转移因子联合治疗。两组均持续治疗12周,比较治疗前后两组乙肝病毒标志物的阴转情况及肝功能指标[总胆红素(TBil)、天冬氨酸转氨酶(AST)、丙氨酸转氨酶(ALT)、总胆汁酸(TBA)],观察两组用药前后的免疫功能变化,并统计比较两组不良反应发生率。结果治疗前,两组乙肝病毒标志物各项指标比较,差异无统计学意义(P>0.05);治疗后,研究组HBV-DNA阴转、HBeAg阴转、HBeAg/抗-HBe人数占比明显高于对照组,差异有统计学意义(P<0.05)。治疗前,两组患者肝功能相关指标对比,差异无统计学意义(P>0.05);治疗后,研究组TBil、AST、ALT、TBA水平低于对照组,差异有统计学意义(P<0.05)。治疗前,两组T淋巴细胞亚群各指标比较,差异无统计学意义(P>0.05);治疗后,两组CD3^(+)、CD4^(+)水平及CD4^(+)/CD8^(+)值高于治疗前,CD8^(+)水平低于治疗前,且研究组的改善幅度大于对照组,差异有统计学意义(P<0.05)。两组不良反应发生率比较,差异无统计学意义(P>0.05)。结论拉米夫定联合抗乙肝胎盘转移因子对慢性乙肝的治疗效果显著,有利于改善肝功能指标,调节免疫功能水平,进一步提高患者的临床治疗效果,且具有一定安全性。

青蒿鞣质抗病毒活性研究

从青蒿水提物中分离得到一种缩合鞣质 (CTA) ,在体外进行了抗HSV 2和HBV活性研究。结果表明 ,CTA具有显著抗HSV 2活性 ,与阳性对照药物阿昔洛韦 (ACV)比较 ,CC50 分别为 6 .84mg/mL和 3.6 9mg/mL ,IC50 分别为 0 .16 2mg/mL和 0 .138mg/mL ,表明CTA的细胞毒性比ACV小 ,最低抗病毒活性浓度相当。用含CTA的培养基培养HepG2 .2 .1.5细胞 ,结果表明在浓度为 2 .5mg/mL时有轻微细胞毒性 ,在浓度为 2 .5～ 0 .15 6mg/mL对HepG2 .2 .1.5细胞表达的HBeAg具有显著的抑制作用 ,0 .6 2 5mg/mL浓度档培养第 12d对HBeAg抑制率达到 90 .4 5 % 。

马蹄金素杂环衍生物的合成及抗乙肝病毒活性

以马蹄金素[N-(N-苯甲酰基-L-苯丙氨酰基)-O-乙酰基-L-苯丙氨醇,MTS]为先导化合物,设计并合成了16个杂环取代的马蹄金素衍生物,其结构经NMR,ESI-MS和元素分析确证.以HepG2 2.2.15细胞为乙肝病毒(HBV)载体,对合成的马蹄金素衍生物进行了抗HBV活性测试.实验结果表明,在测试浓度范围内,化合物5b(IC50=8.20μg/L,SI=10.26),5g(IC50=5.58μg/L,SI=22.78)和5i(IC50=5.07μg/L,SI=16.67)的抗HBV活性较强;在8μg/mL的浓度下,其抑制率分别为56.57%,67.06%和66.83%.

空心莲子草抗病毒活性成分研究

目的 研究空心莲子草Alternanthera philoxeroides中抗HBV活性成分。方法 以硅胶柱、凝胶柱色谱分离，制备HPLC纯化，采用MS、NMR等波谱方法进行结构鉴定；用ELISA法对各化合物进行抗HBV活性检测。结果 从空心莲子草中分离鉴定了10个化合物，分别为齐墩果酸（oleanolic acid，Ⅰ）、3-O-β-D-吡喃葡萄糖醛基齐墩果酸苷（oleanolic acid 3-O-β-D-glucuronopyranoside，Ⅱ）、齐墩果酸-28-O-β-D-吡喃葡萄糖苷（oleanolic acid 28-O-β-D-ducopyranoside，Ⅲ）、竹节参苷Ⅳa甲酯（chikusetsusaponin Ⅳ a methyl ester，Ⅳ）、4，5-dihydroblumenol（Ⅴ）、N-trans-feruloyl 3-methyldopamine（Ⅵ）、N-trans-feruloyltyramine（Ⅶ）、3β羟基豆甾-5-烯-7-酮（3phydrox-ystigmast-5-an-7-one，Ⅶ）、二十四亚甲基环阿尔廷醇（24-methylenecycloartanol，Ⅸ）、环桉烯醇（cycloeucalenol，Ⅹ）。抗HBV活性筛选表明，化合物Ⅰ～Ⅲ，Ⅴ～Ⅶ具有明显的抗HBV活性，化合物Ⅰ和Ⅴ在50μg／mL给药剂量时对HBV-DNA克隆转染的人肝癌细胞HepG2-2．2．15的抑制率分别为85．38％和87．37％。结论 化合物Ⅳ～Ⅶ首次从该植物中分得，阿魏酰胺类化合物为首次从该属植物中分得的结构类型，化合物Ⅰ和Ⅴ具有较强的抗HBV活性。

那西肽脂质体的制备及其体外抗乙肝病毒的研究

目的 制备那西肽脂质体并了解其体外抗乙肝病毒作用。方法 以超声法和脱氧胆酸钠法制备那西肽脂质体,同时考察其包封率和粒径大小的影响因素。以HPLC测定那西肽含量,以透射电镜观察其形态,以激光散射法测定其粒径大小,以HepG2 2. 2. 15细胞模型检测那西肽脂质体对乙肝病毒HBsAg和HBeAg的抑制率。结果 采用氯仿甲醇混合液(2∶1,v/v)作溶剂的那西肽脂质体包封率高于二氧六环作溶剂的。随着那西肽与卵磷脂的重量比增大,脂质体包封率呈下降趋势。脱氧胆酸钠法中的脱氧胆酸钠可以调节脂质体粒径的大小,它与卵磷脂的重量比越大脂质体的粒径越小。添加保护剂的脂质体在-20℃放置2年后基本稳定。脂质体中那西肽质量浓度为1 25, 2 5和5 0μg·mL-1时,对HBsAg和HBeAg的抑制率分别达到(46 .9±2. 6)%, (55. 4±1 .2)%, (65±3)%和(15. 1±2 .3)%, (36 .2±1.7)%, (36 .8±2. 5)%。结论 超声法或脱氧胆酸钠法可以制备那西肽脂质体;那西肽脂质体对乙肝病毒HBsAg和HBeAg抑制效果优于游离那西肽。