Isolation and characterization of 5-carbamoylmethyluridine and 5-carbamoylmethyl-2-thiouridine from human urine.

Two new modified uracil nucleosides, 5-carbamoylmethyuridine (ncm5U, I) and 5-carbamoylmethyl-2-thiouridine (ncm5s2U, II) were isolated from a 24 hr collection of a normal human urine. The structures were assigned on the basis of UV, NMR and mass spectral data and confirmed by comparison of the spectral data and HPLC mobilities with those of authentic samples. On the basis of experimental data it appears possible that 5-carbamoylmethyl-2-thio-uridine (ncm5s2U, II) may be a degradation product produced from a labile precursor by the chemical treatments during the isolation procedure.

Formation of a phosphoramide mustard-nucleotide adduct that is not by alkylation at the N7 position of guanine.

The reaction of 2'-deoxyguanosine 3'-monophosphate with phosphoramide mustard resulted in the formation of several adducts. One of these adducts was formed by linking phosphoramide mustard to the phosphate group of 2'-deoxyguanosine 3'-monophosphate rather than by the generally accepted mechanism involving alkylation at the N7 position of guanine. This adduct served as an acceptor for the transfer of 32p from [gamma 32P]ATP by polynucleotide kinase and thus could be detected by the sensitive 32p-postlabeling assay.

Identification of cis-dichloro-bis-isopropylamine platinum(II) as a major metabolite of iproplatin in humans.

Iproplatin is a quadrivalent second-generation platinum complex undergoing clinical evaluation. In plasma and urine of patients receiving this drug, iproplatin- and platinum-containing metabolites of iproplatin were separated by reverse-phase gradient high performance liquid chromatography. One of the metabolites was identified by cochromatography and electron impact mass spectrometry as cis-dichloro-bisisopropylamineplatinum(II), a metabolite formed by reduction of iproplatin.

Metabolism of 1-methyladenosine.

1-[methyl-8-14C] Adenosine was synthesized and its metabolic fate was determined in intact rat. It was found that approximately 57% of 1-[methyl-8-14C] adenosine administered iv was excreted unchanged in the urine and 33% of the excreted radioactivity in the urine was associated with the major metabolite 1-methyl-hypoxanthine and about 4.5% was associated with 1-methylinosine.

Radioimmunoassay for a novel urinary nucleoside, N6-succinyladenosine.

A sensitive radioimmunoassay has been developed for an unusual urinary nucleoside, N6-succinyladenosine (N6-SAR). Antibodies were raised in rabbits and the antibody specificity was established by binding inhibition radioimmunoassay of compounds with structural similarity to N6-SAR. The assay is sensitive enough to detect up to a nanomole of N6-SAR in 1 ml of urine.

Isolation and characterization of a novel nucleoside, 7-beta-D-ribofuranosylhypoxanthine, from the urine of a chronic myelogenous leukemia patient.

A novel nucleoside, in the amount of 400 micrograms, was isolated from a 24-h collection of urine of a chronic myelogenous leukemia patient. On the basis of ultraviolet, nuclear magnetic resonance, and mass spectrometry and chromatography, its structure was established to be 7-beta-D-ribofuranosylhypoxanthine. The ultraviolet and mass spectral data and the thin layer chromatographic mobilities of the natural material were identical to those of a synthetic sample.

Quantitative aspects of metal ion binding to certain transfer RNA anticodon loop modified nucleosides.

Magnesium and manganese ions bind strongly to the unusual transfer RNA anticodon loop nucleotides, N-[(9-beta-D-ribofuranosyl-9H-purin-6-yl)carbamoyl]-L-threonine 5'-monophosphate (pt6A) and uridine-5-oxyacetic acid 5'-monophosphate (pV). Potentiometric measurements have shown that the delta G for metal ion-pt6A complex formation is 2-3-times more exothermic than for AMP. Electron-nuclear longitudinal dipolar relaxation data yielded manganese-ligand atom distances which permit a three-dimensional construct of the complex in which metal is coordinated to the phosphate, carboxylate of the threonine side-chain (with the nucleotide in the anti glycosidic conformation) and N7 of the adenine ring.

On the function of N-[(9-beta-D-ribofuranosylpurin-6-yl)carbamoyl]threonine in transfer ribonucleic acid. Metal ion binding studies.

The unusual transfer ribonucleic acid (RNA) anticodon adjacent modified nucleoside N-[(9-beta-D-ribofuranosylpurin-6-yl)carbamoyl]threonine, t6A, and its N6-methyl (mt6A) and 5'-phosphate (pt6A) derivatives are efficient ligands for magnesium and manganese ions, as demonstrated by potentiometry and nuclear magnetic resonance spectroscopy. Analysis of the 1H and 13C NMR spectra of t6A in the presence of paramagnetic Mn (II) at 32-34 degrees C has shown that the metal ion binds to the carboxyl group and probably N6 of the side chain as well as the N1 or N7 atom of the adenine ring. A more specific and stronger metal-ligand complex between Mn(II) and pt6A is evident from the 1H, 13C, and 31P NMR data.

Metabolic fate of N6-benzyladenosine and N6-benzyladenosine-5'-phosphate in rats.

The radiolabeled antitumor nucleoside (14C-8)-N6-benzyladenosine and its (14C-8)-5'-phosphate were administered to rats intravenously, and their metabolic fate was studied. Twenty-nine percent of the radioactivity was recovered in the 48-hr urine collection after (14C-8)-N6-benzyladenosine administration. The following metabolites were isolated: unchanged N6-benzyladenosine (20%), adenine (12%), uric acid (5%), and N6-benzyladenine (0.

Synthesis and biological activity of N6-(n-alkylureido)purine ribonucleosides and their 5'-phosphates.

Syntheses and biological activities of 12 N6-(n-alkylureido)purine ribonucleosides (alkyl chain length of 1--10, 16, and 18 carbons) and three N6-(n-alkylureido)purine ribonucleoside 5'-phosphates (chain length of 4, 9, and 10 carbons) are described. The N6-(n-alkylureido)purine ribonucleosides were prepared by a reaction of (2',3',5'-tri-O-acetyl-beta-D-ribofuranosyl)-9H-purine-6-carbamate and n-alkylamine in refluxing pyridine. The 5'-nucleotides were prepared by direct phosphorylation of the corresponding ribonucleoside with phosphorus oxychloride and triethyl phosphate.

Isolation and characterization of N6-succinyladenosine from human urine.

From the urines of colon carcinoma patients and normal subjects we have isolated a nucleoside in which an amino group of aspartic acid is attached to the six position of purine ribonucleoside. The structure, N6-succinyladenosine, N-(9-B-D-ribofuranosylpurin-6-yl)aspartic acid was assigned on the basis of spectral data, chemical degradation, and by synthesis. The ultraviolet and mass spectra, chromatographic and electrophoretic mobilities, and the chemical properties of the naturally occurring nucleoside were identical to those of the synthetic N6-succinyladenosine.

Synthesis and biological activities of some N4-substituted 4-aminopyrazolo(3,4-d)pyrimidines.

Syntheses and biological activities of 26 N4-substituted 4-aminopyrazolo[3,4-d]pyrimidines as analogs of naturally occurring modified nucleic acid bases, N-(purin-6-ylcarbamoyl)-L-threonine and N6-(delta2-isopentenyl)adenine, are described. 4-Aminopyrazolo[3,4-d]pyrimidine was converted into the desired intermediate, ethyl pyrazolo[3,4-d]pyrimidine-4-carbamate (2). 4-Ureidopyrazolo[3,4-d]pyrimidines (6-26) were prepared by displacement of the ethoxy group of the carbamate 2 by amino acids and a variety of amines and by a reaction of 4-aminopyrazolo[3,4-d]pyrimidine (1) with isocyanates.

Purine and pyrimidine inhibitors of arginase.

1. Adenosine, inosine, adenine and uric acid are competitive inhibitors and cytidine and cytosine noncompetitive inhibitors of bovine liver arginase (L-arginine amidinohydrolase, EC 3.5.

Synthesis and biological activities of some N6-(nitro- and -aminobenzyl)adenosines.

Synthesis and biological activities of 12 analogs of N6-benzyladenosine are described. The compounds were prepared by two methods: (1) direct alkylation of adenosine with an appropriately substituted benzyl bromide to give the N1-substituted derivative which was then rearranged in base to give the N6-substituted compound, and (2) by nucleophilic displacement of chlorine in 6-chloropurine ribonucleoside, 6-chloro-2-aminopurine ribonucleoside, and 6-chloro-2-aminopurine with an amine. These analogs were examined for their growth inhibitory effect in cultured leukemic cells and also for their effect on adenosine aminohydrolase activity.

Synthesis properties of the naturally occurring N-[(9-beta-D-ribofuranosylpurin-6-yl)-N-methylcarbamoyl]-L-threonine (mt-6A) and other related synthetic analogs.

The naturally occurring modified nucleoside, N-[(9-beta-D-ribofuranosylpurin-6-yl)-N-methylcarbamoyl]-L-threonine (mt6A), and the corresponding glycine analog mg6A were synthesized from N6-methyl-2',3',5'-tri-O-acetyladenosine and the appropriately blocked isocyanates derived from threonine and glycine. The natural mt6A isolated from Escherichia coli tRNA (F. Kimura-Harada et al.

Synthesis and antitumor activity of 5'-phosphates and cyclic 3',5'-phosphates derived from biologically active nucleosides.

Syntheses and biological activities of 12 N6-substituted adenosine 5'-phosphates and 15 cyclic 3',5'-phosphates are described. Included among these are the cyclic phosphates of the naturally occurring anticodon adjacent modified nucleosides, N6-(delta2-isopentenyl)adenosine and N-(purin-6-ylcarbamoyl)-L-threonine ribonucleoside. Also reported in this paper are the 5'-phosphates and cyclic phosphates of the cytokinins, N6-benzyladenosine, kinetin ribonucleoside, 3-(chloro-trans-2-buten-2-yl)adenosine,6-o-chlorophenylureidopurine ribonucleoside, and 6-allylureidopurine ribonucleoside.