卤代水杨醛N^-4取代缩氨基硫脲衍生物的合成和生物活性研究

以R—NH2 、CS2 、水合肼、水杨醛等为原料设计合成了 6种未见文献报道的卤代水杨醛缩N4 羟基乙基 /邻甲苯基氨基硫脲 ,其组成与结构由元素分析、红外光谱所证实。体外抑菌试验表明 ,合成的化合物均有较强的抑菌活性 ,说明卤代水杨醛N4 取代缩氨基硫脲是一类新型的有重要研究价值的抗菌剂。

畜禽粪便中磺胺类抗生素及其N^4-乙酰代谢物残留特征的分析

建立了超高效液相色谱-串联质谱(UPLC-MS/MS)法测定畜禽粪便中磺胺类及其N^4-乙酰代谢物的分析方法。鸡粪、猪粪和牛粪样品经加速溶剂萃取仪萃取,Oasis HLB固相萃取柱净化,采用电喷雾正离子(ESI+)多反应检测模式(MRM)定量分析。平均回收率达63.8%~103.7%,相对标准偏差小于13%,方法检出限和定量限分别在0.5~5μg·kg-1和1.5~20μg·kg-1之间。通过对畜禽粪便中磺胺类及其代谢物残留特征的分析表明,畜禽粪便中磺胺类及其代谢物的检出率分别为90%和100%,N^4-乙酰代谢物的残留量约为母体磺胺类抗生素残留量的2倍;不同畜禽粪便样品中其残留量的差异较大,其中猪粪>鸡粪>牛粪。实验结果表明,该方法具有较高的准确度和灵敏度,可满足畜禽粪便中磺胺类抗生素及其代谢物残留的检测要求。

N^4-环丙氨-5-氟-β-L-尿苷的合成

报道了N4 环丙氨 5 氟 β L 尿苷的合成新方法 ,以L 阿拉伯糖为起始原料合成了化合物L 吡喃核糖 ( 6) ,然后经三步反应合成了重要中间体 1 氧 -乙酰基 2 ,3 ,5 三 -氧 -苯甲酰基 β L 呋喃核糖 ( 9) ,与硅烷化的 5 氟尿嘧啶缩合得到了带有保护基的 5 氟 β L 尿苷 ( 10 ) ,脱保护后与六甲基二硅氮烷 (HMDS)、环丙氨、硫酸铵 (催化剂 )一锅反应得到了目标化合物N4 环丙氨 5 氟 β L 尿苷 ( 12 ) .产物结构经MS ,UV ,IR ,NMR ,元素分析等确认 .

新型细胞转染素—N^4-精胺胆固醇羰酰氨的合成

目的 合成新型阳离子脂质体 细胞转染素N4 精氨胆固醇羰酰氨 ,作为基因载体 ,用于肺部疾病的基因治疗。方法 用胆固醇羰酰氯为原料 ,与苄氧羰酰基保护的精氨选择性缩合 ,去保护 ,柱层析分离纯化。结果 以 2 5 .46%收率获得目标化合物 ,由波谱分析证实其结构。结论 建立选择性缩合和分离纯化方法 ,获得高纯度目标化合物 。

N^4-烷基-5-甲基-2′-脱氧胞苷的合成

3′,5′- O-二苯甲酰胸苷在三氯氧磷存在的条件下与 1,2 ,4-三唑缩合可得到 1- (3′,5′- O-二苯甲酰 - β- D-呋喃核糖 ) - 4- (1,2 ,4-三唑 - 1-基 ) - 5 -甲基 -嘧啶 - 2 - (1H) -酮 ,再用不同的胺取代核苷 C4上的三唑基团可制备一系列全新的N4-烷基取代的 5 -甲基胞苷。方法简便 ,收率高。

抗恶性肿瘤新药N^4-戊氧碳酰-5’-脱氧-5-氟胞嘧啶的合成

以去氧氟尿苷、盐酸羟胺、氯甲酸正戊酯等为原料合成N4-戊氧碳酰-5’-脱氧-5-氟胞嘧啶,反应经加成、氢化、缩合、酰胺化等步骤得目标产物。产物经元素分析、IR、1H-NMR、13C-NMR、EI-MS确证。五步合成目标产物N4-戊氧碳酰-5’-脱氧-5-氟胞嘧啶总产率20.05%(以去氧氟尿苷计)。

N^4-[二-(4-氟苯)甲基]哌嗪缩氨基硫脲化合物的合成研究

通过CS20H2N-NH2.H2O,(CH3)2SO4反应制得肼基二硫代甲酸甲酯,肼基二硫代甲酸甲酯再与相应的醛或酮,通过缩合反应制得中间体化合物la^ld,产率80%~95%.以乙醇作溶剂,1与N4-[二-(4-氟苯)甲基]哌嗪发生取代反应制得相应的目标化合物2a^2d,产率67%~84%.共计合成目标化合物4个,均为新化合物.以上新化合物均经熔点、质谱、红外光谱、核磁共振氢谱和元素分析确证.

N^+离子注入对锆-4合金耐腐蚀性能的影响

为了研究N+离子注入对锆-4合金耐腐蚀性能的影响,本文使用直线加速器产生的N+离子注入锆-4合金样品,通过对离子注入后样品电化学曲线的测量,分析不同剂量下N+离子注入对锆-4合金钝化电流密度的影响,同时使用透射电子显微镜分析注入层的微观结构。结果表明,随着注入剂量的提高(0—1×1016cm-2),样品钝化电流密度下降,耐腐蚀性能提高,其原因主要归结于样品表层由多晶结构到非晶结构的转变过程。

高效液相色谱法测定小鼠血清中磺胺间甲氧嘧啶及其N^4-乙酰化代谢物含量

磺胺间甲氧嘧啶(SMM)于ICR小鼠哺乳期暴露后,测定其血清中SMM和N4-乙酰化磺胺间甲氧嘧啶(ACSMM)含量。小鼠哺乳期以0、10、50、200 mg/(kg·d)灌胃给药直至21 d,于第22天麻醉,摘眼球取血。血清样品经乙腈沉淀蛋白,超声辅助提取,采用反相C18(250 mm×4.6 mm,5μm)色谱柱,以乙腈:水(含0.2%冰乙酸)为流动相,流速为1 mL/min,柱温35℃,高效液相色谱耦合二极管阵列检测器对目标物检测,SMM和ACSMM检测波长分别为273 nm和267 nm,外标法峰面积定量。SMM和ACSMM血清加标回收率为85.8%~110.2%,精密度(RSD%)均小于8.3%,在0.05~2.5μg/mL和5~30μg/mL(适用母鼠体内SMM)范围内与峰面积呈良好的线性关系,r2>0.990;最低检出限分别为0.023和0.042μg/mL。本法灵敏、快速、准确,可用于血清中SMM和ACSMM的测定。

Targeting N4-acetylcytidine suppresses hepatocellular carcinoma progression by repressing eEF2-mediated HMGB2 mRNA translation

Background:N4-acetylcytidine(ac4C)represents a novel messenger RNA(mRNA)modification,and its associated acetyltransferaseN-acetyltransferase 10(NAT10)plays a crucial role in the initiation and progression of tumors by regulating mRNA functionality.However,its role in hepatocellular carcinoma(HCC)development and prognosis is largely unknown.This study aimed to elucidate the role of NAT10-mediated ac4C in HCC progression and provide a promising therapeutic approach.Methods:The ac4C levels were evaluated by dot blot and ultra-performance liquid chromatography-tandem mass spectrometry with harvested HCC tissues.The expression of NAT10 was investigated using quantitative real-time polymerase chain reaction,western blotting,and immunohistochemical staining across 91 cohorts of HCC patients.To explore the underlying mechanisms of NAT10-ac4C in HCC,we employed a comprehensive approach integrating acetylated RNA immunoprecipitation and sequencing,RNA sequencing and ribosome profiling analyses,along with RNA immunoprecipitation,RNA pulldown,mass spectrometry,and site-specific mutation analyses.The drug affinity responsive targets stability,cellular thermal shift assay,and surface plasmon resonance assays were performed to assess the specific binding of NAT10 and Panobinostat.Furthermore,the efficacy of targeting NAT10-ac4C for HCC treatment was elucidated through in vitro experiments using HCC cells and in vivo HCCmouse models.Results:Our investigation revealed a significant increase in both the ac4C RNA level and NAT10 expression in HCC.Notably,elevated NAT10 expression was associated with poor outcomes in HCC patients.Functionally,silencing NAT10 suppressed HCC proliferation and metastasis in vitro and in vivo.Mechanistically,NAT10 stimulates the ac4C modification within the coding sequence(CDS)of high mobility group protein B2(HMGB2),which subsequently enhances HMGB2 translation by facilitating eukaryotic elongation factor 2(eEF2)binding to the ac4C sites on HMGB2 mRNA’s CDS.Additionally,high-throughput compound library screening revealed Panobinostat as a potent inhibitor of NAT10-mediated ac4C modification.This inhibition significantly attenuated HCC growth and metastasis in both in vitro experiments using HCC cells and in vivo HCC mouse models.Conclusions:Our study identified a novel oncogenic epi-transcriptome axis involving NAT10-ac4C/eEF2-HMGB2,which plays a pivotal role in regulating HCC growth and metastasis.The drug Panobinostat validates the therapeutic potential of targeting this axis for HCC treatment.

DNA4mcEL:基于核苷酸信息特征计算分析与预测DNA N^4-甲基胞嘧啶位点

N^4-甲基胞嘧啶(N^4-methylcytosine,4mC)是一种重要的表观遗传修饰,在DNA的修复、表达和复制中发挥重要作用。准确鉴定4mC位点有助于深入研究其生物学功能和机制,由于4mC位点的实验鉴定即耗时又昂贵,特别是考虑到基因序列的快速积累,迫切需要补充有效的计算方法。因此,提供一个快速、准确的4mC位点在线预测平台十分必要。目前,还未见对构建必要的预测模型所需的不同特征的机器学习(machine learning,ML)方法进行全面的分析和评估。我们构建多组特征集,并且采用5种ML方法 (如随机森林,支持向量机,集成学习等),提出一种称为'DNA4mcEL'的预测方法。在随机10折交叉验证测试下与现有的预测器相比,DNA4mcEL预测C. elegans、D. melanogaster、A. thaliana、E. coli、G. subterraneus、G. pickeringii 6个物种的精度均有提高。基于本方法的预测器DNA4mcEL在这项任务中显著优于现有的预测器。我们希望通过这个综合调查和建立更准确模型的策略,可以作为激发N^4-甲基胞嘧啶预测计算方法未来发展的有用指南,加快新N^4-甲基胞嘧啶的发现。DNA4mcEL的独立版本可以从https://github.com/kukuky00/DNA4mcEL.git免费获得。

Theoretical calculation of photoionization cross sections of B-like ions:N^(2+),O^(3+) and F^(4+)

There can be found some notable discrepancies with regard to the resonance structures when R-matrix calculations from the Opacity Project and other sources are compared with recent absolute experimental measurements of Bizau et al [Astron. Astrophts. 439 387 （2005）] for B-like ions N^2＋, O^3＋ and F^4＋. We performed close-coupling calculations based on the R-matrix formalism for the photoionizations of ions mentioned above both for the ground states and first excited states in the near threshold regions. The present results are compared with experimental ones given by Bizau et al and earlier theoretical ones. Excellent agreement is obtained between our theoretical results and the experimental photoionization cross sections. The present calculations show a significant improvement over the previous theoretical results.

Engagement of N^(6)-methyladenisine methylation of Gng4 mRNA in astrocyte dysfunction regulated by CircHECW2

The N^(6)-methyladenosine(m^(6)A)modification is the most prevalent modification of eukaryotic mRNAs and plays a crucial role in various physiological processes by regulating the stability or function of target mRNAs.Accumulating evidence has suggested that m6A methylation may be involved in the pathological process of major depressive disorder(MDD),a common neuropsychiatric disorder with an unclear aetiology.Here,we found that the levels of the circular RNA HECW2(circHECW2)were significantly increased in the plasma of both MDD patients and the chronic unpredictable stress(CUS)mouse model.Notably,the downregulation of circHECW2 attenuated astrocyte dysfunction and depression-like behaviors induced by CUS.Furthermore,we demonstrated that the downregulation of circHECW2 increased the expression of the methylase WTAP,leading to an increase in Gng4 expression via m^(6)A modifications.Our findings provide functional insight into the correlation between circHECW2 and m^(6)A methylation,suggesting that circHECW2 may represent a potential target for MDD treatment.

Ⅴ.N^4-[5′-(3′-甲硫基-6′-甲基)-1′,2′,4′-三嗪基]-3-甲硫基-5-氧代-6-甲基-4,5-二氢-1,2,4-三嗪的胺解反应

N^4-[5′-(3′-甲硫基-6′-甲基)-1′,2′,4′-三嗪基-3-甲硫基-5-氧代-6-甲基-4,5-二氢-1,2,4-三嗪(Ⅱ)在乙醇中与一系列胺反应,得3-甲硫基-5-取代胺基-6-甲基-1,2,4-三嗪(Ⅲ_(a-h))及3-甲硫基-5-羟基-6-甲基-1,2,4-三嗪(Ⅰ);Ⅱ在同样条件下,与碱性弱或空间障碍较大的对硝基苯胺、邻氯苯胺及邻甲苯胺不发生反应;Ⅱ与氨,异丁胺反应,尚得到3-取代胺基-5-羟基-6-甲基-1,2,4-三嗪(Ⅳ_(a,b)).

Cu-ATSM保护内皮细胞糖氧剥夺再灌注损伤的机制

目的:探讨铜-二乙酰-2(N4-甲基氨基硫脲)(Cu-ATSM)对糖氧剥夺再灌注(OGD/R)后人脐静脉内皮细胞(HUVECs)凋亡的作用及其潜在机制。方法:将体外培养的HUVECs分为Control组、OGD/R组、OGD/R+Cu-ATSM组;采用流式细胞术检测HUVECs的凋亡情况;采用Mito-tracker染色检测线粒体形态;JC-1染色分析各组线粒体膜电位的变化;采用蛋白质印迹法(Western blot)检测各组自噬相关蛋白如ATG5、Beclin1、LC3Ⅰ/Ⅱ的激活情况;采用免疫荧光检测各组细胞LC3Ⅱ的表达情况。结果:流式细胞术结果显示,与Control组比,OGD/R损伤后HUVECs凋亡数目显著上升,而当Cu-ATSM治疗后,凋亡的HUVECs显著减少(均P<0.05)。Mito-tracker及JC-1染色结果显示,与Control组比,OGD/R组损伤后线粒体的形态明显受到损伤,线粒体功能受损,膜电位下降,而OGD/R+Cu-ATSM组治疗后能够改善损伤的线粒体功能,维持线粒体形态和线粒体膜电位(均P<0.05)。Western blot结果显示,与Control组比,OGD/R组损伤后自噬相关信号通路如ATG5、Beclin1、LC3Ⅱ蛋白的表达量增多,而OGD/R+Cu-ATSM组治疗后的自噬相关蛋白表达量较OGD/R组下降(均P<0.05)。免疫荧光结果显示,与Control组比,OGD/R组损伤后LC3Ⅱ的表达显著增多,而OGD/R+Cu-ATSM治疗后的LC3Ⅱ显著减少。结论:Cu-ATSM能够改善HUVECs OGD/R损伤后的线粒体功能,减少HUVECs凋亡,其机制可能是通过抑制自噬实现的。

Transcriptome-wide profiling of RNA N^(4)-cytidine acetylation in Arabidopsis thaliana and Oryza sativa

Acetylation of N^(4)-cytidine(ac^(4)C)has recently been discovered as a novel modification of mRNA.RNA ac^(4)C modification has been shown to be a key regulator of RNA stability,RNA translation,and the thermal stress response.However,its existence in eukaryotic mRNAs is still controversial.In plants,the existence,distribution pattern,and potential function of RNA ac^(4)C modification are largely unknown.Here we report the presence of ac^(4)C in the mRNAs of both Arabidopsis thaliana and rice(Oryza sativa).By comparing two ac^(4)C sequencing methods,we found that RNA immunoprecipitation and sequencing(acRIP-seq),but not ac^(4)C sequencing,was suitable for plant RNA ac^(4)C sequencing.We present transcriptome-wide atlases of RNA ac^(4)C modification in A.thaliana and rice mRNAs obtained by acRIP-seq.Analysis of the distribution of RNA ac^(4)C modifications showed that ac^(4)C is enriched near translation start sites in rice mRNAs and near translation start sites and translation end sites in Arabidopsis mRNAs.The RNA ac^(4)C modification level is positively correlated with RNA half-life and the number of splicing variants.Similar to that in mammals,the translation efficiency of ac^(4)C target genes is significantly higher than that of other genes.Our in vitro translation results confirmed that RNA ac^(4)C modification enhances translation efficiency.We also found that RNA ac^(4)C modification is negatively correlated with RNA structure.These results suggest that ac^(4)C is a conserved mRNA modification in plants that contributes to RNA stability,splicing,translation,and secondary structure formation.

NAT10-mediated ac 4 C-modified ANKZF1 promotes tumor progression and lymphangiogenesis in clear-cell renal cell carcinoma by attenuating YWHAE-driven cytoplasmic retention of YAP1

Background:Lymphatic metastasis is one of the most common metastatic routes and indicates a poor prognosis in clear-cell renal cell carcinoma(ccRCC).N-acetyltransferase 10(NAT10)is known to catalyze N4-acetylcytidine(ac4C)modification of mRNA and participate in many cellular processes.However,its role in the lymphangiogenic process of ccRCC has not been reported.This study aimed to elucidate the role of NAT10 in ccRCC lymphangiogenesis,providing valuable insights into potential therapeutic targets for intervention.Methods:ac4C modification and NAT10 expression levels in ccRCC were assessed using public databases and clinical samples.Functional investigations involved manipulating NAT10 expression in cellular and mouse models to study its role in ccRCC.Mechanistic insights were gained through a combination of RNA sequencing,mass spectrometry,co-immunoprecipitation,RNA immuno-precipitation,immunofluorescence,and site-specific mutation analyses.Results:We found that ac4C modification and NAT10 expression levels increased in ccRCC.NAT10 promoted tumor progression and lymphangiogene-sis of ccRCC by enhancing the nuclear import of Yes1-associated transcriptional regulator(YAP1).Subsequently,we identified ankyrin repeat and zinc fin-ger peptidyl tRNA hydrolase 1(ANKZF1)as the functional target of NAT10,and its upregulation in ccRCC was caused by NAT10-mediated ac4C modifi-cation.Mechanistic analyses demonstrated that ANKZF1 interacted with tyro-sine 3-monooxygenase/tryptophan 5-monooxygenase activation protein epsilon(YWHAE)to competitively inhibit cytoplasmic retention of YAP1,leading to transcriptional activation of pro-lymphangiogenic factors.Conclusions:These results suggested a pro-cancer role of NAT10-mediated acetylation in ccRCC and identified the NAT10/ANKZF1/YAP1 axis as an under-reported pathway involving tumor progression and lymphangiogenesis in ccRCC.

新型阳离子脂质体的制备及其转染效率的测定

目的 制备以新型细胞转染素为阳性成分的阳离子脂质体 ,作为基因载体 ,用于基因转染的实验研究和肺部疾病的基因治疗。方法 用薄膜法制备N4 精胺胆固醇羰酰氨阳离子脂质体 ,通过电镜观察和电泳 ,检测其形态、结构和细胞转染条件 ,并对SPC肿瘤细胞进行转染 ,评估其转染效率。结果 制备的新型阳离子脂质体具有良好的微囊形态 ,当脂质体∶质粒为 (6～ 8)∶1时 ,其转染效率可达 60 %。结论 成功制备了新型阳离子脂质体 ,在适当条件下其具有较高的转染活性 ,为以后的基因转染和基因治疗打下基础。

磺胺甲噁唑及其乙酰化代谢物的药代动力学研究

目的；测定２０名中国男性健康受试者口服复方新诺明片（含磺胺甲噁唑８００ｍｇ）后磺胺甲噁唑（ＳＭＺ）和Ｎ４－乙酰基－磺胺甲噁唑（Ｎ４－Ａｃ－ＳＭＺ）的血药物浓度，进行药代动力学研究。方法：用反相高效液相色谱法同时测定磺胺甲噁唑和Ｎ４－乙酰基－磺胺甲噁唑的血浆浓度，并计算药代动力学参数。结果：磺胺甲噁唑和Ｎ４－乙酰基－磺胺甲噁唑的ｔｍаｘ分别为１．８３±０．７１ｈ和８．１０ ± ２．９４ｈ，Ｃｍаｘ分别为 ３７．５７±１３．５２ｍｇ·Ｌ－１和５．８９±２．９０ｍｇ·Ｌ－１，ｔ１／２分别为１０．２５±１．１６ ｈ和 １２．６５±１．４４ ｈ，ＡＵＣ０－３４分别为４８６．０±１８３．６ ｍｇ·ｈ·Ｌ－１和１３１．７±６４．１ｍｇ·ｈ·Ｌ－１，ＡＵＣ０－８分别为５６２．６±１９６．７ｍｇ·ｈ·Ｌ－１和１６２．２±８２．９ｍｇ·ｈ·Ｌ－１，ＭＲＴ分别为１５．８±１．５ｈ和２１．９±１．９ｈ。结论：磺胺甲■唑的乙酰化代谢途径为生成限速代谢模式，２０人中没有发现明显的快慢代谢分型。

高效液相色谱法测定Cu(ATSM)混悬剂的含量

目的建立测定Cu(ATSM)混悬剂中Cu(ATSM)含量的高效液相色谱法.方法采用RP-HPLC法,用ODS柱,乙腈-水(含0.1%三氟乙酸)的梯度洗脱系统,检测波长为305 nm.结果 Cu(ATSM)在浓度为5~100!g·m L^(-1)范围内线性关系良好(r=0.9967),检测限为2 ng(S/N=3),加样回收率为103.0%,RSD为1.98%(n=6).结论所建方法简便,灵敏度高,准确度好,专属性强,可为Cu(ATSM)制剂的质量评价提供可行的方法参考.