Synthesis and cytotoxic activity of new alkyl[3-(2-chloroethyl)ureido]benzene derivatives

Several alkyl[3-(2-chloroethyl)ureido] (CEU) benzene derivatives were prepared as potential anticancer agents. These new compounds were readily prepared in good yields by addition of anilines to 2-chloroethylisocyanate. Their cytotoxic activity was evaluated on human breast cancer (MDA-MB-231), human colon adenocarcinoma (LoVo) and mouse lymphocytic leukemia (P388D₁) tumor cell lines. Several new CEUs were significantly more cytotoxic than the nitrogen mustard chlorambucil. The biological activity of these aromatic urea derivatives seems to be related to the nature and position of the alkyl substituents on the aromatic ring. Substitution by branched alkyl groups on position 4 of the aromatic ring led to cytotoxic molecules which are up to 5 times more potent than the standard chlorambucil.     

Structure-activity relationships of quaternary protoberberine alkaloids having an antimalarial activity

Seventeen quaternary protoberberine alkaloids related to berberine 1 were tested for antimalarial activity in vitro against Plasmodium falciparum and structure-activity relationships are proposed. The activity of the protoberberine alkaloids was influenced by the type of the oxygen substituents on rings A, C and D and the position of the oxygen functions on ring D. The position of the oxygen functions on ring D and the type of the oxygen substituents at the C-13 position (ring C) strongly influenced the activity. Shifting the oxygen functions at C-9 and C-10 to C-10 and C-11 on ring D resulted in a significant increase in the activity. Compounds bearing a methylenedioxy function at C-2 and C-3 (ring A) or C-9 and C-10 (ring D) showed higher activity than those which have methoxy groups at the same positions. Introduction of a methoxy group into the C-1 position (ring A) decreased the activity. Replacement of a hydroxy group at C-2 or C-3 (ring A) by a methoxy group led to a reduction in the activity. Displacement of a hydroxy function at C-13 (ring C) by the oxygen substituents such as OMe, OEt, OCOOEt, and OCON(Me)₂ reduced the activity. In the same replacement at C-9 (ring D), the activity depended upon the type of the oxygen function. Six protoberberines displayed more potent activity than berberine 1. The activity decreased in the order: 10, 11, 17 and 18 > 7 and 8 > 1.     

Synthesis, spectral studies and antiamoebic activity of new 1-N-substituted thiocarbamoyl-3-phenyl-2-pyrazolines

Thirty new pyrazoline derivatives were synthesized by cyclization of Mannich bases with thiosemicarbazides being substituted by different cyclic and aromatic amines. The structures of the compounds were elucidated by elemental analyses, UV, IR, (1)H and (13)C NMR and ESI-MS spectral data. The in vitro antiamoebic activity was evaluated against Entamoeba histolytica in comparison with metronidazole used as reference substance. Out of the 30 compounds screened for antiamoebic activity, 10 (5, 6, 15, 18, 25-30) were found to be better inhibitors of E. histolytica since they showed lesser IC(50) values than metronidazole. The preliminary results indicated that the presence of 3-chloro or 3-bromo substituent on the phenyl ring at position 3 of the pyrazoline ring enhanced the antiamoebic activity as compared to unsubstituted phenyl ring. The study suggests that the preliminary activity of these compounds may further be explored for the development of new targets for amoebiasis.     

Xenoestrogenic gene expression: structural features of active polycyclic aromatic hydrocarbons

Estrogenicity was assessed using the Saccharomyces cerevisiae-based Lac-Z reporter assay and was reported as the logarithm of the inverse of the 50% molar beta-galactosidase activity (log[EC50(-1)]). In an effort to quantify the relationship between molecular structure of polycyclic aromatic hydrocarbons (PAHs) and estrogenic gene expression, a series of PAHs were evaluated. With noted exceptions, the results of these studies indicate that the initial two-dimensional structural warning for estrogenicity, the superpositioning of a hydroxylated aromatic system on the phenolic A-ring of 17-beta-estradiol, can be extended to the PAHs. This two-dimensional-alignment criterion correctly identified estrogenicity of 22 of the 29 PAHs evaluated. Moreover, the estrogenic potency of these compounds was directly related to the size of the hydrophobic backbone. The seven compounds classified incorrectly by this structural feature were either dihydroxylated naphthalenes or aromatic nitrogen-heterocyclic compounds; all such compounds were false positives. Results with dihydroxylated naphthalenes reveal derivatives that were nonestrogenic when superimposed on the phenolic A-ring of 17-beta-estradiol had the second hydroxyl group in the position of the C-ring or were catechol-like in structure. Structural alerts for nitrogen-heterocyclic compounds must take into account the position of the hydroxyl group and the in-ring nitrogen atom; compounds with the hydroxyl group and nitrogen atom involved with the same ring were observed to be nonactive.     

Synthesis of 9-substituted 2,3,4,9-tetrahydro-1H-carbazole derivatives and evaluation of their anti-prion activity in TSE-infected cells

2,3,4,9-Tetrahydro-9-[2-hydroxy-3-(1-piperidinyl)propyl]-6-methyl-1H-carbazol-1-one (GJP14) is a novel anti-prion compound that we previously discovered by in silico screening and cellular assay. In this study, a variety of GJP14 derivatives were prepared using pyrrole derivatives, (haloalkyl)oxiranes, and amines, and their anti-prion activity was evaluated in TSE-infected cells. It was found that the tricyclic aromatic ring, a hydroxy group at the 2-position and an amino group at the 3-position of the N-propyl group were the basic requirements for anti-prion activity. The derivatives bearing an N-ortho-halobenzyl group exhibited an improved activity, and the most potent derivative was 8 times as effective as the original lead compound, GJP14.     

Structure elucidation and conformational analysis of gonadotropin releasing hormone and its novel synthetic analogue [Tyr(OMe), d-Lys, AzeNHEtGnRH: The importance of aromatic clustering in the receptor binding activity

The conformational properties of gonadotropin releasing hormone (GnRH) in dimethylsulfoxide-d₆ were investigated by nuclear Overhauser effect (nOe) enhancement studies and were compared with the conformational properties of its analogue [Tyr(OMe)⁵GnRH resulting after methylation of the tyrosine hydroxyl. Assignment of all backbone and side-chain protons was possible by combining information from intraresidue nOe studies with two-dimensional correlated spectroscopy (COSY/TOCSY) studies. Saturation of distinct proton resonances of the three aromatic residues Tyr, His, Trp, in clear areas of the NMR spectrum of GnRH resulted in interresidue enhancements of aromatic resonances indicating the proximity of the three aromatic rings. This spatial proximity is not observed in [Tyr(OMe)⁵]GnRH and is correlated with a lower receptor binding affinity in the rat pituitary (K_d = 1.53 ± 0.35 × 10⁻⁶ M) compared with that exerted by GnRH (K_d = 3.69 ± 0.89 × 10⁻⁹ M). However, substitution of Gly at position 6 of [Tyr(OMe)⁵]GnRH with d-Lys⁶ and further replacement of Pro at position 9 with the more rigid Aze residue [Tyr(OMe)⁵, d-Lys⁶, Aze⁹NHEt]GnRH significantly improved the binding affinity (K_d = 0.689 ± 10.15 × 10⁻⁹) and this may be due to the restoration of the ring cluster. Overall, the clustering of the aromatic rings observed in GnRH was not seen in [Tyr(OMe)⁵]GnRH and this conformational difference may be responsible for receptor recognition and higher binding of the parent peptide.     

Structure-stability relationship of anthocyanins under cell culture condition

This work aims to evaluate the structure-stability relationship of anthocyanins in cell culture. An early degradation time (^CT₁₀) and half-degradation time (^CT₅₀) were used to characterise the stability of 10 of the most common anthocyanins, incubated with DMEM at 37?°C, pH?=?7.4, 5% CO₂ for different time periods. According to the glycosylation, the glycosylated forms were more stable than the not glycosylated forms. The methylation at 3’' or 5' position at ring B enhanced their stability; contrarily, the hydroxylation at 3' or 5' position at ring B weakened their stability. Glycosylated forms were much more stable in water than in the culture medium. Although not glycosylated forms were also instable in water, their stability was improved compared with culture medium. Together with the cell culture experiments and, in order to avoid artefacts, stability tests of polyphenols should be performed in parallel experiments with DMEM.     

Design, synthesis and structure-activity relationships of antiproliferative 1,3-disubstituted urea derivatives

Twenty-four new 1,3-disubstituted urea derivatives were synthesized and reported for the first time. The antiproliferative activities of these compounds were evaluated against a panel of one human liver cell line (L02) and two human tumor cell lines (KB and K562) by applying the MTT colorimetric assay. The series of 1,3-disubstituted urea derivatives show good antiproliferative activity against human cancer cell lines (KB and K562) and no antiproliferative activity against liver cell line (L02). The potent in vitro antiproliferative activity of these derivatives and their selectivity for L02 are quite important points for an anticancer drug candidate with fewer side effects. Structure-activity relationships were also discussed based on the obtained experimental data. The hydroxyl groups on the phenyl ring reduced the antiproliferative activities of 1,3-disubstituted urea derivatives. The OH groups could be responsible for a reduction in the permeability of the cell membrane. Generally, an aromatic ring on N-3 seems to be in favor of enhancing the inhibitory activity, compounds introduced a nitro group substituent at C-3 position on the aromatic ring approved to generally decrease activity.     

Synthesis of prenylated benzaldehydes and their use in the synthesis of analogues of licochalcone A

In this paper, a general applicable synthesis of prenylated aromatic compounds exemplified by prenylated benzaldehydes starting from readily available acetophenones is described. The synthesized benzaldehydes are used to prepare a number of novel analogues of Licochalcone A, a known antibacterial compound, and for the exploration of the pharmacophoric elements that are essential for the antibacterial activity. It is shown that the hydroxyl group in the A ring is essential for the activity and that the hydroxyl group in the B ring has no influence on the antibacterial effect of Licochalcone A. Furthermore, it is shown that the prenyl group at the position 5 of the B ring also has a dominating influence on the activity. This aliphatic group can be replaced by other lipophilic long chained substituents in order to maintain the activity.     

Aryl sulfate formation in sea urchins (Strongylocentrotus droebachiensis) ingesting marine algae (Fucus distichus) containing 2,6-dimethylnaphthalene

The metabolism of tritiated 2,6-dimethylnaphthalene (2,6-DMN) was studied in sea urchins (Strongylocentrotus droebachiensis) feeding on marine algae (Fucus distichus). The Fucus accumulated this hydrocarbon from sea water without converting it to metabolites. Most of the tritium accumulated by the sea urchins (e.g., 70.8% after 3 days) from feeding on 2,6-DMN-exposed Fucus was present in the exoskeleton (shell and spines). Moreover, after 3 days feeding, about 90% of the tritium in the total metabolite fraction of the gonads and digestive tract of the sea urchin was present as sulfate derivatives. These metabolites were identified through hydrolysis with aryl sulfatase, followed by thin-layer chromatography of the products. After 14 days of feeding, the tritium associated with the sulfate derivatives decreased in the gonads and digestive tract to 61 and 65%, respectively, of the total metabolite fraction. Hydroxy compounds from sulfatase hydrolysis were chromatographed using multiple elutions with toluene. The hydroxy isomers were separated and the R_f values were compared to those of pure reference compounds. The data indicated that 80% of the 2,6-dimethylnaphthyl sulfate contained the sulfate on the 1 and/or 3 position of the aromatic ring. Moreover, 6-methyl-2-naphthalenemethanol was not detected, which implies that sea urchins, unlike fish, metabolize alkyl-substituted aromatic hydrocarbons primarily through aromatic ring oxidations.     

Amide isosteres in structure-activity studies of antibacterial minor groove binders

Antibacterial minor groove binders related to the natural product, distamycin, are development candidates for novel antibiotics. Alkenes have been found to be effective substitutes for the isosteric amide links in some positions and alkyl groups larger than methyl have been found to increase binding to DNA in both selectivity and affinity. However the impact of other isosteres such as diazenes and the position of an alkyl group with respect to DNA binding and antibacterial activity are not known. The effects of some systematic variations in the structure of polyamide minor groove binders are investigated. Isosteres of the amide link (alkenes and diazenes) are compared: it is shown that all three are competent for binding to DNA but that alkene links give the tightest binding and highest antibacterial activity; no significant antibacterial activity was found for compounds with a diazene link. Within a series of alkene linked compounds, the effect of branched N-alkyl substituents on binding to DNA and antibacterial activity is investigated: it was found that C3 and C4 branched chains are acceptable at the central pyrrole residue but that at the pyrrole ring adjacent to the basic tail group, a C4 branched chain was too large both for DNA binding and for antibacterial activity. The active branched alkyl chain compounds were found to be especially active against Mycobacterium aurum, a bacterium related to the causative agent of tuberculosis.     

Synthesis of 2-substituted beta-cyclodextrin derivatives with a hydrolytic activity against the organophosphorylester paraoxon

Beta-cyclodextrin was substituted by an iodosobenzoic acid derivative to create a catalytic hydrolytic activity against neurotoxic organophosphorus agents. The catalytic moiety was introduced on a secondary hydroxy group at the position 2 of a glucose unit. Several beta-cyclodextrin derivatives were obtained. In these derivatives, the methylene linker occupied all potential positions on the aromatic ring. Kinetic assays were carried out with paraoxon as organophosphate model. Three regioisomers hydrolyzed paraoxon, although the paraoxon-leaving group, para-nitrophenol, was not released from the beta-cyclodextrin torus.     

Naphthalimido derivatives as antifolate thymidylate synthase inhibitors

A new series of N-(substituted)benzyl-1,8-naphthalimides 4, structurally related to the previously reported thymidylate synthase (TS) inhibitor naphthaleins 3, were synthesized and compounds tested for their inhibition of several species of TS. Moreover, their in vitro cytotoxicity together with antimycotic and antibacterial properties were assayed. While no activity was detected in the antibacterial tests, the m-nitro (4ae) and the p-nitro (4af) derivatives were found able to partially inhibit TS at low micromolar concentrations. Introduction of nitro or (substituted)-amino groups in position 4 of the naphthalic ring always led to less active compounds.     

Urinary 1-hydroxypyrene and 2-naphthol concentrations in male Koreans

Objective: Urinary 1-hydroxypyrene (1-OHP) has been used as a biological marker of exposure to polycyclic aromatic hydrocarbons (PAHs), and urinary 2-naphthol is suggested as a new marker for route-specific exposure to airborne PAHs. We analyzed urinary 1-OHP and 2-naphthol concentrations in 292 male Koreans (129 university students and 163 shipyard workers) to define the distribution pattern in Koreans with no or low occupational exposure to PAHs. Method: Histories of cigarette smoking and the eating of PAH-containing foods were obtained by a self-administered structured questionnaire. Urine samples were collected and urinary 1-OHP and 2-naphthol concentrations were measured using high-performance liquid chromatography (HPLC). Results: The arithmetic (geometric) means of urinary 1-OHP and 2-naphthol concentrations for all students, expressed as micromoles per mole of creatinine, were 0.04 (0.04) and 3.12 (2.22), for non-smokers 0.03 (0.03) and 1.78 (1.30) and for smokers 0.05 (0.03) and 4.36 (3.62), respectively. Among shipyard workers, the arithmetic (geometric) means of urinary 1-OHP and 2-naphthol concentrations were 0.69 (0.31) and 4.37 (2.62) for all, 0.27 (0.18) and 2.46 (1.16) for non-smokers, and 0.97 (0.44) and 5.60 (4.44) for smokers, respectively. Mean urinary 1-OHP and 2-naphthol concentrations differed significantly between non-smokers and smokers both in students and in shipyard workers. In smokers, some variables related to smoking habit were positively correlated with urinary 1-OHP and with 2-naphthol concentrations. The latter showed better correlations with the variables related to smoking amount than the former. None of the food-related factors was significantly correlated with urinary 1-OHP or 2-naphthol concentration. Conclusion: These results suggest that urinary 2-naphthol concentration is more sensitively affected by smoking status than urinary 1-OHP concentration and that urinary 2-naphthol is a sensitive marker for low-level inhalation of PAHs. Received: 6 March 2000 / Accepted: 26 July 2000     

Drug design: Le contrôle LUMO du pharmacophore neuroleptique

The aromatic ring of the antagonist of the dopaminergic receptor behaves as an electron acceptor within its interactions with the receptor. The energy of the lower unoccupied molecular orbital (ϵ_LUMO) is correlated with the biological activity for different series of neuroleptic compounds. The principle of interaction LUMO antagonist—HOMO receptor (Principle La—Hr), presented previously for the phenothiazine series, is extended to various series of antagonists of the dopaminergic receptor: thioxanthenes, butyrophenones, benzisoxazoles and benzamides.The Principle La—Hr is refined on a model series of 16 phenothiazines by a multilinear regression giving the biological activity versus 3 variables, with a stepwise procedure. The variable ϵ_LUMO has the strongest contribution, the aliphatic amine chain has a moderate contribution and the variation of lipophilicity induced by the substituent of the aromatic ring in position 2 has only a very weak contribution. The correlation is used to predict, with success, the relative activity of two new phenothiazines: mesoridazine and thioridazine.     

Building a model of interaction at the NK-2 receptors: Polycondensed heterocycles containing the pyrimidoindole skeleton

The pyrazolopyrimidothiazole ring system (compound N, table II) has been previously reported by us as a new competitive antagonist (apparent pA₂ = 7.3 equiv to 55 nM) at NK2-receptors. As part of our investigation on polycondensed heterocycles containing the pyrimidine ring as antagonists of G-protein coupled receptors, pyrimidoindole derivatives were prepared and tested in order to probe the topography of the NK2-receptors and ascertain the pattern of frameworks that result in optimum affinity and specificity. The title indole derivatives 5, 11a-d, 12c, 13a,b and 14b were ‘de novo’ designed or selected from our chemical archives and prepared by up-to-date synthetic routes, thus exploring new synthetic methodologies. According to the established graphic computer model, none of the tested substances exhibited activity as a consequence of the violation of an excluded volume area due the unfavourable position of the aromatic substituents.     

Novel cephalosporin derivatives possessing a substituted cinnamoyl moiety at the 7 beta-position. Synthesis, structural characterization and antibacterial activity of 3-acetoxymethyl cephalosporin derivatives

Twenty 3-acetoxymethyl cephalosporin derivatives, with various cinnamoyl (3-phenyl-2-propenoyl) substituted groups at the 7beta-position, were synthesized and evaluated for antibacterial activity in vitro. Some of these cephalosporin derivatives showed good selective activity against Gram-positive bacteria. Although substitution on the aromatic ring of cinnamoyl moiety generally reduced antimicrobial activity against Staphylococcus sp. and Enterococcus sp., a hydroxy group at the para position, and particularly ortho, para di-chloro substitution, improved the activity against methicillin resistant strains of Staphylococcus aureus (MRSA). Substitution on the double bond alpha position of the cinnamoyl moiety also affected the antimicrobial activity. A cyano group attached to this position increased activity against both negative coagulase Staphylococcus and Enterococcus sp. and extended the antibacterial spectrum towards Gram-negative bacteria.     

Rationalization of physicochemical characters of oxazolyl thiosemicarbazone analogs towards multi-drug resistant tuberculosis: a QSAR approach

The emergence of multi-drug resistant (MDR) strains of Mycobacterium tuberculosis and the continuing pandemic of tuberculosis emphasizes the urgent need for the development of new and potent anti-tubercular agents. In an effort to develop new and more effective agents to treat tuberculosis emphasis was focused on quantification of structure-activity relationship of oxazolyl thiosemicarbazone derivatives. The de novo analysis gave insight to some important structural features i.e. nitro group on phenyl ring at R(1) position is optimal for the activity and might be responsible for electronic interaction, while phenyl ring at R position interact with the hydrophobic pocket more effectively as compared to unsubstituted or methyl substituted analogs. Hansch approach offered the understanding and parameterization of interactions of the inhibitor with receptor. Similarly QSAR analysis gave some important physicochemical properties, i.e. empirical aromatic index (ARR) and 3D-MoRSE code value of scattering angle at 8A(-1). These two physicochemical properties shall be helpful in the development of more potent analogs.     

Pollen-related food allergy: cloning and immunological analysis of isoforms and mutants of Mal d 1, the major apple allergen, and Bet v 1, the major birch pollen allergen

Summary Background: Mal d 1, the major apple allergen, cross-reacts with IgE specific for the major birch pollen allergen, Bet v 1, and is responsible for birch pollen related food allergy to apple. Isoforms of Bet v 1 showing minor sequence variations display different binding capacitiy for specific IgE antibodies from allergic patients. Moreover, strain-dependent variation of allergenicity has been reported for apples. Objective: To investigate the occurence of strain-dependent isoforms of Mal d 1 which may differ in their allergenic potential, to obtain data on structures essential for binding of Mal d 1 to the antibody, and to gain insights into the structures responsible for its IgE cross-reactivity to Bet v 1. Methods: The cDNA of Mal d 1 from various apple strains was amplified by a PCR strategy based on conserved regions of known Mal d 1-sequences, and sequenced. Two major isoforms of Mal d 1were expressed as recombinant proteins and purified, as were different variants of the major birch pollen allergen, Bet v 1. Together with already existing recombinant birch pollen and apple allergens, these were subjected to allergenicity testing by IgE-immunoblotting, enzyme allergo sorbent test and dose related mediator release. “Hot-spots” for IgE-reactivity were identified by site-directed mutagenesis. Results: Twelve Mal d 1-clones were sequenced from 7 apple varieties and compared to 3 known Mal d 1 sequences. The clones were clustered into two groups, each showing a high degree of sequence identity to one of the known sequences and specific differences to the third sequence. No strain-specific sequences were identified. In contrast, apple strains with reported differences in allergenicity showed different expression levels of the major allergen. Immunologic testing of recombinant allergens revealed high IgE binding capacity of 2 major isoforms, named GD26 and GS29, with a slightly higher IgE binding capacity of DG26. Moreover, the allergenicity was similar to another rMal d 1 reported in the literature, representing the isoform divergent from our clones. Mutational analysis of our Mal d 1 allergens identified serine in position 111 as essential for IgE binding. Allergenicity was almost depleted by changing this residue into a proline. Moreover, the corresponding serine residue, present in position 112 of Bet v 1, was in a similar manner crucial for the allergenicity of the birch pollen allergen. Conclusion: We conclude that divergent allergenicity of apple strains mainly depends on differnet expression levels of the major allergen. Introduction of a proline residue in position 111 of Mal d 1 and in position 112 of Bet v 1 led to a drastic reduction of allergenicity of both the pollen and the food allergen, obviously also removing the cross-reactive epitope. Mutants with reduced IgE-reactivity but maintained T-cell reactivity may represent new candidates for a safer specific immunotherapy with reduced side-effects. Received: 7 June 1999, Accepted: 30 July 1999     

Biodegradation of [14C] Ring-Labeled Nonylphenol Ethoxylate

Nonylphenol (NP) and the 9-mole ethoxylate of nonylphenol (NPE9) were synthesized with a uniform radioactive ¹⁴C label in the aromatic ring. The [¹⁴C]NP isomer distribution and [¹⁴C]NPE9 oligomer distribution closely matched that of commercial NPE9. Biodegradation of [¹⁴C]NPE9 was examined under conditions simulating a river water environment, and changes in the oligomer distribution and mineralization to ¹⁴CO₂ were monitored for 128 days. Over 40% of the [¹⁴C]NPE aromatic ring carbon was converted to ¹⁴CO₂ and another 21% was incorporated into the biomass. Primary degradation of NPE (conversion to metabolites other than NP, NPE ethoxylates, and NPE carboxylates) was estimated to be 87–97%. NP was a minor metabolite, accounting for less than 0.4% of the initial NPE. These studies demonstrate that the phenolic ring of NPE is opened, metabolized, and mineralized in the aquatic environment.