Synthesis and fungicidal activity of tubulin polymerisation promoters. Part 2: pyridazines

Special tetrasubstituted pyridazines are potent fungicides by promoting the tubulin polymerisation, hereby disrupting the microtubule dynamics in the fungus. They are monocyclic analogs of similar substituted triazolopyrimidines and pyridopyrazines with the same mode of action. The fungicidal activity of these pyridazines was evaluated against the plant pathogens Botrytis cinerea (grey mould), Mycosphaerella graminicola (wheat leaf blotch) and Alternaria solani (potato and tomato early blight). Structure-activity relationship studies revealed the importance of a methyl and a chlorine substituent next to both ring nitrogen atoms and two aryl or heteroaryl groups in the other two pyridazine positions.     

Structural insights into the design of nonpeptidic isothiazolidinone-containing inhibitors of protein-tyrosine phosphatase 1B

Structural analyses of the protein-tyrosine phosphatase 1B (PTP1B) active site and inhibitor complexes have aided in optimization of a peptide inhibitor containing the novel (S)-isothiazolidinone (IZD) phosphonate mimetic. Potency and permeability were simultaneously improved by replacing the polar peptidic backbone of the inhibitor with nonpeptidic moieties. The C-terminal primary amide was replaced with a benzimidazole ring, which hydrogen bonds to the carboxylate of Asp(48), and the N terminus of the peptide was replaced with an aryl sulfonamide, which hydrogen bonds to Asp(48) and the backbone NH of Arg(47) via a water molecule. Although both substituents retain the favorable hydrogen bonding network of the peptide scaffold, their aryl rings interact weakly with the protein. The aryl ring of benzimidazole is partially solvent exposed and only participates in van der Waals interactions with Phe(182) of the flap. The aryl ring of aryl sulfonamide adopts an unexpected conformation and only participates in intramolecular pi-stacking interactions with the benzimidazole ring. These results explain the flat SAR for substitutions on both rings and the reason why unsubstituted moieties were selected as candidates. Finally, substituents ortho to the IZD heterocycle on the aryl ring of the IZD-phenyl moiety bind in a small narrow site adjacent to the primary phosphate binding pocket. The crystal structure of an o-chloro derivative reveals that chlorine interacts extensively with residues in the small site. The structural insights that have led to the discovery of potent benzimidazole aryl sulfonamide o-substituted derivatives are discussed in detail.     

The causes of the biological action of electrochemically activated solutions by changes in the growth of Escherichia coli cells

To study the causes of the biological effect of electrochemically activated solutions, nutrient growth media M 9 were prepared using catholyte and anolyte solutions containing separate components of the nutrient medium, such as distilled water, phosphate buffer, phosphate buffer with chlorides (NaCl, NH4Cl), and chlorides. The biological activity of different nutrient media was assessed by a comparison with the stimulation or inhibition of the growth of Escherichia coli cells in the catholyte and anolyte of the complete nutrient medium M 9. It was shown that medium M 9 prepared on the catholytes of different initial solutions acquired the stimulating properties only if the initial solution contained salts containing chlorine. The stimulating effect of the initial solution was 18-24%. Electrochemical treatment of solutions containing no chlorides (distilled water, phosphate buffer) and subsequent addition of the components of nutrient medium to exposed solutions had neither a stimulating nor the inhibiting effect on cell growth. The cultivation of cells in a nutrient medium based on the catholyte of preliminarily treated hydrochloric acid showed that it is the presence of chlorine ions in solution during electrolysis that causes the stimulating effect of the nutrient medium based on the catholyte. The formation of oxidizers and the inhibitory effect of the anolyte described previously was also observed if the solution contained chlorine ions during electrolysis. Possible mechanisms of the biological effect of catholytes containing chlorides during electrolysis were discussed.     

Studies on plant growth-regulating substances XXV. The plant growth-regulating activity of cinnamic acids

Effects of ring substitution on the plant growth-regulating activities of trans- and cis-cinnamic acids have been investigated in the wheat cylinder, pea segment and pea curvature tests. Most of the cis- acids were shown to be active. Substitution of fluorine, chlorine or bromine into the ring of cis-cinnamic acid in most cases increased the activity. The results are discussed in relation to mode of action and chemical structure/biological activity relationships: 4-chlorobenzoic acid is shown to act as a competitive antagonist towards 4-chloro-cis-cinnamic acid.     

Biological evaluation of newly synthesized aryl(thio)carbamoyl derivatives of 1- and 1-(2-aminoethyl)-piperazines

Twelve 1-methyl and acetyl-4-substituted piperazines, evaluated as potential herbicides and plant growth regulators, were synthesized by condensation of 1-methyl-piperazine and 1-[2-(acetylamino)ethyl]-4-acetyl-piperazine with the corresponding aryliso(thio)cyanates. These piperazines, which incorporate a piperazine ring and aryl(thio)carbamoyl groups connected directly or through an ethylene group, are new chemical families of herbicides and cytokinin mimics. Structure–activity relationships for the screened compounds were evaluated and discussed. The greatest herbicidal activity against Triticum aestivum was observed with compounds that contained the three structural elements: piperazine ring, ethylene group and 4-fluorophenylcarbamoyl group. Compounds having a combination of two active moieties–piperazine ring and 4-halogenophenylthiocarbamoyl group, also showed high herbicidal activity against T. aestivum. The compound, in which the un-substituted phenylcarbamoyl group was directly connected to the piperazine ring, showed cytokinin-like activity and significantly stimulated betacyanin synthesis in Amaranthus caudatus.     

Halogenated diarylacetylenes repress c-myc expression in cancer cells

Halogenated diarylacetylenes that possess fluorine or chlorine substituents in one aryl ring and N-methylamino or N,N-dimethylamino in the other aryl ring inhibit the proliferation of LS174T colon cancer cells through the repression of c-myc expression and induction of the cyclin-dependent kinase inhibitor-1 (i.e., p21(Wif1/Cip1)) and represent potentially useful antineoplastic agents.     

Estrogen receptor affinity and effects on MCF-7 cell growth of triarylethylene carboxylic acids related to tamoxifen.

The estrogen receptor binding, and growth suppressant and stimulating effects in MCF-7 human breast cancer cells, of four structural variants of the triarylethylene antiestrogen tamoxifen (1) were studied. In these analogs, the dialkylaminoethoxy side chain of 1 was replaced by carboxylic acid or oxyacetic acid substituents. The presence of a p-hydroxy group in the ring geminal to the one bearing the side chain resulted in ligands with estrogen receptor affinities greater than that of 1 but less than that of estradiol. Compared to 1, none of the test compounds were effective suppressants of cell growth. To the contrary, the phenolic oxyacetic acid analog effectively reversed the growth suppressive effect of 1. Also, it was as effective as estradiol, though less potent, in stimulating growth of cells grown in estrogen depleted medium, suggestive of full estrogen agonist activity. Its carboxylic acid counterpart had little or no effect on proliferation. Because the phenolic oxyacetic acid is a metabolite of 1 in animals, its estrogenicity may have therapeutic implications of concern, depending on the extent to which it is formed and distributed in tissues of patients receiving 1.     

Synthesis and fungitoxicity of some pyrimidine derivatives

A series of 12 pyrimidine derivatives were prepared and testedin vitro against growth, sporulation and nucleic acid content ofFusarium oxysporum f. sp.lycopersici andHelminthosporium oryzae. Introduction of a thiazole ring together with two aryl groups into 2-aminopyrimidine brought about drastic toxicity for both fungi. Pyrimidine derivatives with aryl groups alone were less toxic. Nitro groups were found to enhance the toxicity of the pyrimidine derivatives especially when substituted in the ortho-position of the aryl groups. Inhibition of nucleic acid synthesis of both fungi was attributed mainly to the presence of the thiazole ring.     

A Structure-Activity Study with Aryl Acylamidases

We examined the relationship between chemical structure and biodegradability of acylanilide herbicides by using a set of model compounds. Four bacterial isolates (one gram-negative and three gram-positive) that grew on acetanilide were used. These soil isolates cleaved the amide bond of acetanilide via an aryl acylamidase reaction, producing aniline and the organic acid acetate. A series of acetanilide analogs with alkyl substitutions on the nitrogen atom or the aromatic ring were tested for their ability to induce aryl acylamidase activity and act as substrates for the enzyme. The substrate range, in general, was limited to those analogs not disubstituted in the ortho position of the benzene ring or which did not contain an alkyl group on the nitrogen atom. These same N-substituted compounds did not induce enzyme activity either, whereas the ortho-substituted compounds could in some cases.     

Isolation and identification of prostaglandin I 2 stimulating factor from human plasma

To isolate and identify the plasma factor which stimulates prostaglandin I 2 production by rat aortic ring, a human plasma fraction which showed a major stimulating activity on prostaglandin I 2 production was purified by ultrafiltrate, Sephadex G-10 gel filtration and QAE-Sephadex column chromatography. The purified plasma factor was identified as uric acid by its ultraviolet and infrared absorption spectroscopy, and 1H nmr and 13C nmr spectroscopy. The stimulating activity of the purified plasma factor and that of authentic uric acid coincided with each other. The stimulating potency of uric acid at its physiological concentration in human plasma (about 50 micrograms/ml) was half of the deproteinized human plasma, and was about 30 fold stronger than that of L-tryptophan, a cofactor of prostaglandin hydroperoxidase.     

Associative diazotrophs of pearl millet (Pennisetum glaucum) from semi arid region--isolation and characterization

Diversity of the native diazotrophs associated with the rhizosphere of pearl millet (P. glaucumn), grown in nutritionally poor soils of semi-arid regions was studied with a view to isolate effective nitrogen fixing and plant growth stimulating bacteria with root associative characteristics. The native population varied from 10(3)-10(4) g(-1) of rhizosphere soil after 40 d growth and belonged to genera Azospirillum, Azotobacter and Klebsiella. Another non-diazotrophic root associative group was Pseudomonas sp., which also produced IAA and enhanced plant growth. Some of these rhizobacteria showed high in vitro acetylene reduction activity along with production of indole acetic acid. Out of 11 selected diazotrophs used as seed inoculants, M10B (Azospirillum sp.), M11E (Azotobacter sp.) and M12D4 (Klebsiella sp.) resulted in significant increase in total root and shoot nitrogen at 45 and 60 days of plant growth under pot culture conditions.     

Effects of water stress, organic amendment and mycorrhizal inoculation on soil microbial community structure and activity during the establishment of two heavy metal-tolerant native plant species

Our aim was to examine the effect of water stress on plant growth and development of two native plant species (Tetraclinis articulata and Crithmum maritimum) and on microbial community composition and activity in the rhizosphere soil, following the addition of an organic amendment, namely sugar beet residue (SBR), and/or the inoculation with an arbuscular mycorrhizal (AM) fungus, namely Glomus mosseae, in a non-sterile heavy metal-polluted soil. The AM inoculation did not have any significant effect on plant growth of both species. In T. articulata, SBR increased shoot growth, foliar P, total phospholipid fatty acids (PLFA), fungi-related PLFA, AM fungi-related neutral lipid fatty acid, bacterial gram-positive/gram-negative PLFA ratio and the β-glucosidase and dehydrogenase activities. SBR and AM inoculation increased phosphatase activity in T. articulata plants grown under drought conditions. In both plants, there was a synergistic effect between AM inoculation and SBR on mycorrhizal colonisation under drought conditions. In C. maritimum, the increase produced by the SBR on total amounts of PLFA, bacterial gram-positive-related PLFA and bacterial gram-negative-related PLFA was considerably higher under drought conditions. Our results suggest that the effectiveness of the amendment with regard to stimulating microbial communities and plant growth was largely limited by drought, particularly for plant species with a low degree of mycorrhizal colonisation.     

Lead optimization of methionine aminopeptidase-2 (MetAP2) inhibitors containing sulfonamides of 5,6-disubstituted anthranilic acids

A series of aryl sulfonamides of 5,6-disubstituted anthranilic acids were identified as potent inhibitors of methionine aminopeptidase-2 (MetAP2). Small alkyl groups and 3-furyl were tolerated at the 5-position of anthranilic acid, while -OCH(3), CH(3), and Cl were found optimal for the 6-position. Placement of 2-aminoethoxy group at the 6-position enabled interaction with the second Mn(2+) but did not result in enhancement in potency. Introduction of a tertiary amino moiety at the ortho-position of the sulfonyl phenyl ring gave reduced protein binding and improved cellular activity, but led to lower oral bioavailability.     

Isolation and identification of prostaglandin I 2 stimulating factor from human plasma

To isolate and identify the plasma factor which stimulates prostaglandin I 2 production by rat aortic ring, a human plasma fraction which showed a major stimulating activity on prostaglandin I 2 production was purified by ultrafiltrate, Sephadex G-10 gel filtration and QAE-Sephadex column chromatography. The purified plasma factor was identified as acid by its ultraviolet and infrared absorption spectroscopy, and ¹H nmr and ¹³C nmr spectroscopy. The stimulating activity of the purified plasma factor and that of authentic uric acid coincided with each other. The stimulating potency of uric acid at its physiological concentration in human plasma (about 50 μg/ml) was half of the deproteinized human plasma, and was about 30 fold stronger than that of L-tryptophan, a cofactor of prostaglandin hyperoxidase.     

Synthesis of structurally diverse benzosuberene analogues and their biological evaluation as anti-cancer agents

Diversely functionalized, fused aryl–alkyl ring systems hold a prominent position as well-established molecular frameworks for a variety of anti-cancer agents. The benzosuberene (6,7 fused, also referred to as dihydro-5H-benzo[7]annulene and benzocycloheptene) ring system has emerged as a valuable molecular core component for the development of inhibitors of tubulin assembly, which function as antiproliferative anti-cancer agents and, in certain cases, as vascular disrupting agents (VDAs). Both a phenolic-based analogue (known as KGP18, compound 39) and its corresponding amine-based congener (referred to as KGP156, compound 45), which demonstrate strong inhibition of tubulin assembly (low micromolar range) and potent cytotoxicity (picomolar range for KGP18 and nanomolar range for KGP156) are noteworthy examples of such benzosuberene-based compounds. In order to extend the structure–activity relationship (SAR) knowledge base related to benzosuberene anti-cancer agents, a series of eleven analogues (including KGP18) were prepared in which the methoxylation pattern on the pendant aryl ring as well as functional group incorporation on the fused aryl ring were varied. The synthetic approach to these compounds featured a sequential Wittig olefination, reduction, Eaton’s reagent-mediated cyclization strategy to achieve the core benzosuberone intermediate, and represented a higher-yielding synthesis of KGP18 (which we prepared previously through a ring-expansion strategy). Incorporation of a fluorine or chlorine atom at the 1-position of the fused aryl ring or replacement of one of the methoxy groups with hydrogen (on the pendant aryl ring of KGP18) led to benzosuberene analogues that were both strongly inhibitory against tubulin assembly (IC₅₀ approximately 1.0 μM) and strongly cytotoxic against selected human cancer cell lines (for example, GI₅₀ = 5.47 nM against NCI-H460 cells with fluoro-benzosuberene analogue 37). A water-soluble phosphate prodrug salt of KGP18 (referred to as KGP265, compound 44) and a water-soluble serinamide salt (compound 48) of KGP156 were also synthesized and evaluated in this study.     

Molecular design, chemical synthesis, and biological evaluation of '4-1' pentacyclic aryl/heteroaryl-imidazonaphthalimides

A novel series of '4-1' pentacyclic naphthalimides, where the chromophore consists of a naphthalimide moiety, fused to an imidazole ring containing an unfused aryl or heteroaryl ring, were synthesized and evaluated for in vitro antitumor activity. In general, the new derivatives showed an improved cytotoxic activity over amonafide. DNA binding experiments supported that this class of compounds behaves as effective DNA-intercalating agents.     

Correlation between Plant Growth Regulator Release Rate and Bioactivity for the Series of Newly Synthesized Phytoactive Polymers

Phytoactive polymers are high molecular weight systems in which a plant growth regulator (PGR) unit is attached to the polymeric chain by a hydrolyzable chemical bond. The release rate of the PGR is linked to the biological activity of the phytoactive polymer and can be controlled by properties inherent in the whole macromolecular system. In this study the correlation of biological activity and plant growth regulator hydrolytic release rate was investigated for the series of newly synthesized 2,4-dichlorophenoxyacetic acid (2,4-D) polymeric esters. The polymers synthesized differ in their molecular weight, side group structure, and 2,4-D residue content. The influence of these polymer characteristics on the 2,4-D hydrolytic release was investigated, and it was demonstrated that hydrolysis rate substantially depends on the polymer molecular weight, side group structure, and 2,4-D residue content. It was also demonstrated that phytoactive polymer bioactivity depends on the hydrolysis rate of the polymers, and in dependence of this parameter can provide stimulating or inhibiting activity. Biological activity was illustrated by the elongation of wheat and barley coleoptiles.     

Modified 2,4-diaryl-5H-indeno[1,2-b]pyridines with hydroxyl and chlorine moiety: Synthesis,anticancer activity,and structure–activity relationship study

As a part of ongoing studies in developing novel anticancer agents, a series of modified 2,4-diaryl-5H-indeno[1,2-b]pyridines were designed, and synthesized by introducing hydroxyl and chlorine moieties. They were evaluated for topoisomerase inhibitory activity and cytotoxicity against HCT15, T47D, and HeLa cancer cell lines. This modification allowed us to demonstrate structure–activity relationship (SAR) study with respect to the non-substituted 2,4-diaryl-5H-indeno[1,2-b]pyridines. Compounds (2, 3, 4, 5, 8, and 9) with meta or para hydroxyl group on 2 or 4-phenyl ring have enhanced topo I and II inhibitory activity and cytotoxicity. However, additional substitution of chlorine group on furyl or thienyl ring (11, 12, 14, 16–18) generally reduced topo I and II inhibitory activity but improved cytotoxicity. The observation of cytotoxic properties and SAR study according to the position of hydroxyl and chlorine group will provide valuable insight for further study of development of novel anticancer agents with related scaffolds.     

Design, synthesis, and biological evaluation of 4-arylmethyl-1-phenylpyrazole and 4-aryloxy-1-phenylpyrazole derivatives as novel androgen receptor antagonists

A series of 4-arylmethyl-1-phenylpyrazole and 4-aryloxy-1-phenylpyrazole compounds B were designed, synthesized, and evaluated for their potential as new-generation androgen receptor (AR) antagonists therapeutically effective against castration-resistant prostate cancer (CRPC). Introduction of a bulky amide substituent (R(2)) to the terminal aryl ring of the 4-arylmethyl group favored the reduction of agonistic activity and improved the pharmacokinetic (PK) properties. Similarly, introduction of a bulky substituent in the 4-aryloxy derivatives also resulted in improved PK properties. Compounds 28 h and 44b exhibited potent antitumor effects against a CRPC model of LNCaP-hr cell line in a mouse xenograft model. On the contrary, bicalutamide showed only partial suppression of tumor growth. These results suggest that the novel pyrazole derivatives are new-generation AR antagonists, different from the 'first-generation' antagonists such as bicalutamide in a CRPC treatment model.     

Comparative Effects of Ancymidol and Its Analogs on Growth of Peas and Ent-Kaurene Oxidation in Cell-Free Extracts of Immature Marah macrocarpus Endosperm

The plant growth retardant alpha-cyclopropyl-alpha-(4-methyoxyphenyl)-5-pyrimidine methyl alcohol (ancymidol) and a series of analogs of this substance in which one or more of the substituents were varied were tested for their comparative biological activity. The compounds were tested as inhibitors of internode elongation in peas and as inhibitors of the oxidation of ent-kaurene catalyzed by microsomal preparations from the liquid endosperm of Marah macrocarpus seeds. The relative effectiveness of a substance was generally the same as an inhibitor of the two processes. Ancymidol was the most effective. Substitution of the alcohol group of ancymidol by either methoxy or hydrogen groups reduced the activity only slightly. Substitution of the cyclopropyl group by an isopropyl moiety also had little effect on the activity. However, substitution of the cyclopropyl group with a phenyl or other aryl substituent greatly reduced the effectiveness of the analog as an inhibitor. Replacement of the 4-methoxyphenyl substituent with a similar substituent such as 4-chlorophenyl had little effect on activity, but replacement with a 2-methoxyphenyl group greatly reduced activity. Analogs in which the pyrimidyl moiety of ancymidol was modified were inactive in whole plants, but moderately active in the cell-free ent-kaurene oxidation system. The application of gibberellic acid can overcome the growth inhibitions due to treatment of the test plants with 10(-5)m or lower concentrations of the inhibitors. However, the inhibitory effects of 10(-4)m or higher concentrations of inhibitors on test plants were not overcome by the applications of exogenous gibberellic acid. These results support the idea that the effects of low concentrations of these substances on plant growth are primarily a consequence of their ability to inhibit ent-kaurene oxidation and gibberellin biosynthesis. Other modes of inhibition may operate at higher inhibitor concentrations.