ABSTRACT
Trans-activator (Tat)-mediated human immunodeficiency virus type 1 (HIV-1) transcription is essential for the replication of HIV-1 and is considered a potent therapeutic target for HIV-1 inhibition. In this study, the Library of Pharmacologically Active Compounds (LOPAC1280) was screened using our dual-reporter screening system for repositioning as Tat-inhibitory compounds. Consequently, two compounds were found to be potent, with low cytotoxicity. Of these two compounds, Roscovitine (CYC202) is already known to be a Tat inhibitor, while gemcitabine has been newly identified as an inhibitor of Tat-mediated transcription linked to viral production and replication. In an additional screening using the ribonucleoside analogues of gemcitabine, two analogues (2'-C-methylcytidine and 3-deazauridine) showed a specific Tat-inhibitory effect linked to their anti-HIV-1 activity. Interestingly, these compounds did not affect Tat protein directly, while the mechanism underlying their inhibition of Tat-mediated transcription was linked to pyrimidine biosynthesis, rather than to alteration of the dNTP pool, influenced by the inhibition of ribonucleotide reductase. Taken together, the proposed functional screening system is a useful tool for the identification of inhibitors of Tat-mediated HIV-1 transcription from among a large number of compounds, and the inhibitory effect of HIV-1 transcription by gemcitabine and its analogues may suggest a strategy for developing a new class of therapeutic anti-HIV drugs.
Subject(s)
Anti-HIV Agents/pharmacology , HIV-1/drug effects , tat Gene Products, Human Immunodeficiency Virus/antagonists & inhibitors , 3-Deazauridine/pharmacology , Cell Line , Cytidine/analogs & derivatives , Cytidine/pharmacology , Deoxycytidine/analogs & derivatives , Deoxycytidine/pharmacology , Drug Repositioning , HIV-1/genetics , HIV-1/physiology , High-Throughput Screening Assays , Humans , Roscovitine/pharmacology , Small Molecule Libraries , Transcription, Genetic/drug effects , Virus Replication/drug effects , tat Gene Products, Human Immunodeficiency Virus/genetics , tat Gene Products, Human Immunodeficiency Virus/metabolism , GemcitabineABSTRACT
A series of 3-deazauridines (3-DU) analogues were synthesized and evaluated in vitro for their antiherpetic activity against HSV-1 on Vero cell lines by cell viability. A first campaign of tests suggested that C3-arylated-3-DU derivatives could constitute a novel family of antiherpetic agents. A second campaign of biological evaluations led to the discovery of two potent anti-HSV-1 agents with comparable activity than acyclovir.
Subject(s)
3-Deazauridine/pharmacology , Antiviral Agents/pharmacology , Herpesvirus 1, Human/drug effects , 3-Deazauridine/analogs & derivatives , 3-Deazauridine/chemical synthesis , Animals , Antiviral Agents/chemical synthesis , Antiviral Agents/chemistry , Cell Survival/drug effects , Chlorocebus aethiops , Dose-Response Relationship, Drug , Microbial Sensitivity Tests , Molecular Structure , Structure-Activity Relationship , Vero CellsABSTRACT
Pyrazofurin (NSC 143095) as the monophosphate derivative is a potent inhibitor of orotidine 5'-monophosphate (OMP) decarboxylase of the pyrimidine pathway and has been proposed to inhibit 5-aminoimidazole-4-carboxamide ribotide (AICAR) transformylase (EC 2.1.2.3) of the purine pathway (J. F. Worzalla, and M. J. Sweeney, Pyrazofurin inhibition of purine biosynthesis via 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranosyl 5'-monophosphate formyltransferase. Cancer Res., 40: 1482-1485, 1980). Measurement of levels of pyrimidine and purine intermediates in cultured mouse L1210 leukemia cells has shown that 25 microM pyrazofurin induces an 8-fold accumulation of OMP and large accumulations of intermediates proximal to the blockade with abrupt decreases in uridine and cytidine nucleotides. Considerable increases in the cellular concentrations of N-succino-AICAR (SAICAR), AICAR, 5-formamidoimidazole-4-carboxamide ribotide (FAICAR), IMP, XMP, and GMP at later times indicate that AICAR transformylase is not significantly inhibited in cultured cells; rather the purine pathway and the GMP branch are stimulated. However, addition of 25 microM 3-deazauridine (NSC 126849) to leukemia cells did result in inhibition of AICAR transformylase: AICAR and SAICAR accumulated, IMP disappeared and there was a large accumulation of guanosine nucleotides. Blockade of pyrimidine biosynthesis by derivatives of pyrazofurin or 3-deazauridine spares 5-phosphoribosyl-1-pyrophosphate and L-glutamine, elevated concentrations of which may stimulate initial reactions of purine biosynthesis and the reaction XMP----GMP.
Subject(s)
3-Deazauridine/pharmacology , Antibiotics, Antineoplastic/pharmacology , Hydroxymethyl and Formyl Transferases , Leukemia L1210/metabolism , Purines/biosynthesis , Pyrimidines/biosynthesis , Ribonucleosides/pharmacology , Uridine/analogs & derivatives , Acyltransferases/antagonists & inhibitors , Amides , Animals , Bicarbonates/metabolism , Mice , Orotidine-5'-Phosphate Decarboxylase/antagonists & inhibitors , Phosphoribosylaminoimidazolecarboxamide Formyltransferase , Pyrazoles , Ribose , Tumor Cells, CulturedABSTRACT
Because alterations in cell membrane sialoglycoconjugates can affect the behavior of neoplastic cells, we investigated the effects of in vitro treatment with antimetabolites used in cancer therapy on the expression of membrane sialic acid in cultured HL-60 leukemic cells. In these studies, cells were incubated with Vibrio cholerae neuraminidase to remove surface sialic acid. Reappearance of membrane sialic acid during drug treatment was followed (a) by measuring changes in radioactive surface labeling of viable cells with sodium metaperiodate-sodium[3H]-borohydride, (b) by measuring the decline in accessible surface galactosyl receptor sites which occurred coincident with membrane sialic acid replacement, and (c) by measuring the incorporation of [3H]glucosamine into membrane-associated neuraminidase-labile sialic acid. We were especially interested in learning whether drugs that affect intracellular pools of cytidine triphosphate (CTP), an important nucleotide intermediate in sialylation reactions, could inhibit regeneration of membrane sialic acid. 3-Deazauridine, a competitive inhibitor of CTP synthetase, depleted CTP pools and curtailed surface membrane resialylation with little or no effect on synthesis of de novo sialic acid from precursor sugars. The addition of cytidine restored CTP pools and sialic acid regeneration. Acivicin, a glutamine antagonist, also depleted CTP pools and curtailed surface membrane resialylation. In addition, it retarded de novo synthesis of sialic acid. The addition of cytidine restored intracellular CTP pools and sialic acid regeneration. However, both cytidine and guanosine were required to restore sialic acid synthesis from precursor sugars. 1-beta-D-Arabinofuranosylcytosine, a competitive inhibitor of sialic acid synthetase and of sialyltransferase, inhibited both de novo sialic acid synthesis and membrane resialylation. Only the latter effect was reversed by the addition of exogenous cytidine. Hydroxyurea, an agent shown previously to inhibit glycoconjugate production in hamster fibroblasts, curtailed membrane resialylation and de novo synthesis of sialic acid without depleting CTP pools. Doxorubicin, at levels that caused marked arrest of cell proliferation, had no effect on sialic acid synthesis or expression on the membrane surface. These data suggest that antimetabolites, apart from their cytotoxic effects or effects on cellular growth, may directly inhibit the expression of membrane sialic acid.(ABSTRACT TRUNCATED AT 400 WORDS)
Subject(s)
Antimetabolites, Antineoplastic/pharmacology , Leukemia, Myeloid, Acute/metabolism , Pyrimidines/antagonists & inhibitors , Sialic Acids/metabolism , 3-Deazauridine/pharmacology , Agglutination , Cell Line , Cytarabine/pharmacology , Cytidine Triphosphate/analysis , Doxorubicin/pharmacology , Glucosamine/metabolism , Humans , Isoxazoles/pharmacology , Lectins , N-Acetylneuraminic AcidABSTRACT
The antitumor agent 3-deazauridine (DAU) was administered rapidly to four patients before surgical removal of intracerebral tumor. Tumor, adjacent brain tissue, and temporalis muscle were assayed for DAU by high-pressure liquid chromatography. DAU penetrated comparably into tumor, brain, and muscle; in one patient, tissue concentrations were higher than concurrent plasma concentrations. The active metabolite 3-deazauridine 5'-triphosphate was quantitated in one tumor sample and greatly exceeded its Ki for cytidine 5'-triphosphate synthetase. DAU was also present in autopsy brain specimens from two patients treated shortly antemortem. Cerebrospinal fluid concentrations were 22.1 and 59.0%, respectively, of concurrent plasma concentrations during continuous infusion of DAU in two patients. Cerebrospinal fluid concentration was 3.1 microgram/ml 2 hr after a 30-min infusion of 1.5 g of drug per sq m and fell to 1.9 microgram/ml at 16 hr. Thus, DAU is capable of penetrating into intracerebral tumor, brain, and cerebrospinal fluid and is worthy of investigation in the treatment of intracerebral and meningeal neoplasms.
Subject(s)
3-Deazauridine/metabolism , Brain Neoplasms/metabolism , Brain/metabolism , Uridine/analogs & derivatives , 3-Deazauridine/cerebrospinal fluid , 3-Deazauridine/pharmacology , Blood-Brain Barrier , Brain Neoplasms/cerebrospinal fluid , Brain Neoplasms/drug therapy , Humans , Meningeal Neoplasms/drug therapy , Muscles/metabolismABSTRACT
When incubated with 1-beta-D-arabinofuranosylcytosine (ara-C), RPMI 6410 cells formed a hitherto unrecognized ara-C metabolite, 1-beta-D-arabinofuranosylcytosine diphosphate choline. This compound was characterized by (a) chromatographic behavior, (b) chemical and enzymatic hydrolysis, (c) phosphorus content, and (d) incorporation of [5-3H]ara-C and [methyl-14C]choline. Formation of 1-beta-D-arabinofuranosylcytosine diphosphate choline by RPMI 6410 cells was enhanced in the presence of 3-deazauridine (DU) and was preceded by that of 1-beta-D-arabinofuranosylcytosine triphosphate. The antiproliferative effects of ara-C and DU toward RPMI 6410 cells were potentiated when the agents were present together. The anabolism of ara-C during a 24-hr interval of culture was markedly enhanced by the presence of DU; cellular concentrations of 1-beta-D-arabinofuranosylcytosine triphosphate and 1-beta-D-arabinofuranosylcytosine diphosphate choline were 5- and 15-fold higher than those in the absence of DU. This enhancement appears to be the basis of the potentiation of cytotoxicity resulting from combination of the agents. Pretreatment of RPMI 6410 cells with DU resulted in enhanced rates of cellular uptake of ara-C. ara-C uptake under these circumstances was blocked by the inhibitor of nucleoside transport, nitrobenzylthioinosine.
Subject(s)
Choline/analogs & derivatives , Cytarabine/analogs & derivatives , Cytidine Diphosphate Choline/analogs & derivatives , Leukemia, Experimental/metabolism , 3-Deazauridine/pharmacology , Animals , Cell Division/drug effects , Cells, Cultured , Cytarabine/biosynthesis , Cytarabine/metabolism , Cytarabine/pharmacology , Cytidine Diphosphate Choline/biosynthesis , Drug Synergism , Humans , Hydrolysis , Leukemia, Experimental/drug therapy , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/metabolismABSTRACT
1-beta-D-Arabinofuranosylcytosine diphosphate choline was formed from 1-beta-D-arabinofuranosylcytosine (ara-C) during incubation in vitro of peripheral myeloblasts from patients with acute myelogenous leukemia and cultured cells (nonleukemic human lymphocytes, mouse lymphoma L5178Y, and HeLa); as well, 1-beta-D-arabinofuranosylcytosine diphosphate choline was formed in vivo in mouse leukemia L1210 cells and mouse liver. 3-Deazauridine enhanced the anabolism of ara-C in nonleukemic lymphocytes in vitro and leukemia L1210 cells in vivo but did not influence ara-C anabolism in the other cell types. In acute myelogenous leukemia myeloblasts incubated in vitro with ara-C, concentrations of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate were maximal after 8 hr of incubation and formation of the latter preceded that of 1-beta-D-arabinofuranosylcytosine diphosphate choline.
Subject(s)
Choline/analogs & derivatives , Cytarabine/analogs & derivatives , Cytidine Diphosphate Choline/analogs & derivatives , Leukemia, Experimental/metabolism , 3-Deazauridine/pharmacology , Animals , Cytarabine/biosynthesis , Cytarabine/metabolism , Cytarabine/pharmacology , Cytidine Diphosphate Choline/biosynthesis , Drug Synergism , Female , HeLa Cells/metabolism , Leukemia L1210/metabolism , Leukemia, Experimental/drug therapy , Leukemia, Myeloid, Acute/metabolism , Liver/metabolism , Lymphocytes/metabolism , Mice , Mice, Inbred StrainsABSTRACT
Therapeutic nucleoside analogue 3-deazauridine (DU) exerts cytotoxic activity against cancer cells by disruption of DNA synthesis resulting in cell death. The present study evaluates whether DU alone at doses 2.5-15 microM or in combination with all trans retinoic acid (RA) or dibutyryl cAMP (dbcAMP) is effective against myelogenous leukemia. The data of this study indicate that DU induces dose-dependent cell death by apoptosis in myeloid leukemia cell lines HL-60, NB4, HEL and K562 as demonstrated by cell staining or flow cytometry and agarose gel electrophoresis. 24h-treatment with DU produced dose-dependent HL-60 cell growth inhibition and dose-independent S phase arrest that was not reversed upon removal of higher doses of DU (10-15 microM). Exposition to nontoxic dose of DU (2.5 microM) for 24h followed by RA or dbcAMP and 96 h-cotreatment with DU significantly enhanced RA- but not dbcAMP-mediated granulocytic differentiation. Cell maturation was paralleled with an increase in the proportion of cells in G1 or G2+M phase. We conclude that, depending on the dose or the sequence of administration with RA, an inhibitor of DNA replication, DU triggers a process of either differentiation or apoptosis in myeloid leukemia cells.
Subject(s)
3-Deazauridine/pharmacology , Apoptosis/drug effects , Cell Differentiation/drug effects , Granulocytes/drug effects , Tretinoin/pharmacology , Bucladesine/pharmacology , Cell Cycle/drug effects , Cell Line, Tumor , Cell Survival/drug effects , DNA Fragmentation/drug effects , Dose-Response Relationship, Drug , Drug Synergism , Flow Cytometry , Granulocyte Precursor Cells/drug effects , HL-60 Cells , Humans , K562 Cells , Leukemia, Myeloid/drug therapy , Leukemia, Myeloid/pathologyABSTRACT
The cytosine analogue of neplanocin A, cyclopentenylcytosine (CPE-C, 3), has significant antitumor and antiviral activity commensurate with the drug's ability to produce a significant depletion of cytidine triphosphate (CTP) levels that result from the potent inhibition of cytidine triphosphate synthetase. Another important antitumor agent, previously identified as a potent inhibitor of the same enzyme, is 3-deazauridine (2). The synthesis of the cyclopentenyl nucleosides 3-deaza-CPE-C (5) and 3-deaza-CPE-U (6) was undertaken in order to investigate the effects of a modified 3-deaza pyrimidine aglycon moiety on the biological activity of the parent CPE-C. These compounds were synthesized via an SN2 displacement reaction on cyclopenten-1-ol methanesulfonate (10) by the sodium salt of the corresponding aglycon. In each case, separation and characterization of the corresponding N- and O-alkylated products was necessary before final removal of the blocking groups. The target compounds were devoid of in vitro antiviral activity against the HSV-1 and human influenza viruses. Although 3-deaza-CPE-C was nontoxic to L1210 cells in culture, 3-deaza-CPE-U displayed significant cytotoxicity against murine L1210 leukemia in vitro.
Subject(s)
3-Deazauridine/analogs & derivatives , Antimetabolites, Antineoplastic/chemical synthesis , Antiviral Agents/chemical synthesis , Cytidine/analogs & derivatives , Cytidine/chemical synthesis , Uridine/analogs & derivatives , Uridine/chemical synthesis , 3-Deazauridine/chemical synthesis , 3-Deazauridine/chemistry , 3-Deazauridine/pharmacology , Animals , Cytidine/chemistry , Cytidine/pharmacology , Drug Screening Assays, Antitumor , Indicators and Reagents , Leukemia L1210 , Mice , Molecular Structure , Structure-Activity Relationship , Uridine/chemistry , Uridine/pharmacologyABSTRACT
In view of the marked antitumor activity of 3-deazauridine, the synthesis of 4-(beta-D-ribofuranosyl)-1,3-dihydroxybenzene (1,3-dideazauridine) and its dibenzyl derivative was carried out. 4-Bromo-1,3-dihydroxybenzene was converted to its dibenzyl derivative, which, upon reaction with n-butyllithium followed by treatment with anhydrous cadmium chloride, gave bis(1,3-dibenzyloxyphenyl-4)cadmium. Condensation of this intermediate with 2,3,5-tri-O-benzoyl-D-ribofuranosyl chloride in refluxing toluene, and subsequent removal of the protecting benzoyl groups, afforded 4-(beta-D-ribofuranosyl)-1,3-dibenzyloxybenzene which, upon catalytic hydrogenation over Pd/C, furnished the desired 4-(beta-D-ribofuranosyl)-1,3-dihydroxybenzene. The beta configuration at the anomeric center was established by NMR and hydrogen bonding studies. 4-(Beta-D-ribofuranosyl)-1,3-dibenzyloxybenzene inhibited the growth of leukemia L1210 cells by 50% at 7 x 10(-6) M, and that of mammary carcinoma TA3 cells at 5 x 10(-5) M. Dideazauridine itself was less active, inhibiting the leukemia L1210 but not the TA3 cells at 1 x 10(-4) M, but the compound was significantly active against herpes simplex (type I) virus in vitro.
Subject(s)
3-Deazauridine/analogs & derivatives , Uridine/analogs & derivatives , 3-Deazauridine/chemical synthesis , 3-Deazauridine/pharmacology , Animals , Cells, Cultured , Escherichia coli/drug effects , Escherichia coli/growth & development , Female , Leukemia L1210/metabolism , Magnetic Resonance Spectroscopy , Mammary Neoplasms, Experimental/metabolism , Mass Spectrometry , Mice , Simplexvirus/drug effects , Streptococcus/drug effects , Streptococcus/growth & developmentABSTRACT
Novel 3-substituted analogues of 4-amino-1-beta-D-ribofuranosyl-2(1H)-pyridinone (3-deazacytidine, 3) and 4-hydroxy-1-beta-D-ribofuranosyl-2(1H)-pyridinone (3-deazauridine, 4) have been synthesized and tested for antitumor and antiviral activity. Thus the 3-chloro (9a), 3-bromo (9b), and 3-nitro (9c) analogues of 3 and the 3-chloro (9d), 3-bromo (9e), and 3-nitro (9f) analogues of 4 were prepared by standard glycosylating procedures. Novel requisite heterocycles 4-amino-3-chloro-2(1H)-pyridinone (7a) and 4-amino-3-bromo-2(1H)-pyridinone (7b) were prepared by halogenating 4-amino-2(1H)-pyridinone (5). Requisite heterocycles 4-amino-3-nitro-2(1H)-pyridinone (7c), 3-chloro-4-hydroxy-2(1H)-pyridinone (7d), 3-bromo-4-hydroxy-2(1H)-pyridinone (7e), and 4-hydroxy-3-nitro-2(1H)-pyridinone (7f) were synthesized by known procedures from 4-hydroxy-2(1H)-pyridinone (6). Structure proof of target nucleosides was provided by independent synthesis, 1H NMR, and UV. Compounds 9a-f were devoid of activity against intraperitoneally implanted L1210 leukemia in mice. Compound 9f displayed significant activity against rhinovirus type 34 grown in WISH cells. 4-Amino-3-fluoro-1-beta-D-ribofuranosyl-2(1H)-pyridinone (1) displayed good activity against intraperitoneally implanted P388 leukemia in mice, but it was devoid of activity against M5076 sarcoma, amelanotic (LOX) melanoma xenograft, and subrenal capsule human mammary carcinoma MX-1 xenograft in mice. Compound 1 also displayed significant activity against rhinovirus type 34.
Subject(s)
3-Deazauridine/analogs & derivatives , 3-Deazauridine/chemical synthesis , Antineoplastic Agents/chemical synthesis , Antiviral Agents/chemical synthesis , Cytidine/analogs & derivatives , Uridine/analogs & derivatives , 3-Deazauridine/pharmacology , 3-Deazauridine/therapeutic use , Animals , Cell Line , Cell Survival/drug effects , Cytidine/chemical synthesis , Cytidine/pharmacology , Cytidine/therapeutic use , Female , Humans , Indicators and Reagents , Leukemia L1210/drug therapy , Leukemia P388/drug therapy , Mice , Microbial Sensitivity Tests , Molecular Structure , Neoplasm Transplantation , Rhinovirus/drug effects , Rhinovirus/growth & development , Structure-Activity Relationship , Subrenal Capsule Assay , Transplantation, Heterologous , Tumor Cells, Cultured/cytology , Tumor Cells, Cultured/drug effectsABSTRACT
Exposure of cultured L1210 cells to [2-14C]deoxycytidine and analysis or radioactivity incorporated into DNA-pyrimidines revealed that 2.7--5.5-fold more radioactivity is incorporated into DNA-thymine than into cytosine bases. Thus the pathway involving deamination of deoxycytidylate to deoxyuridylate and methylation to thymidylate is highly favored over successive phosphorylation to dCTP. Several modified and endogenous pyrimidines altered the labelling of DNA-thymine and DNA-cytosine with [2-14C]deoxycytidine. 3-Deazauridine at 0.1 mM caused a 56% increase in the labelling of DNA-thymine. Both thymidine and 3-deazauridine (greater than or equal to 10 microM) increased the specific activity of DNA-cytosine by 4-fold. Cytosine arabinoside (ara-C) (greater than or equal to 10 microM) reduced the labelling of both DNA-cytosine and DNA-thymine. Excess cytidine (0.1 mM) reduced the labelling of DNA-cytosine by 40%. Tetrahydrouridine at concentrations up to 1 mM had no effect.
Subject(s)
DNA/metabolism , Deoxycytidine/metabolism , Thymidine/metabolism , 3-Deazauridine/pharmacology , Animals , Carbon Radioisotopes , Cytarabine/pharmacology , Cytidine/pharmacology , Leukemia L1210/metabolism , Mice , Tetrahydrouridine/pharmacology , Thymidine/pharmacologyABSTRACT
Cytotoxic effects of 3-deazaguanosine (3-DGUO) result from the inhibition of DNA synthesis and incorporation of the drug into DNA. Synergistic antiproliferative effects of a combination of 3-deazaguanosine and 2-beta-D-ribofuranosylthiazole-4-carboxamide, a potent inhibitor of inosine monophosphate dehydrogenase, was observed in human tumor cells. Inosine reversed the antiproliferative effects of the 3-deazaguanosine but not 2-beta-D-ribofuranosylthiazole-4-carboxamide. 3-Deazaguanosine monophosphate was shown to inhibit the activity of the de novo purine synthesis enzyme, 5'-phosphoribosyl-5-aminoimidazole-4-carboxamide transformylase. The data suggested a cytotoxic effect of 3-DGUO associated with the inhibition of de novo purine synthesis by drug nucleotides, an effect which may account for the synergistic action noted.
Subject(s)
Antimetabolites, Antineoplastic , Neoplasms/metabolism , 3-Deazauridine/pharmacology , Cell Division/drug effects , Cells, Cultured , Drug Synergism , Humans , Inosine/pharmacology , Ribavirin/analogs & derivatives , Ribavirin/pharmacologyABSTRACT
The effect of 3-deazauridine on the synthesis of uracil nucleotides by de novo and salvage pathways was investigated in intact cultured L1210 cells. De novo pyrimidine biosynthesis, as measured by sodium [14C]bicarbonate incorporation into uracil nucleotides, was inhibited 40-85% at intracellular 3-deazauracil nucleotide concentrations of 1-6 nmoles/10(6) cells. The inhibition was not due to an increase in the size of the uracil nucleotide pool since this pool was only 97-66% of control level at 3-deazauracil nucleotide concentrations of 1-6 nmoles/10(6) cells. Furthermore, intracellular 3-deazauracil nucleotide concentrations of 0.5 to 5.2 nmoles/10(6) cells inhibited the salvage of [14C]uridine by 25-75%. The data indicate that 3-deazauridine may potentiate its own inhibition of CTP synthetase by reducing the concentration of competing uracil nucleotides by inhibiting de novo pyrimidine biosynthesis and pyrimidine salvage. It is postulated that the biochemical mechanism by which 3-deazauridine inhibits uracil nucleotide synthesis is by acting as a fraudulent allosteric regulator of carbamyl phosphate synthetase II and uridine/cytidine kinase.
Subject(s)
3-Deazauridine/pharmacology , Leukemia L1210/metabolism , Pyrimidines/biosynthesis , Uracil Nucleotides/biosynthesis , Uridine/analogs & derivatives , Animals , Bicarbonates/metabolism , Carbon Radioisotopes , Chromatography, High Pressure Liquid , Kinetics , Mice , Sodium Bicarbonate , Uridine/metabolismABSTRACT
Mycophenolic acid, an inhibitor of the enzyme IMP dehydrogenase, antagonizes the CTP synthetase inhibitor 3-deazauridine in its anti-proliferative effects on MOLT-3 human T leukemia cells. No depletion of CTP occurred, and decreased amounts of 3-deazuridine-triphosphate were measured in cells incubated with mycophenolic acid and 3-deazuridine. Most probably, these phenomena are related to the increased amounts of PRPP observed, which can result in an increased pyrimidine biosynthesis de novo and, as a consequence, a decreased metabolism of 3-deazauridine via the salvage pathway.
Subject(s)
3-Deazauridine/pharmacology , Carbon-Nitrogen Ligases , Enzyme Inhibitors/pharmacology , IMP Dehydrogenase/antagonists & inhibitors , Ligases/antagonists & inhibitors , Mycophenolic Acid/pharmacology , Phosphoribosyl Pyrophosphate/metabolism , 3-Deazauridine/antagonists & inhibitors , Cell Division/drug effects , Cell Survival/drug effects , Dose-Response Relationship, Drug , Humans , Leukemia , Ribonucleotides/biosynthesis , Tumor Cells, CulturedABSTRACT
3-Nitro-3-deazauridine (3N-3DU) is a new synthetic nucleoside having activity against members of 5 RNA virus families including: paramyxoviruses (parainfluenza, PIV), picornaviruses (rhino-, RV), rhabdoviruses (vesicular stomatitis, VSV), togaviruses (Semliki Forest, SFV) and bunyaviruses (Punta Toro, PTV). In this report, we evaluate and compare its activity with the parent nucleoside, 3-deazauridine (3DU) and ribavirin as drug standards. Comparison of drug activities utilizes observations of antiviral indices, which are determined by the following formula: maximum tolerated dose (MTD)/minimum inhibitory concentration (MIC). The antiviral index (AI) of 3N-3DU (AI 15.3) was comparable to ribavirin and much higher than 3DU when evaluated against PIV. The 3N-3DU was the most active of the three when tested against RV (AI 24.1), SFV (AI 76.9) or VSV (AI 50). In contrast to the RV activity, 3N-3DU (AI 0.5) and 3DU (AI less than 0.1) were less active than ribavirin (AI 1.3) when evaluated against poliovirus, type 1 (PoV). Ribavirin (AI 10.0) was more active than 3N-3DU (AI 2.4) and 3DU (AI less than 0.1) against PTV. 3N-3DU exhibited comparable toxicity to ribavirin in KB cells, was 4-fold less toxic in WISH cells and 4-fold more toxic in LLC-MK2 cells. Overall, 3N-3DU is markedly less toxic than its parent nucleoside, 3DU. It appears from this study that the structural modification of 3DU resulting from the addition of the nitro group in the 3 position of the base reduces toxicity and enhances the antiviral activity.
Subject(s)
3-Deazauridine/analogs & derivatives , Antiviral Agents/pharmacology , RNA Viruses/drug effects , Uridine/analogs & derivatives , 3-Deazauridine/pharmacology , 3-Deazauridine/toxicity , Animals , Antiviral Agents/toxicity , Cells, Cultured , Drug Evaluation , Humans , In Vitro Techniques , Microbial Sensitivity Tests , Ribavirin/toxicityABSTRACT
A variety of antiviral compounds were examined for their inhibitory effect on measles (SSPE) virus plaque formation in VERO cells. The following compounds inhibited SSPE virus (strain Niigata-1) replication at concentrations that were significantly lower than their minimum cytotoxic concentrations: neplanocin A, neplanocin C, carbocyclic 3-deazaadenosine, 9-(trans-2', trans-3'-dihydroxycyclopent-4'-enyl)adenine, 9-(trans-2',trans-3'-dihydroxycyclopent-4'-enyl)-3-deazaadenine, (RS)-3-adenin-9-yl-2-hydroxypropanoic acid isobutyl ester, carbodine, cyclopentenyl cytosine, 3-deazaguanine, pyrazofurin, ribavirin and 6-azauridine. As the most selective inhibitors of SSPE virus replication emerged pyrazofurin, 3-deazaguanine, 6-azauridine and ribavirin. These compounds were further examined for their relative potency against a number of measles (SSPE) virus strains. Their order of (decreasing) potency was pyrazofurin greater than 6-azauridine approximately 3-deazaguanine greater than ribavirin. Amantadine, inosiplex and glycyrrhizin, that were also included in these assays, did not show appreciable activity against any of the measles (SSPE) virus strains.
Subject(s)
Antiviral Agents/pharmacology , Measles virus/drug effects , 3-Deazauridine/pharmacology , Amides , Animals , Azauridine/pharmacology , Measles virus/physiology , Nucleosides/pharmacology , Pyrazoles , Ribavirin/pharmacology , Ribonucleosides/pharmacology , Ribose , Vero Cells , Viral Plaque Assay , Virus Replication/drug effectsABSTRACT
Deazauridine inhibited growth of tumor cells in culture and in culture and in vivo; this agent was significantly more effective against L1210/AraC than against the parent sensitive line. Inhibition of growth of tumor cells in culture was prevented by uridine and cytidine and was partially alleviated by deoxycytidine, but not by deoxyuridine or thymidine. DeazaUR inhibited nucleic acid synthesis but not protein synthesis in tumor cells in culture; deoxycytidine alleviated inhibition of nucleic acid synthesis. The labeling of pyrimidine ribonucleotides by 6-14C-orotic acid was inhbited by deazaUR. DeazaUR treatment of tumor cells in culture resulted in increased uptake of cytidine-3H into RNA, whereas uridine-3H uptake into RNA was inhibited. Labelling of DNA by uridine-3H/ and cytidine-H was inhibited by deazaUR. Pools of CMP, CDP, and CTP decreased markedly during deazaUR treatment of L1210 cells in culture and in vivo. These observations in growing cells pointed to deazaUR inhibition of the synthesis of cytidylic acid. Deazauridine 5'-triphosphate was found to be an inhibitor of the synthesis of CTP from UTP catalyzed by enzyme preparations from L1210 cells. This observation is in agreement with those of McPartland et al.19 that deazaUTP inhibited CTP synthetase purified from calf liver. Deazauridine treatment of L1210 cells in culture stimulated the uptake of deoxycytidine-3H into DNA while inhibiting the uptake of 3H-labeled deoxyuridine, thymidine, deoxyadenosine, and deoxyguanosine. Intracellular pools of dCTP were decreased by deazauridine treatment in L1210 cells in culture and in vivo. Deazauridine 5'-diphosphate inhibited the enzymatic reduction of pyrimidine ribonucleoside 5'-diphosphates to the corresponding deoxyribonucleotides. These results are consistent with the view that deazauridine, after its uptake and intracellular phosphorylation, strongly inhibits the formation of CTP. This is considered to be the primary metabolic effect of the analog. A secondary effect appears to be an inhibition of dCTP formation.
Subject(s)
3-Deazauridine , Adenocarcinoma/metabolism , Cytarabine/pharmacology , Leukemia L1210/metabolism , Liver/enzymology , Uridine/analogs & derivatives , 3-Deazauridine/pharmacology , 3-Deazauridine/therapeutic use , Animals , Cell Line , DNA, Neoplasm/biosynthesis , Deoxycytidine/therapeutic use , Deoxyribonucleotides/biosynthesis , Drug Resistance , Leukemia L1210/drug therapy , Mice , Neoplasms, Experimental/metabolism , Orotic Acid/metabolism , Phosphotransferases/metabolism , Pyrimidine Nucleotides/biosynthesis , RNA, Neoplasm/biosynthesis , Ribonucleotide Reductases/metabolism , Ribonucleotides/biosynthesis , Uracil NucleotidesABSTRACT
The effect of high concentrations of exogenous dCyd on the growth inhibitory properties of several inhibitors of de novo pyrimidine biosynthesis (dThd, 3-DAU, PALA, PF) was examined in three cultured human leukemic cell lines (HL-60, K-562, KG-1), and a dCyd kinase-deficient, Ara-C-resistant variant (HL-60/Ara-C). In the presence of dCyd concentrations (10(-3) M), far exceeding normal human plasma levels (0.5 to 4.0 X 10(6) M), substantial but partial reversal of pyrimidine antagonist-mediated growth inhibition and restoration of intracellular dCTP levels was noted in all cell types except HL-60/Ara-C. When high concentrations of dCyd (10(-3) M) were combined with low levels of uridine or cytidine (10(-5) M), full restoration of growth was observed in sensitive cell lines. When exposed to supraphysiologic concentrations of dCyd, HL-60/Ara-C cells were more sensitive to the growth inhibitory effects of pyrimidine antagonists than parent HL-60 cells; this phenomenon was maximal at 10(-4) M dCyd and was not observed in the presence of dCyd concentrations of 10(-6) M or lower. These studies suggest that in the presence of low concentrations of uridine or cytidine, perturbations in intracellular dCTP pools may play a critical role in determining the in vitro antiproliferative response of human leukemic myeloid cells to diverse inhibitors of de novo pyrimidine biosynthesis. They also raise the possibility that modulation of exogenous dCyd concentrations may improve the therapeutic efficacy of pyrimidine antagonists toward certain salvage pathway-deficient, drug-resistant leukemic cells.
Subject(s)
Deoxycytidine/pharmacology , Leukemia, Experimental/metabolism , Pyrimidines/biosynthesis , 3-Deazauridine/pharmacology , Amides , Aspartic Acid/analogs & derivatives , Aspartic Acid/pharmacology , Cell Division/drug effects , Cell Line/drug effects , Deoxycytidine Kinase/deficiency , Drug Interactions , Humans , Phosphonoacetic Acid/analogs & derivatives , Phosphonoacetic Acid/pharmacology , Pyrazoles , Pyrimidines/antagonists & inhibitors , Ribonucleosides/pharmacology , Ribose , Thymidine/pharmacologyABSTRACT
The incorporation model of Sanyal et al. has been used to understand the biological activity of the cytostatic compound 3-deazauridine. The interaction energies of various types of binding pattern of the enterant molecule with nucleic acid fragments have been computed. The energy values and the sites of association of the analogous base, obtained by optimization of energy values as well as the sites of association of nucleic acid bases during the transcription process have been compared. The specificity of the binding of the interacting molecule has been discussed, along with the inhibitory effect of 3-deazauridine. They are in agreement with the experimentally observed evidence.