Strand-scission of HeLa cell deoxyribonucleic acid by bleomycin in vitro and in vivo

The ability of bleomycin (BLM) to cause strand breaks of DNA in vitro has been confirmed by means of alkaline sucrose sedimentation. BLM-induced breakage of extracted HeLa S3 cell DNA occurs extensively after dialysis following reaction with 50 μg/ml of BLM. 2-Mercaptoethanol (0·003 M) is not obligatory for BLM-induced degradation of DNA in vitro, but enhances it. The presence of 0·025 M EDTA in the reaction mixture completely prevents single-strand breaks of DNA by BLM. In good agreement with the in vitro results, the DNA from the HeLa cell lysate prepared in a lysing medium containing 0·015 M of EDTA is fragmented a little after a 30-min treatment with 25 μg/ml of BLM, while nonspecific and extensive degradation of DNA occurs when the cells are lysed in the absence of EDTA. A 30-min treatment with BLM also provokes a small amount of unscheduled incorporation of ³H-thymidine into non-S phase cells, indicating that a small number of single-strand breaks induced are repair-patched. Moreover, 25 μg/ml of BLM exerts a somewhat inhibitory effect on the joining of short segments of replicating DNA after a 30-min ³H-thymidine pulse, but the joining ability is soon resumed. These data suggest that BLM may either hardly enter HeLa S3 cells or may be readily inactivated.     

Tallysomycin, a new antitumor antibiotic complex related to bleomycin. V. Production, characterization and antitumor activity of tallysomycin S10b, a new biosynthetic tallysomycin derivative

Tallysomycin S10b is a biosynthetic derivative of tallysomycin B obtained by precursor amine-feeding fermentation. Tallysomycin S10b contains 1,4-diaminobutane as the terminal amine moiety in place of spermidine of tallysomycin B, and its assigned structure was verified by carbon-13 NMR spectrum. The antitumor activity of tallysomycin S10b was comparable to that of tallysomycin A against sarcoma 180 but less active than the latter against leukemia P388. Tallysomycin S10b was less toxic than tallysomycin A in terms of acute and subacute LD50 values. The nephrotoxic potential of tallysomycin S10b in rats was less than that of tallysomycin A.     

Damage of rat liver deoxyribonucleic acid by bleomycin

Bleomycin has been shown by other investigators to react with DNA in vitro causing the release of free bases and strand scission. We have examined the possibility of damage to rat liver DNA in vivo by bleomycin. Thirty min after injection of bleomycin (20 mg/kg body wt), the DNA was shifted to the top of an alkaline sucrose gradient, suggesting damage in vivo. By 3 hr after injection of bleomycin, a normal gradient pattern was observed. Perfusion of the liver prior to squashing (preparation of the liver DNA for gradient analysis) abolished the effect of bleomycin on the DNA, indicating that bleomycin did not damage the DNA in vivo, but is present in the blood and reacts with DNA after squashing and/or lysing of the cells.     

Tallysomycin, a new antitumor antibiotic complex related to bleomycin. IV. New biosynthetic derivatives of tallysomycin

A series of new biosynthetic derivatives of tallysomycins A and B were obtained by precursor amine-feeding fermentation. Certain diamines were incorporated into tallysomycins as the terminal amine moiety affording two classes of biosynthetic derivatives, with or without the Beta-lysine moiety in the subterminal position. Among 21 derivatives prepared, tallysomycin S10b was selected for further studies in view of its favorable therapeutic indices.     

Mode of action of saframycin antitumor antibiotics: sequence selectivities in the covalent binding of saframycins A and S to deoxyribonucleic acid

The sequence-preferential reversible covalent binding of certain saframycin antitumor antibiotics from Streptomyces lavendulae has been examined by complementary-strand methidium propyl-EDTA (MPE) footprinting on an EcoRI/HindIII restriction fragment of pBR322 DNA under several experimental conditions. A buffer at pH 7.4 and in the presence of 9.5 mM dithiothreitol at 37 degrees C was found to be optimum for the interaction of these antibiotics with DNA. At r' = 0.6 both saframycins A and S exhibited footprints in the regions 4244-4257 (CAAATAGGGTTCC) and 4265-4286 (TTCCCCAAAAGTGCCACCTG) and a weak footprint in the region 4297-4302 (AACCAT). The binding locations identified that are common to saframycins A and S are (all 5'----3') GGGG (4250-4253), CCCC (4268-4271), and GCC (4279-4281), and weak interaction locations are ACC (4282-4284 and 4298-4300) (underlined bases are shared by two adjacent binding sites). Both the antibiotic saframycins A and S show preference for 5'-GGG or 5'-GGC sequences. It appears that saframycin A has no affinity for 5'-CGG while saframycin S shows a strong footprint at this sequence. Neither of the saframycins recognizes alternating CG sequences. Saframycin S also binds to 5'-CTA, which suggests that molecular recognition processes involving the parent antibiotics are also important, and not only recognition by, and covalent binding of, the common iminium species to the DNA. The protection sites at 5'-GCC and 5'-ACC suggest that saframycins A and S recognize 5'-GGPy sequences. However, between the two pyrimidine bases, C is preferred to T. Enhancement of cleavage by both saframycins is observed in the AT-rich region of 4301-4318.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enhancement of antitumor activity of bleomycin by benzamide in vitro and in vivo

The cytotoxic effects of bleomycin on HeLa cells in culture were enhanced by incubation of the cells with benzamide, a potent inhibitor of poly(ADP-ribose) polymerase, at concentrations at which benzamide alone did not show any cytotoxicity. Benzamide plus bleomycin display enhanced therapeutic effects against Ehrlich ascites tumor cells in vivo. On daily treatment with various doses of bleomycin plus benzamide for 10 days, mice with Ehrlich ascites tumors survived longer than mice on treatment with bleomycin alone.     

In vitro susceptibilities of clinical isolates of coagulase-negative staphylococci to glycopeptide antibiotics

Two hundred and thirteen clinical strains of coagulase-negative staphylococci isolated in Japan between 1980 and 1997 were analyzed for glycopeptide susceptibility by determining MIC using both Mueller-Hinton agar (MHA) and Brain Heart Infusion agar (BHIA) plates. Of 37 Staphylococcus epidermidis strains isolated between 1980 and 1981, all were susceptible to vancomycin and teicoplanin on both MHA and BHIA. However, of 122 isolates of Staphylococcus epidermidis isolated between 1994 and 1997, 1 (0.8%) was intermediate to vancomycin on MHA and 39 (32%) were intermediate on BHIA, while 3 (2.5%) and 27 (22.1%) were intermediate or resistant to teicoplanin on MHA and BHIA, respectively. It was demonstrated that the susceptibilities of the strains in 1990s to vancomycin and teicoplanin were significantly decreased compared with those in 1980s. Population analysis was performed with six strains each of Staphylococcus epidermidis and Staphylococcus haemolyticus (three with vancomycin MIC > or = 8 micrograms/ml and three with vancomycin MIC < or = 4 micrograms/ml using BHIA). The population curves of the Staphylococcus epidermidis strains showed a homogeneous pattern of susceptibility. Whereas, those for two Staphylococcus haemolyticus strains (vancomycin MIC = 8 micrograms/ml using BHIA) showed a typical heterogeneous pattern. Vancomycin-resistant mutants (MIC > or = 32 micrograms/ml) were obtained with a high frequency of 10(-4)-(-5) from the strains by one-step selection with 16 micrograms/ml of vancomycin.     

Studies on the effects of the antitumor agent camptothecin and derivatives on deoxyribonucleic acid: Mechanism of the scission of deoxyribonucleic acid by photoactivated camptothecin

Aqueous solutions of the antitumor alkaloid camptothecin, its sodium salt, or a number of derivatives, when irradiated with 360 nm light in the presence of covalently closed circular DNA, produced single strand breaks in the DNA. The chromophore essential for the scission reaction consisted of intact rings A, B, C and D. The overall DNA breakage showed an inverse dependence on oxygen, and there is evidence for at least two reaction mechanisms. Photosensitization of the alkaloid may generate radicals which attack DNA or, in the presence of oxygen, generate hydroperoxy radicals. The photolytic reaction of camptothecin itself proceeded via formation of the photolabile camptothecin hemiacetal 4 which was isolated and characterized, and which suggests an alternative mechanism. In the anaerobic pathway, photodecarbonylation of the alkaloid may generate a diradical which can collapse to the racemized 4 or abstract hydrogen atoms from DNA leading to strand scission. In the presence of oxygen, the alternative aerobic pathway, in which hydroperoxy radicals are generated, can supervene leading to the generation of hydrogen peroxide and, then. the principal reactive species, the hydroxyl radical, which can then attack DNA. The intermediacy of these three species was proven unambiguously by (i) selective inhibition of scission with Superoxide dismutase, (ii) selective inhibition with catalase, and (iii) spin-trapping and e.s.r. spectroscopy respectively. Certain antibiotics, which show a preference for binding to A,T rich regions of DNA, showed an enhancement in the extent of photoinitiated camptothecin cleavage. By contrast, antibiotics which have a preference for binding to G,C rich regions had a suppressing effect on such photodynamic DNA breakage. Natural negatively supercoiled CCC-DNA showed a greater extent of photoinitiated camptothecin breakage than DNA which had been relaxed with a topoisomerase. These results may indicate a weak intercalative interaction of the alkaloid into the G, C regions of DNA. The relative efficiencies of several synthetic analogues of camptothecin in promoting photoinduced DNA cleavage were compared.     

Role of the glycopeptide framework in the antibacterial activity of hydrophobic derivatives of glycopeptide antibiotics

The antibacterial properties of glycopeptide antibiotics are based on their interaction with the d-Ala-d-Ala containing pentapeptide of bacterial peptidoglycan. The hydrophobic amides of vancomycin (1), teicoplanin (2), teicoplanin aglycon (3), and eremomycin (4) were compared with similar amides of minimally or low active des-(N-methyl-d-leucyl)eremomycin (5), eremomycin aglycon (6), des-(N-methyl-d-leucyl)eremomycin aglycon (7), and a teicoplanin degradation product TB-TPA (8). All hydrophobic amides of 1, 3, 4, and 6 were almost equally active against glycopeptide-resistant enterococci (GRE) [minimum inhibitory concentrations (MIC) 相似文献   

The binding of the anthelmintic pyrvinium cation to deoxyribonucleic acid in vitro

Pyrvinium pamoate, an anthelmintic drug with mutagenic activity, binds to duplex DNA in vitro. Binding markedly enhances the intrinsic fluorescence of the drug, permitting a characterization of the binding reaction to be performed directly through fluorimetric analysis. The monopyrvinium salt, pyrvinium iodide, binds cooperatively to native calf thymus DNA with an intrinsic binding constant of 1.1 X 10(4) M-1 as determined fluorimetrically and 1.4 X 10(4) M-1 as determined by equilibrium dialysis. The respective binding site was estimated to be 2.5 base pairs. A change in the absorption of bound drug as a function of the binding ratio identifies secondary, nonfluorescent binding which is essentially ionic in character. The binding of pyrvinium removes negative supercoils from PM2-ccc-DNA with an experimental efficiency greater than that observed for the intercalating drug chloroquine. Electron microscopy shows that molecules of pBR322 DNA treated with pyrvinium increase in length by as much as 18% over controls, consistent with an increase in DNA base pair separation upon binding. The evidence indicates that the primary interaction of pyrvinium with DNA involves intercalation.     

积雪草酸衍生物的合成及体外抗肿瘤活性研究

目的设计合成积雪草酸衍生物并检测其体外抗肿瘤活性。方法采用计算机辅助药物设计的方法设计并筛选目标化合物的结构;以天然产物积雪草酸为起始原料,对其C-2、C-3、C-23位羟基以及C-28位羧基进行结构改造;采用MTT法测试目标化合物对人肝癌细胞(HepG2)和人肺癌细胞(A549)的体外抗肿瘤活性。结果目标化合物对这两种细胞株的抑制活性均优于母体积雪草酸,其中化合物Ⅰ_8和Ⅱ_2表现出较强的抗肿瘤活性,且分子对接模拟也显示化合物Ⅰ_8和Ⅱ_2与Survivin蛋白的结合力较强。结论经结构改造后的积雪草酸衍生物具有一定的抗肿瘤活性,值得进一步研究。     

第二代糖肽类抗生素的研究进展

随着细菌耐药的不断升级，耐万古霉素肠球菌、高度耐糖肽类抗生素的金葡菌的出现，给临床抗感染治疗带来极大困难。开发新型糖肽类抗生素刻不容缓。人们对万古霉素等天然糖肽类抗生素进行化学修饰，得到了一系列有价值的衍生物，其中oritavancin、dalbavancin和telavancin已进入Ⅲ期临床和新药注册。本文对第二代糖肽类抗生素的结构特征、药效学与药动学特征以及临床应用作一综述。     

聚乙二醇缀合熊果酸衍生物的合成及体外抗癌活性

目的:合成具有一定水溶性的聚乙二醇(PEG)缀合熊果酸衍生物并考察其体外抗癌活性。方法:以熊果酸(ursolic acid,UA)作为母核,采用PEG、琥珀酸对其C28位进行接合修饰,合成一系列PEG缀合熊果酸衍生物并考察其水溶性和抗癌活性;采用流式细胞术探讨其抗癌机制。结果:所合成系列PEG缀合熊果酸衍生物的水溶性较熊果酸有明显改善,同时具有更显著的体外抗癌活性;化合物8c对5株受试肿瘤细胞的IC50(48 h)均小于10μmol.L-1,将AGS细胞阻滞于G2/M期并具有诱导凋亡作用(凋亡率高于70%)。结论:将熊果酸C28位进行PEG修饰是保持和提高其抗癌活性的有效途径之一。     

Antitumor effects of bis-thiosemicarbazones— inhibition of deoxyribonucleic acid synthesis in vitro

A group of bis-thiosemicarbazones was evaluated for potential antitumor activity, using the L1210 murine leukemia in cell culture. Drug levels required to inhibit DNA synthesis by 50 per cent, under standard conditions, were determined. The most potent of the agents examined had the structure X[CH₂CR₁=NNHCSNHR₂]₂ where X = C or S and R₁ = R₂ = CH₃. Optimal activity was also obtained with R₁ = H and R₂ = CH₃ only when X = S. The most potent derivatives inhibited DNA synthesis by 50 per cent within 10 min at 10^?6 M levels (id₅₀). Metal chelates of several compounds tested were extremely potent inhibitors of DNA synthesis ($\text{id}{}_{50}\text{= 10}{}^{\mathrm{?7}}\text{M or less}$). Insolubility in water and short duration of action in vivo may limit effectiveness of the bis-thiosemicarbazones.     

A procedure for examining deoxyribonucleic acid synthesis in regenerating liver in vitro

When hepatic slices prepared from rats subjected to a 70% partial hepatectomy 16 hr earlier are incubated in vitro, they will progress through a wave of DNA synthesis in a manner similar to that seen in vivo. However, slices prepared at 14 hr post-hepatectomy will not initiate DNA synthesis when incubated in vitro. These results were not altered by the addition of either 5% or 20% serum (calf serum or fetal calf serum) to the incubation medium. These results are interpreted as indicating that between 14 hr and 16 hr after partial hepatectomy there exists a critical period during which exposure to a systemic factor(s) commits hepatic cells to enter a round of DNA synthesis. The experimental procedures outlined in this report provide a means to study, in vitro, progression through the S-phase in a normal mammalian cell population. In addition, experiments employing slices prepared at 14 hr post-hepatectomy might aid in discerning factors initiating the G₁ to S-phase conversion. The experimental protocol described is applicable to studies aimed at assessing selected aspects of the effects of drugs and putative toxicants on hepatic function.