Pyrimidine benzamide-based thrombopoietin receptor agonists

A series of pyrimidine benzamide-based thrombopoietin receptor agonists is described. The lead molecule contains a 2-amino-5-unsubstituted thiazole, a group that has been associated with idiosyncratic toxicity. The potential for metabolic oxidation at C-5 of the thiazole, the likely source of toxic metabolites, was removed by substitution at C-5 or by replacing the thiazole with a thiadiazole. Potency in the series was improved by modifying the substituents on the pyrimidine and/or on the thiazole or thiadiazole pendant aryl ring. In vivo examination revealed that compounds from the series are not highly bioavailable. This is attributed to low solubility and poor permeability.     

Effect of water soluble vitamins and their analogues on growth of Candida albicans II. Vitamin B12, thiamine,oxythiamine, neopyrithiamine,substituted pyrimidines and thiazoles

Summary By employing wide ranges in vitamin concentrations in biotin basal mineral synthetic medium, it was demonstrated that vitamin B₁₂ markedly stimulated the growth ofCandida albicans, the organism showing a partial dependency upon this vitamin. Growth inhibition by 5-fluorouracil was reversed non-competitively by vitamin B₁₂, suggesting that B₁₂ has a role in nucleic acid biosynthesis of the organism. Thiamine was growth stimulatory, the organism being partially dependent upon this vitamin as well. Neopyrithiamine and oxythiamine were growth inhibitory in thiamine-free biotin basal mineral synthetic medium although the halves of each inhibitor compound were non-inhibitory. Neopyrithiamine inhibition was reversed by intact thiamine but not by pyrimidine thiamine or thiazole thiamine; while oxythiamine inhibition was reversed by thiamine and pyrimidine thiamine but not by thiazole thiamine, the inference being drawn that oxythiamine selectively blocks utilization of pyrimidine thiamine. Twenty-seven different substituted pyrimidines, thiazoles and related thiamine compounds were all utilizable byC. albicans in thiamine-free basal synthetic mineral medium, the organism presumably synthesizing thiamine when presented with the constituent parts of these thiamine analogues. Substitution of sulfur of the thiazole ring with oxygen, as in -methyloxazolium, failed to produce an inhibitory compound forC. albicans. Acetylthiamine, allithiamine, cocarboxylase, tetrahydrothiamine and dihydrothiamine were equally as growth stimulatory as thiamine.     

Observations on the biosynthesis of thiamine in yeast

1. Methods are described for the isolation of radioactively pure thiamine from yeast and its degradation on a small scale to its cyclic components. 2. A degradation of the pyrimidine ring and a thin-layer method for the separation of thiamine, its derivatives and pyrimidine and thiazole residues are described. 3. [(14)C]Formate is more effectively incorporated into the pyrimidine residue than into the thiazole residue, whereas the reverse is true with l-[Me-(14)C]methionine. 4. Experiments with [Me-(14)C,(35)S]methionine demonstrate that methionine provides an intact unit for the biosynthesis of the thiazole ring. 5. [6-(14)C]Orotic acid is insignificantly incorporated into the pyrimidine residue of thiamine. 6. Experiments with [1-(14)C]- and [2-(14)C]-acetate indicate that it is incorporated as a unit into the thiazole residue, but that only C-2 is incorporated into the pyrimidine residue. 7. l-[U-(14)C]Alanine is also effectively incorporated into the thiazole residue. 8. These results are discussed in relation to possible pathways of biosynthesis of the two ring components of the thiamine molecule.     

Anticancer profile of newly synthesized BRAF inhibitors possess 5-(pyrimidin-4-yl)imidazo[2,1-b]thiazole scaffold

In this work, a new series of imidazo[2,1-b]thiazole was designed and synthesized. The new compounds are having 3-fluorophenyl at position 6 of imidazo[2,1-b]thiazole and pyrimidine ring at position 5. The pyrimidine ring containing either amide or sulphonamide moiety attached to a linker (ethyl or propyl) at position 2 of the pyrimidine ring. The final compounds were selected by NCI for in vitro cytotoxicity screening. Most derivatives showed cytotoxic activity against colon cancer and melanoma cell lines. In addition, IC₅₀s of the target compounds were determined over A375 and SK-MEL-28 cell lines using sorafenib as positive control. Compounds12b, 12c, 12e, 12f, 15a, 15d, 15f, 14g and 15h exhibited superior activity when compared to sorafenib. The most potent compounds were tested against wild type BRAF, v600e BRAF, and CRAF. Compound 15h exhibited a potential inhibitory effect against^V600EBRAF (IC₅₀?=?9.3?nM).     

GENETIC BLOCKS IN THE THIAMINE SYNTHESIS OF THE ANGIOSPERM ARABIDOPSIS,,

Five thiamine-requiring mutants were obtained at two loci. Two are blocked in the synthesis of the pyrimidine part of the vitamin, the other three have lost the ability to make the thiazole moiety. None of the tested substances suggested as possible or likely precursors of the pyrimidine or the thiazole components of thiamine displayed any activity in the mutants. These conditional lethals responded to remarkably small supplements of thiamine. The pyrimidine-requiring mutants utilized to some extent the anti-vitamin neopyrithiamine. The thiazole-less mutants grew on basal media supplemented only with the analog, oxythiamine. Thiamine deficiency, irrespective of the position of the genetic block in the synthesis, results in a characteristic anomaly of pigmentation. The position of the py locus in the second linkage group has been determined. Allelic complementation has not been detected. The frequency of mutations affecting thiamine synthesis appears about the same in Arabidopsis as in fungi. The general frequency of reparable genetic lesions is, however, one to two orders of magnitude lower in Arabidopsis than that in fungi or bacteria.     

Synthesis and Studies of 3′-C-Trifluoromethyl-β-D-ribonucleosides Bearing the Five Naturally Occurring Nucleic Acid Bases

Abstract

3′-C-Trifluoromethyl-β-D-ribonucleoside derivatives bearing the five naturally occurring nucleic acid bases have been synthesized. All these derivatives were prepared by glycosylation reactions of purine and pyrimidine bases with a suitable peracylated 3-C-trifluoromethyl ribofuranose precursor. After deprotection, the resulting title nucleoside analogues were tested for their inhibitory properties against the replication of HIV, HBV and several RNA viruses. However, none of these compounds showed significant antiviral activity.     

ORCHID MYCORRHIZA: VITAMIN PRODUCTION AND REQUIREMENTS BY THE SYMBIONTS

A Rhizoctonia species isolated from Cymbidium has been cultured successfully on a defined medium consisting of minerals, sugar, thiamine, and folic acid. Thiamine can be replaced by its thiazole component, which is probably produced by germinating orchids. The fungus apparently produces the pyrimidine moiety of thiamine, a compound which may enhance growth of certain orchid seedlings. Niacin is also provided by the fungus. Para-aminobenzoic acid, a constituent of folic acid, produced and released by orchid seeds, can satisfy the vitamin requirement of the fungus. These findings point to the possibility that orchids and their fungi may have coevolved with respect to vitamin requirements. The data also suggest that exchanges of vitamins or their components between orchids and endophytes are important aspects of the symbiotic relationship.     

Differential susceptibility ofSclerotium cepivorum Berk. to some synthesized visnagin sulfonamide derivates

Twenty-five visnagin sulfonamide derivatives were testedin vitro against sclerotial germination, growth and cellulolytic activity ofSclerotium cepivorum Berk. The effectiveness of the derivatives depends on the concentration and the substituent introduced to the title compounds. The introduction of SO₂Cl₂ to C₉ of visnagin induced high toxicity than introducing SO₂NH₂. Compounds with sulfonyl piperidine or sulfonyl morpholine gave small toxicity only at 30 and 75 μg cm⁻³. Addition of N-aryl ring to visnagin-9-sulfonamide rendered the title compound to be more toxic. The substitution of the N-aryl ring bym-CH₃,m-Cl orp-Cl enhanced the toxicity, while its substitution witho-CH₃,p-CH₃,p-Br,o-OCH₃ orm-OCH₃ caused a drop in the toxicity as compared to compounds with unsubstituted aryl ring. Visnagin sulfonamide derivatives having azole rings were strongly inhibitory for sclerotial germination, growth, sclerotial formation and cellulolytic activity, even when applied at 4 μg cm⁻³. The most toxic one was that having dimethyl isoxazole. The cleavage of γ-pyrone ring led to a decline in the toxicity as compared with the other sulfonamide derivatives. Communicated by J. ŠPAK     

Nucleotide,nucleoside and base nutritional requirements of the mosquito Culex pipiens

Nucleic acid subcomponents needed to satisfy the dietary nucleic acid requirement of Culex pipiens were studied in growth experiments using synthetic media in which nucleosides, bases and alternative nucleotides were variously substituted in mixtures of 3 nucleotides (adenylic acid, thymidylic acid, and either cytidylic or uridylic acid) previously shown to be adequate replacements for whole nucleic acid. Any or all 3 nucleotides could be replaced by corresponding nucleosides without adverse effect, except that adenosine substitution moderately delayed pupation. All base substitutions were unsatisfactory: substitution of thymine for thymidylic acid allowed development to the adult stage but at a greatly reduced rate; single substitution of adenine, cytosine or uracil for the corresponding nucleotides allowed scarcely more development than in the total absence of nucleic acid derivatives. Inosinic acid or inosine were adequate substitutes for adenylic acid, but orotic acid or orotidine were ineffective in place of the pyrimidine ribonucleotides, cytidylic or uridylic acids. Deoxyadenylic acid could take the place of adenylic acid, though inefficiently, but deoxycytidylic and deoxyuridylic acids were very poor replacements for the corresponding ribonucleotides. The minimal required nucleic acid derivatives thus appear to be a purine ribonucleotide (adenylic or inosinic acids), a pyrimidine ribonucleoside (either uridine or cytidine), and the pyrimidine deoxyribonucleoside, thymidine.     

Studies on non-steroidal inhibitors of aromatase enzyme; 4-(aryl/heteroaryl)-2-(pyrimidin-2-yl)thiazole derivatives

Steroidal and non-steroidal aromatase inhibitors target the suppression of estrogen biosynthesis in the treatment of breast cancer. Researchers have increasingly focused on developing non-steroidal derivatives for their potential clinical use avoiding steroidal side-effects.Non-steroidal derivatives generally have planar aromatic structures attached to the azole ring system. One part of this ring system comprises functional groups that inhibit aromatization through the coordination of the haem group of the aromatase enzyme. Replacement of the triazole ring system and development of aromatic/cyclic structures of the side chain can increase selectivity over aromatase enzyme inhibition.In this study, 4-(aryl/heteroaryl)-2-(pyrimidin-2-yl)thiazole derivatives were synthesized and physical analyses and structural determination studies were performed. The IC₅₀ values were determined by a fluorescence-based aromatase inhibition assay and compound 1 (4-(2-hydroxyphenyl)-2-(pyrimidine-2-yl)thiazole) were found potent inhibitor of enzyme (IC₅₀:0.42?nM). Then, their antiproliferative activity over MCF-7 and HEK-293 cell lines was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Compounds 1, 7, 8, 13, 15, 18, 21 were active against MCF-7 breast cancer cells. Lastly, a series of docking experiments were undertaken to analyze the crystal structure of human placental aromatase and identify the possible interactions between the most active structure and the active site.     

Pyrimidine metabolism during somatic embryo development in white spruce (Picea glauca)

Pyrimidine metabolism was investigated at various stages ofsomatic embryo development of white spruce (Picea glauca). The contribution of thede novo and the salvage pathways of pyrimidine biosynthesis to nucleotide and nucleic acid formation and the catabolism of pyrimidine was estimated by the exogenously supplied [6-¹⁴C]orotic acid, an intermediate of thede novo pathway, and with [2-¹⁴C]uridine and [2-¹⁴C]uracil, substrates of the salvage pathways. Thede novo pathway was very active throughout embryo development. More than 80 percnt; of [6-¹⁴C]orotic acid taken up by the tissue was utilized for nucleotide and nucleic acid synthesis in all stages of this process. The salvage pathways of uridine and uracil were also operative. Relatively high nucleic acid biosynthesis from uridine was observed, whereas the contribution of uracil salvage to the pyrimidine nucleotide and nucleic acid synthesis was extremely limited. A large proportion of uracil was degraded as ¹⁴CO₂, probably via β-ureidopropionate. Among the enzymes of pyrimidine metabolism, orotate phosphoribosyltransferase was high during the initial phases of embryo development, after which it gradually declined. Uridine kinase, responsible for the salvage of uridine, showed an opposite pattern, since its activity increased as embryos developed. Low activities of uracil phosphoribosyltransferase and non-specific nucleoside phosphotransferase were also detected throughout the developmental period. These results suggest that the flux of thede novo and salvage pathways of pyrimidine nucleotide biosynthesisin vivo is roughly controlled by the amount of these enzymes. However, changing patterns of enzyme activity during embryo development that were measuredin vitro did not exactly correlate with the flux estimated by the radioactive precursors. Therefore, other fine control mechanisms, such as the fluctuation of levels of substrates and/or effectors may also participate to the real control of pyrimidine metabolism during white spruce somatic embryo development.     

Oxidation and ring cleavage of dibenzofuran by the filamentous fungus <Emphasis Type="Italic">Paecilomyces lilacinus</Emphasis>

The ability of the imperfect soil fungus Paecilomyces lilacinus to transform the environmental pollutant dibenzofuran was investigated. Transformation of dibenzofuran and related derivatives lead to 14 products, which were identified by UV spectroscopy, mass spectrometry, and proton nuclear magnetic resonance spectroscopy. Biotransformation was initiated by two separate hydroxylation steps, leading to the accumulation of 4-monohydroxylated and 4-dihydroxylateddibenzofurans. Hydroxylation at both aromatic rings produced 2,7-dihydroxydibenzofuran, 3,7-dihydroxydibenzofuran, and 2,8-dihydroxydibenzofuran. Further oxidation yields ring cleavage of dibenzofuran, which has not been described before for filamentous fungi. The ring fission products were identified as benzo[b]furo[3,2-d]-2-pyrone-6-carboxylic acid and [2-(1-carboxy-methylidene)-benzofuran-3-ylidene]-hydroxy-acetic acid and its derivatives hydroxylated at carbon 7 and 8 at the non-cleaved ring. Other metabolites were riboside-conjugates of 2-hydroxydibenzofuran and 3-hydroxydibenzofuran. The results showed that P. lilacinus transforms the hydrophobic compound dibenzofuran by phase I/phase II reactions to produce hydroxylated products and excretable sugar conjugates.     

2,4-Diacetylphloroglucinol suppresses zoosporogenesis and impairs motility of Peronosporomycete zoospores

2,4-Diacetylphloroglucinol (DAPG) produced by Pseudomonas fluorescens, shows toxicity to many microorganisms including fungi, bacteria, and peronosporomycetes. Zoosporogenesis and motility of zoospores are critical for a complete disease cycle and pathogenicity of the peronosporomycete phytopathogens. The aim of this study was to test the effects of DAPG and its derivatives on zoosporogenesis and motility of zoospores of a downy mildew pathogen, Plasmopara viticola, and a damping-off pathogen, Aphanomyces cochlioides. In both cases, DAPG inhibited zoosporogenesis (5 μg/ml) and the motility of zoospores (10 μg/ml) in a dose-dependent manner. Generally, zoospores became immotile shortly after exposure to DAPG followed by lysis. However, a fraction of DAPG treated A. cochlioides zoospores formed round cystospores instead of lysis and then germinated with excessively-branched germ tubes. All derivatives of DAPG had similar inhibitory activities but at varying doses. Among them, 2,4-dipropylphloroglucinol exerted the highest inhibitory activity against both zoosporogenesis and motility of zoospores. This revealed that the degree of hydrogen atoms substitution in the benzene ring by acyl groups and the length of substituted acyl groups were related to the level of bioactivity. This is the first report of inhibitory activities of DAPG and its derivatives against zoosporogenesis and motility of zoospores of two important peronosporomycete phytopathogens.     

Biosynthesis of thiamin in Bacillus subtilis. Isolation of mutants accumulating 4-amino-5-hydroxymethyl-2-methylpyrimidine phosphate.

Thiamin-deficient mutants of Bacillus subtilis were characterized by their growth responses to the pyrimidine and thiazole moieties of the vitamin molecule and by cross-feeding tests. All mutants growing on the thiazole moiety and all mutants with an absolute requirement for thiamin fed all those growing on the pyrimidine moiety. No other cross-feeding effects were observed. From the culture fluid of a mutant growing on the thiazole moiety, two compounds were isolated which supported growth of mutants requiring the pyrimidine moiety. These compounds were identified by chromatographic, bioautographic and spectrophotometric procedures as 4-amino-5-hydroxymethyl-2-methylpyrimidine and its monophosphate derivative.     

Synthesis and Antitumor Activity of N-Sulfonyl Derivatives of Nucleobases and Sulfonamido Nucleoside Derivatives

Abstract

The introduction of sulfonamido group on the C-2 position of pyrimidine nucleosides was achieved by ring opening of 2,2′- and 2,3′-anhydronucleosides. N-sulfonyl derivatives of nucleobases and sulfonamido derivatives of nucleosides were assayed for in vitro antitumor activity.     

Partial Asymmetric Synthesis in α-Alkylation of Enamine with Electrophilic Olefin

About 30 dipeptides and some tripeptides were led to new benzimidazole derivatives by incorporating their carboxyl groups into benzimidazole ring by the reaction with o-phenylenediamine. The ring closure to benzimidazole was well achieved by heating mildly at a moderate temperature in acetic acid.

Some benzimidazole derivatives of peptides had remarkable phytotoxicities.     

Adaptive Biosynthesis of Thiazole and Bypassing of the Thiazole Requirement in Exobasidium by Acetyl Phosphate, Tween and Ethanol

The tested strain of Exobasidium vaccinii (Fuck.) Woron. required thiamine or its two moieties, thiazole and pyrimidine, for good growth. When grown on pyrimidine medium, rapid growth or thiazole synthesis appears after 600–1000 hours. As methionine, a supposed precursor in the biosynthesis of thiazole, stimulates this inductive growth, the influence of methionine precursors or metabolites stimulating methionine synthesis were tested for their ability to shorten the lag phase of E. vaccinii cultured on a thiamine-deficient synthetic medium. None of the tested methionine precursors replaced methionine or significantly stimulated the inductive growth. This result does not exclude acetylhomoserine as a central metabolite in methionine synthesis. Acetyl phosphate, alcohols and the fatty acids of Tween, possible deliverers of the active acetyl groups, induced growth in thiamine-free media. This growth seems to be possible by utilization of the compounds through a metabolic pathway not requiring thiamine. No evidence for participation of the glyoxylate cycle was found.     

Degradation and oligomerization of syringic acid by distinctive ecological groups of fungi

Forty-four terrestrial and aeroaquatic and aquatic fungi, including fifteen species causing white-rot, four species causing brown-rot, and some species causing soft-rot of wood, were tested for their ability to degrade the monomer syringic acid, which is released during decay of angiosperm lignin. None of the white- or brown-rot species caused any detectable degradation of syringic acid under the test conditions; however, six typical white-rot fungi strongly oligomerized syringic acid, both with and without cosubstrate. The main polymerization product was identified as a 1,3-dimethylpyrogallol oligomer by¹³C-NMR. Other minor metabolic products were methylated and hydroxylated derivatives. Oligomerization depended on the presence of 1 or 2 methoxy groups in ortho position to the hydroxy group of the substrate. Among the remaining fungi,Exophiala jeanselmei, Fusarium eumartii, andPaecilomyces variotii completely and rapidly degraded syringic acid (5 g/liter) within 48 to 100 hours. A further seven species were able to degrade syringic acid to some extent when glucose was added. Methylated and demethylated metabolic intermediates were identified by GC/MS.     

Inhibition studies on a panel of human carbonic anhydrases with N1-substituted secondary sulfonamides incorporating thiazolinone or imidazolone-indole tails

Being the primary sulfonamide among the most efficient zinc binding group (ZBG) to design inhibitors for the metallo-enzymes carbonic anhydrases (CA, EC 4.2.1.1), herein, we propose an investigation on four physiologically important human (h) CAs (hCA I, II, IV, and IX) with N¹-substituted secondary sulfonamides incorporating thiazolinone or imidazolone-indole tails. The effect of the functionalisation of the sulfonamide group with five different substitution patterns, namely acetyl, pyridine, thiazole, pyrimidine, and carbamimidoyl, was evaluated in relation to the inhibition profile of the corresponding primary sulfonamide analogues. With most of these latter being nanomolar inhibitors of all four considered isoforms, a totally counterproductive effect on the inhibition potency can be ascribed to N¹-functionalisations of the ZBG primary sulfonamide structure with pyridine, thiazole, and pyrimidine moieties. On the other hand, incorporation of less hindered groups, such as sulfonylacetamides and sulfonylguanidines, maintained a certain degree of activity dependent on the tailing moiety, with K_Is spanning in the low micromolar range.     

Effect of lignin-related phenols and their methylated derivatives on the growth of eight white-rot fungi

When various lignin-related para-phenolic benzoic acids, para-phenolic cinnamic acids, para-phenolic phenylpropionic acids, the corresponding unsubstituted and 4-O-methylated derivatives, and 4-hydroxyl substituted benzaldehydes were tested on the growth of eight white-rot fungi, methylation of the 4-hydroxy substituent resulted, in most cases, in increased inhibition of fungal growth. This effect was most notable with monomethoxylated compounds. When the aromatic ring contained additional methoxyl substituents, the toxicity of the 4-O-methylated derivative was less pronounced. Marked inhibition of fungal growth was also observed with aromatic compounds lacking a para-substituent. Higher concentrations of aromatic aldehydes were manifestly more toxic than the corresponding carboxylic acid.J.A. Buswell is with the Department of Biology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong. K.-E.L. Eriksson is with the Department of Biochemistry, The University of Georgia, Athens, Georgia, 30602, USA.