胞核嘧啶氢键二聚体的电子拓扑性质研究

采用GAUSSIAN程序包,在B3LYP/6-311++G**水平下,对胞核嘧啶及其7种氢键二聚体进行了结构优化和能量分析。使用AIM2000程序,在分子中的原子理论框架下对这些体系进行了详尽的电子拓扑性质的理论研究。从理论的角度证明了这些氢键的存在并探讨了其性质,比较了各种不同类型的氢键及二聚体的稳定性,为实验研究提供了一定的理论参考数据。

含腙结构的农药及其开发方向

主要介绍了目前已商品化的含腙类结构农药,以及20世纪90年代以来世界各大公司研制的具农药活性的腙类化合物。

骨骼肌P2受体的功能与疾病

嘌呤和嘧啶核苷酸作用的P2受体分为G蛋白偶联型的P2Y受体和配体门控离子通道型的P2X受体。研究表明,骨骼肌细胞有多种P2X和P2Y受体亚型表达。嘌呤信号在肌肉萎缩症的肌肉再生中具有重要的作用,与肌肉萎缩症、肌腱炎等骨骼肌疾病有关,并可能为治疗与肌肉有关的疾病提供新的治疗策略。

Eucaryotic DNA Methylation and Gene Mutation

5-methylcytosine (m5C) as a rare base exists in eucaryotic genomes, it is a normal constituent of many eucaryotic DNA, whose existence is a character of eucaryotic DNA. In the regular physiological conditions, cytosine residue of eucaryotic DNA is methylated to be popular. Up to the present, many people consider that the m5C may be mutation hotspots by the m5C deamination leading to gene mutation. Our theoretical investigations indicated that the spontaneous mutation caused by the transition of G - C-A - T, in eukaryotic DNA, may be a result caused by the tautomer changing base pairs and may also be caused by other factor actions, however it could not be caused by the deamination of m5C.

M-RNA碱基复合物(M=Ca^+和Mg^+)特征振动的密度泛函理论分析

采用密度函数理论的B3LYP交换关联能泛函在6-311+G(2df,2p)基组水平上,优化计算了Ca+,Mg+与RNA碱基嘧啶各同分异构体形成稳定复合物的结构,发现其中C1M,T1M和U1M(M=Ca+和Mg+)为最稳定复合物,并对这些复合物红外振动进行了计算。计算结果显示碱基嘧啶单体主要存在的两个红外特征振动,是由环振动和环外氧原子及与其成键的环上碳原子之间产生伸缩振动引起的。当形成复合物时,由于离子的参与,使单体碱基分子的振动情况发生了改变,主要表现在离子倾向于与环外负电子原子N和O原子成键,使与离子直接作用环外原子参与的特征振动频率减小,谱线红移;不与离子直接作用环外原子引起的特征振动频率增大,谱线蓝移。

Antineoplastic Effect of Calcium Channel Blocker-Verapamil and 5-Fluorouracil Intraperitoneal Chemotherapy on Hepatocarcinoma-Bearing Rats

Objective To study the antineoplastic effect of the calcium channel blocker verapamil and 5-fluorouracil intraperitoneal chemotherapy onhepatocarcinoma-bearing rats, and examine the action between calcium channel blockers and cytotoxic drugs.Methods We adopted the method of subcapsular implantation of carcinoma tissues of walker-256 in the left liver lobe as a model of livercarcinoma-bearing rats. All experimental animals were divided into four groups. On the sixth day post implantation, in group A (controlgroup) 6 ml of saline was injected intraperitoneally once a day for 3 days. In group B (single chemotherapy group) 6 ml of 5-Fu 75 mg/kg was injected intraperitoneally once a day for 3 days. In group C (combination of treatment group) both 5-Fu (75 mg/kg) and verapamil(25 mg/kg) were administered simultaneously as in A and B. In group D (simple verapamil group) only 6 ml of verapamil (25 mg/kg)was administered as above.Results Compared with groups A, B and D, The volume of cancer and the contents of liver cancer DNA and protein were significantlyreduced. The rates of inhibiting cancer (89.9% in group C and 35.4% in group B) were significantly increased in group C. Group C hadsignificantly long survival time compared to groups A, B and D ( P < 0.05) . By light microscopy, a number of focal necroses were foundin cancer tissue in group C.Conclusion Calcium channel blockers can enhance the antineoplastic effect of 5-Fu intraperitoneal chemotherapy to liver cancer ; Theuse of verapamil can not increase the toxicity of 5-Fu.

Synthesis of Aminoglucose Conjugates of 5-Fluorouracil-1-acetic Acid and 5-Fluorouracil-1-propanoic Acid and Their Antitumor Activities

Six aminoglucose conjugates were synthesized by the reaction of aminoglucose with 5-fluorou-racil-1-acetic acid or 5-fluorouracil-1-propanoic acid and confirmed by IR, 1H NMR and elemental analyses. Their antitumor activities against A2780 cells and PC-14 cells were also evaluated.

Specific CEA-producing colorectal carcinoma cell killing with recombinant adenoviral vector containing cytosine deaminase gene

AIM: To kill CEA positive colorectal carcinoma cells specifically using the E coli cytosine deaminase (CD) suicide gene, a new replication-deficient recombinant adenoviral vector was constructed in which CD gene was controlled under CEA promoter and its in vitro cytotoxic effects were evaluated. METHODS: Shuttle plasmid containing CD gene and regulatory sequence of the CEA gene was constructed and recombined with the right arm of adenovirus genome DNA in 293 cell strain. Dot blotting and PCR were used to identify positive plaques. The purification of adenovirus was performed with ultra-concentration in CsCl step gradients and the titration was measured with plaque formation assay. Cytotoxic effects were assayed with MTT method, The fifty percent inhibition concentration (IC(50)) of 5-FC was calculated using a curve-fitting parameter. The human colorectal carcinoma cell line, which was CEA-producing, and the CEA-nonproducing Hela cell line were applied in cytological tests. An established recombinant adenovirus vector AdCMVCD, in which the CD gene was controlled under CMV promoter, was used as virus control. Quantitative results were expressed as the mean +/- SD of the mean. Statistical analysis was performed using ANOVA test. RESULTS: The desired recombinant adenovirus vector was named AdCEACD. The results of dot blotting and PCR showed that the recombinant adenovirus contained CEA promoter and CD gene. Virus titer was about 5.0 X 10(14)pfu/L(-1) after purification. The CEA-producing Lovo cells were sensitive to 5-FC and had the same cytotoxic effect after infection with AdCEACD and AdCMVCD (The IC(50) values of 5-FC in parent Lovo cells, Lovo cells infected with 100 M.O.I AdCEACD and Lovo cells infected with 10 M.O.I AdCMVCD were 】15000, 216.5+/-38.1 and 128.8+/-25.4 micromol.L(-1), P【0.001, respectively), and the cytotoxicity of 5-FC increased accordingly when the m.o.i of adenoviruses were enhanced (The value of IC(50) of 5-FC was reduced to 27.9+/-4.2 micromol.L(-1) in 1000 M.O.I AdCEACD infected Lovo cells and 24.8+/-7.1 micromol.L(-1) in 100 M.O.I AdCMVCD infected Lovo cells, P【0.05, P【0.01, respectively). The CEA-nonproducing Hela cells had no effect after infection with AdCEACD, but Hela cells had the cytotoxic sensitivity to 5-FC after infection with AdCMVCD (The IC(50) of 5-FC in parent Hele cells and Hela cells infected with AdCMVCD at 10 M.O.I was 】15000 and 214.5+/-31.3 micromol.L(-1), P【0.001). AdCEACD/5-FC system also had bystander effect, and the viability was about 30 percent when the proportion of transfected cells was only 10 percent. CONCLUSION: The recombinant adenovirus vector AdCEACD has the character of cell type-specific gene delivery. The AdCEACD/5-FC system may become a new, potent and specific approach for the gene therapy of CEA-positive neoplasms, especially colon carcinoma.

Systemic gemcitabine combined with intra-arterial low-dose cisplatin and 5-fluorouracil for advanced hepatocellular carcinoma: Seven cases

The combination of intra-arterial low-dose cisplatin and 5-fluorouracil (5-FU) is effective against advanced hepatocellular carcinoma (HCC). Systemic gemcitabine chemotherapy seems effective in many cancers. We report the results of combination therapy with systemic gemcitabine, intra-arterial low-dose cisplatin and 5-FU (GEMFP). Seven patients with non-resectable advanced HCC were treated with GEMFP. One course of chemotherapy consisted of daily intra-arterial cisplatin (20 mg/body weight/hour on d 1, 10 mg/body weight per 0.5 h on d 2-5 and 8-12), followed by 5-FU (250 mg/body weight per 5 h on d 1-5 and 8-12) via an injection port. Gemcitabine at 1000 mg/m2 was administered intravenously at 0.5 h on d 1 and 8. The objective response was 57%. The response to GEMFP was as follows: complete response (no patients), partial response (four patients), stable disease (three patients), and progressive disease (no patients). The median survival period was 8 mo (range, 5-55). With regard to the National Cancer Institute Common Toxicity Criteria (NCI-CTC) grade 3 or 4 adverse reactions, seven (100%), seven, six (86%) and one (14%) patients developed leukopenia, neutropenia, thrombocytopenia and anemia, respectively. GEMFP may potentially be effective for non- resectable advanced HCC, but it has severe hematologic toxicity.

Induction of nucleoside phosphorylase in Enterobacter aerogenes and enzymatic synthesis of adenine arabinoside

Nucleoside phosphorylases (NPases) were found to be induced in Enterobacter aerogenes DGO-04, and cytidine and cytidine 5′-monophosphate (CMP) were the best inducers. Five mmol/L to fifteen mmol/L cytidine or CMP could distinctly increase the activities of purine nucleoside phosphorylase (PNPase), uridine phosphorylase (UPase) and thymidine phosphorylase (TPase) when they were added into medium from 0 to 8 h. In the process of enzymatic synthesis of adenine arabinoside from adenine and uracil arabinoside with wet cells of Enterobacter aerogenes DGO-04 induced by cytidine or CMP, the reaction time could be shortened from 36 to 6 h. After enzymatic reaction the activity of NPase in the cells induced remained higher than that in the cells uninduced.

Medium optimization for enhanced production of cytosine-substituted mildiomycin analogue (MIL-C) by Streptoverticillium rimofaciens ZJU 5119

Cytosine-substituted mildiomycin analogue （MIL-C） was produced effectively by supplementing cytosine into the culture of Streptoverticillium rimofaciens. In order to improve the yield of MIL-C, statistically-based experimental designs were applied to optimize the fermentation medium for S. rimofaciens ZJU 5119. Fifteen culture conditions were examined for their significances on MIL-C production using Plackett-Burman design. The Plackett-Burrnan design and one-variable-at-a-time design indicated that glucose and rice meal as the complex carbon sources, and peanut cake meal and NH4NO3 as the complex nitrogen sources were beneficial for MIL-C production in S. rimofaciens ZJU 5119. The results of further central composition design （CCD） showed that the optimal concentration of glucose, rice meal and peanut cake meal were 18.7 g/L, 64.8 g/L and 65.1 g/L, respectively. By using this optimal fermentation medium, the MIL-C concentration was increased up to 1336.5 mg/L, an approximate 3.8-fold improvement over the previous concentration （350.0 mg/L） with un-optimized medium. This work will be very helpful to the large-scale production of MIL-C in the future.

Re-evaluation of antitumor effects of combination chemotherapy with interferon-α and 5-fluorouracil for advanced hepatocellular carcinoma

AIM： To evaluate the efficacy of combination chemotherapy with interferon-α （IFNα） and 5-fluorouracil （5-FU） in patients with advanced hepatocellular carcinoma （HCC）. METHODS： Twenty-eight HCC patients in advanced stage were enrolled in the study. They were treated with IFNα/ 5-FU combination chemotherapy. One cycle of therapy lasted for 4 wk. IFNα （3×10^6 units） was subcutaneously injected thrice weekly on days 1, 3, and 5 for 3 wk, and 5-FU （500 mg/d） was administered via the proper hepatic artery for 5 consecutive days per week for 3 wk. No drugs were administered during the 4th wk. The effect of combination chemotherapy was evaluated in each patient alter every cycle based on the reduction of tumor volume. RESULTS： Alter the 1^st cycle of therapy, 16 patients showed a partial response （PR, 57.1%） but none showed a complete response （CR, 0%）. At the end of therapy, the number of patients who showed a CR, PR, or no response （NR） was 1, 10, and 17, respectively. The response rate for therapy （CR＋PR） was 21.5%. Biochemical tests before therapy were compared between responsive （CR＋PR） and non-responsive （NR） patients, but no significant differences were found for any of the parameters examined, indicating that no reasonable predictors could be identified in our analysis. CONCLUSION： Attempts should be made to discriminate between responders and non-responders by evaluating tumor size alter the first cycle of IFNα/5-FU combination chemotherapy. For non-responders, therapy should not proceed to the next cycle, and instead, different combination of anticancer drugs should be explored. 2005 The WJG Press and Elsevier Inc. All rights reserved

Methyl-CpG binding proteins in the nervous system

Classical methyl-CpG binding proteins contain the conserved DNA binding motif methyl-cytosine binding domain(MBD), which preferentially binds to methylated CpG dinucleotides. These proteins serve as transcriptional repressors,mediating gene silencing via DNA cytosine methylation. Mutations in methyl-CpG binding protein 2 (MeCP2) have beenlinked to the human mental retardation disorder Rett syndrome, suggesting an important role for methyl-CpG bindingproteins in brain development and function. This mini-review summarizes the recent advances in studying the diversefunctions of MeCP2 as a prototype for other methyl-CpG binding proteins in the development and function of thevertebrate nervous system.

Inhibition of human gastric carcinoma cell growth by atofluding derivative N_3-o-toluyl-fluorouracil

AIM： To evaluate the growth inhibition efficacy of atofluding derivative N3-o-toluyl-fluorouracil （TFU） on human gastric carcinoma cell lines SGC-7901 and MKN-45. METHODS： Cell growth inhibition by TFU was measured by MTT and clonogenic assays without or with liver microsomal enzymes. Xenografts of cancer cells in nude mice were employed to study the anti-proliferative effects of TFU in vivo. RESULTS： TFU inhibited the growth of SGC-7901 and MKN-45 cells. However, the inhibitory effects of TFU on cell growth were not significant. The inhibition rates were enhanced in the presence of liver microsomal enzymes, ranging 4.73%-48.57% in SGC-7901 cells and 9.0%-62.02% in MKN-45 cells. In v/vo, TFU delayed the growth of SGC-7901 and MKN-45 cells in nude mice. The inhibition rates were 40.49%, 63.24%, and 75.98% in SGC-7901 cells and 40.76%, 61.41%, and 82.07% in MKN-45 cells when the oral doses were 25, 50, and 100 mg/kg, respectively. TFU treatment was generally well tolerated by mice with less than 20% reduction in body weight. CONCLUSION： TFU inhibits the growth of human gastric carcinoma cells. The inhibition rates are increased in the presence of liver microsomal enzymes. The efficacy of TFU may be associated with the sustaining release of 5-fluorouracil （5-FU） mediated by the enzymes.

Spindle cell carcinoma of the breast as complex cystic lesion: a case report

Spindle cell carcinoma of the breast is a rare tumor. This tumor can proliferate rapidly and cause cystic changes because of internal tissue necrosis. We evaluated a 54-year-old woman with right breast lump. Mammography showed a category four mass with a diameter of 2.5 cm. Ultrasonography(US) revealed a complex cystic lesion, and fine-needle aspiration(FNA) cytology demonstrated bloody fluid and malignant cells. Partial breast resection and sentinel lymph node biopsy were performed. Immunohistology revealed spindle cells with positive results for cytokeratin(AE1/AE3) and vimentin, partially positive results for s-100, and negative results for desmin and α-actin. The pathological stage was IIA, and biochemical characterization showed that the tumor was triple negative. Six courses of FEC-100 chemotherapy(5-fluorouracil 500 mg/m2, epirubicin 100 mg/m2, and cyclophosphamide 500 mg/m2) were administered. Radiotherapy was performed. This case is discussed with reference to the literature.

Effect of targeted magnetic nanoparticles containing 5-FU on expression of bcl-2, bax and caspase 3 in nude mice with transplanted human liver cancer

AIM: To investigate the anti-tumor effect and mechanisms of magnetic nanoparticles targeting hepatocellular carcinoma. METHODS: Human hepatocellular carcinoma was induced in nude mice, and the mice were randomly divided into group A receiving normal saline, group B receiving magnetic nanoparticles containing 5-fluorouracil (5-FU), group C receiving 5-FU, and group D receiving magnetic nanoparticles containing 5-FU with a magnetic field built in tumor tissues. The tumor volume was measured on the day before treatment and 1, 4, 7, 10 and 13 d after treatment. Tumor tissues were isolated for examination of the expression of bcl-2, bax and caspase 3 by immunohistochemical method, reverse transcription polymerase chain reaction and Western blotting. RESULTS: The tumor volume was markedly lower in groups C and D than in groups A and B (group C or D vs group A or B, P < 0.01). The volume was markedly lower in group D than in group C (P < 0.05). The expression of protein and mRNA of bcl-2 was markedly lower in groups C and D than in groups A and B (group C or D vs group A or B, P < 0.01), and was markedly lower in group D than in group C (P < 0.01). The expression of bax and caspase 3 in groups C and D was signif icantly increased, compared with that in groups A and B (P < 0.01). CONCLUSION: The targeted magnetic nanoparticles containing 5-FU can improve the chemotherapeutic effect of 5-FU against hepatocellular carcinoma by decreasing the expression of bcl-2 gene, and increasing the expression of bax and caspase 3 genes.

Rosiglitazone enhances fluorouracil-induced apoptosis of HT-29 cells by activating peroxisome proliferator-activated receptor γ

AIM： To examine whether and how rosiglitazone enhances apoptosis induced by fluorouracil in human colon cancer （HT-29） cells. METHODS： Human colon cancer HT-29 cells were cultured in vitro and treated with fluorouracil and/or rosiglitazone. Proliferation and growth of HT-29 cells were evaluated by MTF assay and trypan blue exclusion methods, respectively. The apoptosis of HT-29 cells was determined by acridine orange/ethidium bromide staining and flow cytometry using PI fluorescence staining. The expressions of peroxisome proliferator-activated receptor γ （PPARγ）, Bcl-2 and Bax in HT-29 cells were analyzed by Western blot. RESULTS： Although rosiglitazone at the concentration below 30 μmol/L for 72 h exerted almost no inhibitory effect on proliferation and growth of HT-29 cells, it could significantly enhance fluorouracil-induced HT-29 cell proliferation and growth inhibition. Furthermore, 10 μmol/L rosilitazone did not induce apoptosis of HT-29 cells but dramatically enhanced fluorouracil-induced apoptosis of HT-29 cells. However, rosiglitazone did not improve apoptosis induced by fluorouracil in HT-29 cells pretreated with GW9662, a PPARγ antagonist. Meanwhile, the expression of Bax and PPAR7 was upregulated, while the expression of Bcl-2 was down regulated in HT-29 cells treated with rosiglitazone in a time-dependent manner. However, the effect of rosiglitazone on Bcl-2 and Bax was blocked or diminished in the presence of GW9662. CONCLUSION： Rosiglitazone enhances fluorouracilinduced apoptosis of HT-29 cells by activating PPARγ.

N-terminal and C-terminal cytosine deaminase domain of APOBEC3G inhibit hepatitis B virus replication

AIM: To investigate the effect of human apolipoprotein B mRNA-editing enzyme catalytic-polypeptide 3G (APOBEC3G) and its N-terminal or C-terminal cytosine deaminase domain-mediated antiviral activity against hepatitis B virus (HBV) in vitro and in vivo. METHODS: The mammalian hepatoma cells HepG2 and HuH7 were cotransfected with APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain expression vector and 1.3-fold-overlength HBV DNA as well as the linear monomeric HBV of genotype B and C. For in vivo study, an HBV vector-based mouse model was used in which APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain expression vectors were co-delivered with 1.3-fold-overlength HBV DNA via high-volume tail vein injection. Levels of hepatitis B virus surface antigen (HBsAg) and hepatitis B virus e antigen (HBeAg) in the media of the transfected cells and in the sera of mice were determined by ELISA.The expression of hepatitis B virus core antigen (HBcAg) in the transfected cells was determined by Western blot analysis. Core-associated HBV DNA was examined by Southern blot analysis. Levels of HBV DNA in the sera of mice as well as HBV core-associated RNA in the liver of mice were determined by quantitative PCR and quantitative RT-PCR analysis, respectively. RESULTS: Human APOBEC3G exerted an anti-HBV activity in a dose-dependent manner in HepG2 cells, and comparable suppressive effects were observed on genotype B and C as that of genotype A. Interestingly, the N-terminal or C-terminal cytosine deaminase domain alone could also inhibit HBV replication in HepG2 cells as well as Huh7 cells. Consistent with in vitro results, the levels of HBsAg in the sera of mice were dramatically decreased, with more than 50 times decrease in the levels of serum HBV DNA and core-associated RNA in the liver of mice treated with APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain as compared to the controls. CONCLUSION: Our findings provide probably the first evidence showing that APOBEC3G and its N-terminal or C-terminal cytosine deaminase domain could suppress HBV replication in vitro and in vivo.

Activity and safety of pegylated liposomal doxorubicin,5-fluorouracil and folinic acid in inoperable hepatocellular carcinoma:A phase Ⅱ study

AIM: To improve the results of New therapeutic strategies in hepatocellular carcinoma (HCC). We have conducted a phase Ⅱ study with pegylated liposomal doxorubicin (PLD), 5-fluorouracil (5FU) and folinic acid (FA). METHODS: Thirty-one patients with hystologically- confirmed, inoperable HCC, received combination chemotherapy with PLD 25 mg/mq on d 1, 5FU 1200 mg/mq in 48 h continuous infusion, and oral FA 30 mg on d 1 and 2 every 3 wk until disease progression or intolerable toxicity. RESULTS: The median age was 65 years (range 41-82) and 28 patients were hepatitis C virus seropositive (90%). The majority of patients were Child-Pugh Class B (55%). Two patients showed a partial response (PR), and 16 had stable disease (SD). With a median follow-up of 14 mo, the median time to progression of all evaluable patients was 4 mo (95% CI 1.7-7). Median overall survival was 9 mo (95% CI 3-24 mo). After 1 year, 9 of 18 PR/SD patients were alive. Chemotherapy was well tolerated. CONCLUSION: PLD/FU/FA combination seems capable of achieving durable stabilization of HCC. The manageable toxicity supports a role for combination with other anticancer agents.

Theoretical Investigation on the Absence of Spore Photoproduct Analogue at Cytosine-Thymine Site

As a well known DNA photolesion product, the special UV induced pyrimidine dimmer called spore photoproduct （SP）, has aroused strong research interests. The SP formation was reported solely between two adjacent thymidine residues. It remains unclear in pervious experimental observations why there is an absence of the cytosine-derived SP-like photoprod- uct formation at the cytosine containing DNA strand, although the cytosine residue holds great similarity to thymine in terms of molecular structure. From a theoretical perspec- tive, we have explored this issue in this work by means of density functional theory at the B3LYP/6-311＋＋G（d,p） //B3LYP/6-31G（d,p） level for the DNA dinucleotide fragment, cy- tosine phosphate thymine （CpT）. Key factors blocking the formation of the SP-like product between two adjacent cytosine and thymidine residues are revealed. Instead of undergoing photochemical SP reaction, a photophysical deactivation pathway back to the ground state turns out to be favorable for the CpT dinucleotide fragment.