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PubMed1990

Preparation of a mixture of nucleoside triphosphates suitable for use in synthesis of nucleotide phosphate sugars from ribonucleic acid using nuclease P1, a mixture of nucleoside monophosphokinases and acetate kinase.

This paper (1) describes the enzymatic synthesis of a mixture of adenosine, guanosine, cytidine, and uridine triphosphates (ATP, GTP, CTP, and UTP) from ribonucleic acid (RNA). RNA was hydrolyzed by nuclease P1 to a mixture of 5'-nucleoside monophosphates. This mixture was converted to the nucleoside triphosphates using a mixture of nucleoside monophosphate kinases and acetate kinase, with acetyl phosphate as the ultimate phosphoryl donor. The nucleoside monophosphokinases were extracted from brewer's yeast in a four-step procedure. The specific activity of the yeast enzyme preparation after gel permeation chromatography was sufficiently high that the yeast kinases could be immobilized in volumes that were practical for laboratory scale syntheses. Conversions from NMP to NTP in a mixture containing 0.34 mol of total nucleoside phosphates were: ATP, 90%; GTP, 90%; CTP, 60%; and UTP, 40%.

Acetate Kinase↗
PubMed1983

Complete analysis of tRNA-modified nucleosides by high-performance liquid chromatography: the 29 modified nucleosides of Salmonella typhimurium and Escherichia coli tRNA.

A high-performance liquid chromatography (HPLC) method has been developed to quantify the major and modified nucleoside composition of total, unfractionated transfer RNA. The method is rapid and sensitive and offers a high degree of chromatographic resolution suitable for quantifying both stable and unstable modified nucleosides. It is nondestructive and allows the recovery of nucleosides for further characterization. We apply the method in the analysis of the 29 modified nucleosides in tRNA from Salmonella typhimurium (and Escherichia coli) and show it to be useful in examining changes in the modified nucleoside content of tRNA. Such changes may be important in regulation.

Chemical Phenomena↗
PubMed2005

Post mortem degradation of nucleosides in the brain: comparison of human and rat brains for estimation of in vivo concentration of nucleosides.

There is an increasing attention paid for nucleoside metabolism and changes of nucleoside concentrations in human brain because of its pathological and physiological relevance. In order to determine the post mortem degradation of nucleosides and nucleoside metabolites, the concentrations of four nucleosides and three nucleobases were measured in rat and neurosurgical human cerebral cortical samples with 30s to 24h post mortem delay. Adenosine degradation coefficient (a multiplying factor for calculating concentrations of investigated substances for the living state) was 0.886 for human brain at 2 h post mortem time, while it was 1.976 for rats. Hypoxanthine, an adenosine degradation product had coefficients 0.564 for human brain and 0.812 for the rat brain. We provide data and degradation coefficients for the concentrations of adenosine, guanosine, inosine, uridine, uracil, hypoxanthine and xanthine with 2, 4, 6 and 24 h post mortem delay. We also report a method how to validate human neurosurgical brain samples in terms of sample preparation and statistical analysis.

Aged↗
PubMed1985

Nucleosides. 133. Synthesis of 5-alkenyl-1-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)cytosines and related pyrimidine nucleosides as potential antiviral agents.

The synthesis of 1-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)cytosines with a halovinyl or vinyl substituent at C-5 was accomplished from the corresponding 5-iodo (FIAC, 1) and/or 5-chloromercuri nucleoside analogues with use of Li2PdCl4- and Pd(OAc)2-mediated coupling reactions. Thiation of the benzoylated derivative of the 5-ethyluracil nucleoside 3 followed by S-methylation and then ammonolysis provided 5-ethyl-2'-fluoro-ara-C. 5-Ethynyl-2'-fluoro-ara-C (19a) and 5-ethynyl-2'-fluoro-ara-U (19b) were also obtained from the persilylated 5-iodo nucleosides 1 and 16, respectively, by PdII/CuI catalyzed coupling with (trimethylsilyl)acetylene. With use of selective sugar deprotection of the initial coupling products with H2O/Me2SO, the corresponding 5-[2-(trimethylsilyl)ethynyl] derivatives 18a and 18b could be isolated. Most of the new compounds showed activity in vitro against both HSV-1 and HSV-2, as did the known corresponding 5-alkenyluracil nucleosides synthesized earlier. The 5-vinylcytosine and -uracil nucleosides 10 and 24, respectively, were highly effective against HSV-1 (ED90 = 0.40 and 0.043 microM, respectively) and HSV-2 (ED90 = 0.59 and 0.56 microM, respectively). Unlike BVDU, the 2'-fluoroarabinosyl derivatives of 5-(halovinyl)cytosine and -uracil showed activity against both types of herpes simplex virus. The therapeutic indices of these compounds are in some cases superior to those of 2'-fluoro-5-methyl-ara-U (FMAU, 2). Moderate antileukemic activity was observed in vitro for the 5-alkynyl and 5-vinyl compounds. The competition of these compounds with thymidine for viral-induced thymidine kinases was also studied.

Animals↗
PubMed2004

Facile deprotection of O-Cbz-protected nucleosides by hydrogenolysis: an alternative to O-benzyl ether-protected nucleosides.

[reaction: see text] Because of side-reactions encountered during hydrogenolysis, benzyl ethers are usually not an effective protecting group for nucleosides. Benzyloxycarbamates provide an alternative to traditional benzyl ethers for protection of nucleoside hydroxyl groups, as they are much more labile to hydrogenolysis. Deprotection conditions using transfer hydrogenolysis are described that avoid the reduction of the pyrimidine nucleobase during deblocking of O-Cbz-protected nucleosides. Additionally, an experiment is described that suggests the nucleobase component of a nucleoside is responsible for the sluggish hydrogenolysis of nucleosides.

Benzyl Compounds↗
PubMed1995

Regulation of nucleoside diphosphate kinase and an alternative kinase in Escherichia coli: role of the sspA and rnk genes in nucleoside triphosphate formation.

We have previously reported that two genes cloned from a cosmid library of Escherichia coli can restore mucoidy to an algR2 mutant of Pseudomonas aeruginosa. AlgR2 is a protein involved in the regulation of nucleoside diphosphate kinase (Ndk) as well as alginate synthesis in P. aeruginosa. One of the E. coli genes, rnk, encodes a 14.9 kDa protein with no homology to any other proteins. The other gene, sspA, encodes the stringent starvation protein, a regulatory protein involved in stationary-phase regulation and the stringent response of E. coli. While both rnk and sspA restored alginate production to the P. aeruginosa algR2 mutant, only rnk restored Ndk activity to the mutant. In this report, we have examined the effect of mutations in rnk and sspA on the levels of Ndk in E. coli. We find that a mutation in rnk drastically reduces the level of Ndk in E. coli. A mutation in sspA, however, affects the level of another nucleoside diphosphate kinase distinct from Ndk. The proteins can be easily distinguished from each other by their different affinities for nucleoside diphosphates (NDPs) and also by the differential effect of anti-Ndk antibodies on the reactions they catalyse. The ability of either of these two proteins to restore alginate synthesis in the algR2 mutant of P. aeruginosa demonstrates the importance of nucleoside triphosphate synthesis and energy metabolism for alginate synthesis. Additionally, a role for the stringent starvation protein (SspA) in the modulation of nucleoside triphosphate (NTP) levels in E. coli is also suggested from these experiments.

Adhesins, Bacterial↗
PubMed2004

Interaction of nucleoside inhibitors of HIV-1 reverse transcriptase with the concentrative nucleoside transporter-1 (SLC28A1).

Human concentrative nucleoside transporter-1 (hCNT1) (SLC28A1) is a widely expressed, high-affinity, pyrimidine-preferring, nucleoside transporter implicated in the uptake of naturally occurring pyrimidine nucleosides as well as a variety of derivatives used in anticancer treatment. Its putative role in the uptake of other pyrimidine nucleoside analogues with antiviral properties has not been studied in detail to date. Here, using a hCNT1 stably transfected cell line and the two-electrode voltage-clamp technique, we have assessed the interaction of selected pyrimidine-based antiviral drugs, inhibitors of HIV-1 reverse transcriptase such as zidovudine (AZT), stavudine (d4T), lamivudine (3TC) and zalcitabine (ddC), with hCNT1. hCNT1 transports AZT and d4T with low affinity, whereas 3TC and ddC are not translocated, the latter being able to bind the transporter protein. Selectivity appears to rely mostly upon the presence of a hydroxyl group in the 3'-position of the ribose ring. Thus, hCNT1 cannot be considered a broad-selectivity pyrimidine nucleoside carrier; in fact, very slight changes in substrate structure provoke a dramatic shift in selectivity.

Animals↗
PubMed1983

Selective inhibition of anti-nucleoside-specific antibody production by nucleoside-ricin A conjugate.

The effect of the preincubation of peripheral blood lymphocytes, from SLE patients, with nucleoside-ricin A conjugates on spontaneous A,G,C,T antibody production was examined. Enhanced spontaneous anti-nucleoside-specific antibody (anti-A,G,C,T antibody) production by SLE B cells was selectively inhibited by pretreatment in vitro with nucleosides conjugated to the ricin A chain. The selective suppression was demonstrated by the lack of suppression of the anti-DNP response or of polyclonal IgG production by pretreatment that did suppress anti-A,G,C,T production by the lymphocytes of eight patients with SLE. Furthermore, pretreatment of B cells, but not of T cells, with nucleoside-ricin A conjugates inhibited the A,G,C,T antibody response by these B cells. Thus, (A,G,C,T)-BGG-ricin A conjugates bind directly to the nucleoside-specific B cells via their antigen receptors. This demonstration of the selective elimination of B cells might have therapeutic applications in SLE.

Antibodies, Antinuclear↗
PubMed2005

Residue 33 of human equilibrative nucleoside transporter 2 is a functionally important component of both the dipyridamole and nucleoside binding sites.

Human equilibrative nucleoside transporters 1 and 2 (hENT1 and hENT2) differ functionally in that hENT2 generally displays lower affinity for its nucleoside permeants and is less sensitive to inhibition by the coronary vasodilators dilazep and dipyridamole. In previous work, we demonstrated that mutation of residues 33 (Met versus Ile) of hENT1 and hENT2 altered sensitivity to dilazep and dipyridamole and that the hENT2 mutant (I33M) displayed a K(m) value for uridine that was lower than that of hENT2 and similar to that of hENT1 (J Biol Chem 277:395-401, 2002). In this study, we report results of an in-depth investigation of the role of residue 33 in hENT2. We found that hENT2-I33M displayed decreased K(m) values for both pyrimidine and purine nucleosides and increased V(max) values for purine nucleosides. Cys or Ser at position 33 had similar effects on the kinetic parameters of hENT2 as Met, indicating that hydrophobic (Met and Cys) or hydrogen-bonding energy (Ser) contributed to permeant binding by these residues. hENT2-I33M and I33C displayed increased sensitivities to dipyridamole compared with wild-type hENT2, hENT2-I33A, and hENT2-I33S, suggesting interaction of the sulfur atom of Met and Cys with aromatic moieties on dipyridamole. hENT2-I33C was inhibited by the membrane-impermeant sulfhydryl reactive reagent p-chloromercuribenzyl sulfonate, and uridine, adenosine, and dipyridamole protected against inhibition. Our results indicated that residue 33 resides in an extracellular domain as predicted by the current hENT2 topology model and suggested that it is a functionally important component of both the permeant and dipyridamole binding sites.

Adenosine↗
PubMed1999

Hydroxyurea potentiates the antiherpesvirus activities of purine and pyrimidine nucleoside and nucleoside phosphonate analogs.

Hydroxyurea has been shown to potentiate the anti-human immunodeficiency virus activities of 2',3'-dideoxynucleoside analogs such as didanosine. We have now evaluated in vitro the effect of hydroxyurea on the antiherpesvirus activities of several nucleoside analogs (acyclovir [ACV], ganciclovir [GCV], penciclovir [PCV], lobucavir [LBV], (R)-9-[4-hydroxy-2-(hydroxymethyl)butyl]guanine [H2G], and brivudin and nucleoside phosphonate analogs (cidofovir [CDV] and adefovir [ADV]). When evaluated in cytopathic effect (CPE) reduction assays, hydroxyurea by itself had little effect on CPE progression and potentiated in a subsynergistic (herpes simplex virus type 1 [HSV-1]) to synergistic (HSV-2) fashion the antiviral activities of ACV, GCV, PCV, LBV, H2G, ADV, and CDV. Hydroxyurea also caused marked increases in the activities of ACV, GCV, PCV, LBV, and H2G (compounds that depend for their activation on a virus-encoded thymidine kinase [TK]) against TK-deficient (TK(-)) HSV-1. In fact, in combination with hydroxyurea the 50% effective concentrations of these compounds for inhibition of TK(-) HSV-1-induced CPE decreased from values of 20 to > or = 100 microg/ml (in the absence of hydroxyurea) to values of 1 to 5 microg/ml (in the presence of hydroxyurea at 25 to 100 microg/ml). When evaluated in a single-cycle virus yield reduction assay, hydroxyurea at a concentration of 100 microg/ml inhibited progeny virus production by 60 to 90% but had little effect on virus yield at a concentration of 25 microg/ml. Under these assay conditions hydroxyurea still elicited a marked potentiating effect on the antiherpesvirus activities of GCV and CDV, but this effect was less pronounced than that in the CPE reduction assay. It is conceivable that the potentiating effect of hydroxyurea stems from a depletion of the intracellular deoxynucleoside triphosphate pools, thus favoring the triphosphates of the nucleoside analogues (or the diphosphates of the nucleoside phosphonate analogues) in their competition with the natural nucleotides at the viral DNA polymerase level. The possible clinical implications of these findings are discussed.

Animals↗
PubMed1981

Comparison of pyrimidine and purine nucleoside secretion and nucleoside kinase expression in resident and elicited peritoneal macrophages.

Thymidine was found in culture supernatants from both normal peritoneal macrophages and macrophages elicited in vivo with LPS under conditions in which there was no detectable TK activity. Abrogation of thymidine secretion was observed only when elicited macrophages were cultured in the presence of macrophage growth factor (MGF), whereas resident macrophages continued secreting under similar conditions. Similarly, TK activity was induced with MGF only in elicited, but not in resident, macrophages; LPS added in vitro failed to render resident macrophages susceptible to stimulation by MGF. Tests for adenosine and deoxyadenosine in macrophage supernatants proved that these nucleosides were not secreted by these cells. Although AK was consistently expressed in both resident and elicited macrophages, no significant dAK levels could be detected in either cell population. In contrast to TK, both purine nucleoside kinases could not be modulated or induced with MGF. The data indicate different reactivity to MGF by resident and elicited macrophages and a difference in pyrimidine vs purine nucleoside secretion as well as nucleoside kinase expression by these cells.

Animals↗
PubMed2002

Osteosarcoma cells, resistant to methotrexate due to nucleoside and nucleobase salvage, are sensitive to nucleoside analogs.

PURPOSE: To test a novel strategy for overcoming intrinsic resistance to methotrexate (MTX) in osteosarcoma (OS) due to nucleoside and nucleobase salvage (NS). METHODS: Four OS cell lines, found to be highly resistant to MTX, were tested to determine the dominant mechanism of resistance. Sensitivity to MTX was tested in the presence of dialyzed serum or the transport inhibitor dipyridamole (DP) to confirm the contribution of NS to MTX resistance. We then investigated whether increased NS activity could be exploited using cytotoxic nucleoside analogs. RESULTS: Like other cell types, OS cells are capable of circumventing inhibition of de novo nucleotide synthesis by relying on NS. MTX, at concentrations as high as 1 m M did not inhibit cell growth in culture medium supplemented with undialyzed serum. In contrast, when NS was inhibited by DP or in medium depleted of nucleosides and nucleobases, sensitivity to MTX was seen at nanomolar concentrations. In medium with dialyzed serum, thymidine and hypoxanthine provided dose-dependent protection from MTX toxicity at concentrations similar to those seen in human plasma. No evidence of other significant mechanisms of resistance were found. All four cell lines were sensitive to 3-day exposures to cytarabine (IC50 0.22 to 2.88 micro M) and vidarabine (IC50 0.09 to 0.95 micro M). CONCLUSIONS: Salvage of de novo nucleotide synthesis inhibition by extracellular thymidine and hypoxanthine, at physiologically relevant concentrations, contributes to resistance to MTX in OS. However, this same process may impart a collateral sensitivity to nucleoside analogs. These findings support clinical trials for patients with OS using nucleoside analogs, either alone or in combination.

Animals↗
PubMed1975

13C magnetic resonance investigation of mercury (II) binding to nucleosides and thiolated nucleosides in dimethyl sulfoxide.

Natural abundance, proton-decoupled 13C magnetic resonance spectroscopy is shown to be a useful technique for identifying the mercury (II) binding sites on nucleosides and especially thiolated nucleosides. Measurements made on dimethyl sulfoxide-d6 solutions, 0.5 M in nucleoside and 0.15 M in mercury, reveal that both CH3 HgCl and HgCl2 bind principally to the sulfur atoms of s6 Guo and s8 Guo. The 13C NMR spectra of the unthiolated nucleosides in the presence of excess (4:1) mercury reveal that HgCl2 binds to N-3 of cytidine, to more than one site on adenosine and guanosine, but not strongly to uridine. Excess HgCl2 shifts the thiocarbonyl carbon atoms in s6 Guo and s8 Guo approx. 16 ppm upfield compared to the free nucleosides, and there is evidence for additional coordination to N-7 of s6 Guo. Binding to the ribose hydroxyl groups is clearly ruled out. At least in these instances, 13C NMR proves to be useful for assigning the mercury (II) binding sites, complementing the results of proton magnetic resonance studies. Proton NMR data for the binding of CH3 HgCl and HgCl2 to s6 Guo and s8 Guo are also presented.

Binding Sites↗
PubMed1980

Molecular orbital studies on the structure of nucleoside analogs. III. Conformation of 3-deazapyrimidine nucleosides.

PCILO computations ahve been carried out on the conformational properties of 3-deazapyrimidine nucleosides namely: 3-deazauridine and 3-deazacytidine. These nucleoside analogs result as a consequence of the replacement of N(3) by a carbon atom and they become nucleoside antibiotics having cytostatic and antiviral properties. Both C(2')-endo and C(3')-endo sugar geometries have been considered and the results indicate that the conformational preferences of these nucleoside antibiotics are very similar to those of their parent nucleosides and more particularly so in the situations that occur in aqueous solutions. The important biological significance of the results has been discussed.

3-Deazauridine↗
PubMed1997

New nucleoside-5'-alkylphosphonophosphates and related compounds containing 2'-deoxycytidine, thymidine and adenosine as nucleoside component. Syntheses and their effects on tumor cell growth in vitro.

Recent studies have shown that phosphono analogs of cytidine-5'-diphosphate diacylglycerol (CDP-DAG) possessing a structurally modified lipid moiety exhibit antiproliferative activity in vitro. As an extension of our previous work we tried to elucidate whether the presence of the cytidine component is necessary for cytostatic activity. In this context we have synthesized similarly structured nucleoside-phospholipid conjugates containing nucleoside components other than cytidine, which also do not exhibit cytostatic properties as such. The compounds include 5'-alkyldiphosphates and 5'-alkylphosphonophosphates of 2'-deoxycytidine, thymidine and adenosine with different alkyl chain length as well as selected 3-hexadecyl-2-chloro-2-deoxyglycero-(1)-diphosphates and -phosphonophosphates of these nucleosides. The chemical structures of the newly synthesized nucleoside-phospholipid conjugates were confirmed by fast atom bombardment (FAB) and electrospray ionization (ESI) mass spectrometry. It was found that these compounds also inhibit the cell growth of different human cell lines, i.e. the presence of the cytidine component is not a necessary prerequisite for the antiproliferative activity of these nucleoside-phospholipid conjugates.

Adenosine↗
PubMed1986

Binding of cis- and trans-diamminedichloroplatinum(II) to deoxyribonucleic acid exposes nucleosides as measured immunochemically with anti-nucleoside antibodies.

We report the use of anti-nucleoside antibodies to probe for local denaturation of calf thymus DNA upon binding of the antitumor drug cis-diamminedichloroplatinum(II), cis-DDP, and the biologically inactive analogues trans-diamminedichloroplatinum(II), trans-DDP, and chloro(diethylenetriamine)platinum(II) chloride, [Pt(dien)Cl]Cl. These antibodies specifically recognize each of the four DNA nucleosides. They bind well to denatured DNA, but not to native DNA in which the bases are less accessible owing to Watson-Crick duplex structure. At relatively high levels of modification (D/N approximately 0.1), cis-DDP causes significant disruption of DNA base pairing as reflected by the increased binding of anti-cytidine, anti-adenosine, and anti-thymidine antibodies. At lower levels of platinum adduct formation, however, all four anti-nucleoside antibodies bind more to DNA modified with trans-DDP. This result indicates that adducts formed by trans-DDP disrupt the DNA structure to a greater extent than those formed by cis-DDP at low D/N ratios. Modification of DNA by the monofunctional complex [Pt(dien)Cl]Cl does not affect its recognition by anti-nucleoside antibodies, demonstrating that base pair disruption is a consequence of bifunctional binding. The relative anti-nucleoside antibody recognition of cis-DDP-modified DNA is anti-cytosine greater than anti-adenosine approximately anti-thymidine much greater than anti-guanosine, consistent with the major adduct being an intrastrand d(GpG) cross-link. These results reveal that base pair disruption in a naturally occurring DNA modified by either cis-DDP or trans-DDP is sufficient to be detected by protein (antibody) binding. The relevance of these findings to current ideas about the molecular mechanism of action of cis-DDP is discussed.

Animals↗
PubMed1995

Nucleosides and nucleotides. 141. Chemical stability of a new antitumor nucleoside, 2'-C-cyano-2'-deoxy-1-beta-D-arabino-pentofuranosylcytosine in alkaline medium: formation of 2'-C-cyano-2'-deoxy-1-beta-D-ribo-pentofuranosylcytosine and its antitumor activity.

We have designed 2'-C-cyano-2'-deoxy-1-beta-D-arabino- pentofuranosylcytosine (CNDAC) as a potential mechanism-based DNA-strand-breaking nucleoside, which showed potent tumor cell growth inhibitory activity against various human tumor cell lines in vitro and in vivo. When measuring the pKa of the 2' alpha-proton of CNDAC, we found that CNDAC epimerized to 2'-C-cyano-2'-deoxy-1-beta-D-ribo-pentofuranosylcytosine (CNDC) with concomitant degradation of both CNDAC and CNDC to cytosine and 1,4-anhydro-2-C-cyano-2-deoxy-D-erythro-pent-1- enitol. Kinetic analysis of these reactions showed that abstraction of the acidic 2'-proton of CNDAC and CNDC initiated the reactions, which quickly reached an equilibrium. In the equilibrium, a concentration ratio of CNDAC and CNDC was about 3:5. Concomitant degradation of these nucleosides was found to be rather slow. Deuterium incorporation experiments with CNDAC in a D2O buffer suggested the mechanism of the beta-elimination reactions is an E1cB type. These epimerization and degradation reactions were found even in neutral conditions (pH 7.5) and also occurred in RPMI 1640 cell culture medium. The discovery of which nucleoside possesses the predominate tumor cell growth inhibitory activity was important. While both nucleosides showed potent tumor cell growth inhibitory activity against three human tumor cell lines (colon carcinoma WiDr, small cell lung carcinoma SBC-5, and stomach carcinoma MKN-74 cells) in 48 h of incubation, in 20 min of incubation, CNDAC was 11-50 times more effective than CNDC. In vivo antileukemic activity of these nucleosides against a mouse P388 model, CNDAC was obviously superior to CNDC.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenocarcinoma↗
PubMed1996

Nucleosides and nucleotides. 158. 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)-cytosine, 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)uracil, and their nucleobase analogues as new potential multifunctional antitumor nucleosides with a broad spectrum of activity.

We previously designed 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)uracil (EUrd) as a potential multifunctional antitumor nucleoside antimetabolite. It showed a potent and broad spectrum of antitumor activity against various human tumor cells in vitro and in vivo. To determine the structure-activity relationship, various nucleobase analogues of EUrd, such as 5-fluorouracil, thymine, cytosine, 5-fluorocytosine, adenine, and guanine derivatives, were synthesized by condensation of 1-O-acetyl-2,3,5-tri-O-benzoyl-3-C-ethynyl-alpha,beta-D-ribo-pentofur anose (6) and the corresponding pertrimethylsilylated nucleobases in the presence of SnCl4 or TMSOTf as a Lewis acid in CH3CN followed by debenzoylation. The in vitro tumor cell growth inhibitory activity of these 3'-C-ethynyl nucleosides against mouse leukemia L1210 and human nasopharyngeal KB cells showed that 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)cytosine (ECyd) and EUrd were the most potent inhibitors in the series, with IC50 values for L1210 cells of 0.016 and 0.13 microM and for KB cells of 0.028 and 0.029 microM, respectively. 5-Fluorocytosine, 5-fluorouracil, and adenine nucleosides showed much lower activity, with IC50 values of 0.4-2.5 microM, while thymine and guanine nucleosides did not exhibit any activity up to 300 microM. We next evaluated the tumor cell growth inhibitory activity of ECyd and EUrd against 36 human tumor cell lines in vitro and found that they were highly effective against these cell lines with IC50 values in the nanomolar to micromolar range. These nucleosides have a similar inhibitory spectrum. The in vivo antitumor activities of ECyd and EUrd were compared to that of 5-fluorouracil against 11 human tumor xenografts including three stomach, three colon, two pancreas, one renal, one breast, and one bile duct cancers. ECyd and EUrd showed a potent tumor inhibition ratio (73-92% inhibition relative to the control) in 9 of 11 and 8 of 11 human tumors, respectively, when administered intravenously for 10 consecutive days at doses of 0.25 and 2.0 mg/kg, respectively, while 5-fluorouracil showed potent inhibitory activity against only one tumor. Such excellent antitumor activity suggests that ECyd and EUrd are worth evaluating further for use in the treatment of human cancers.

Animals↗