Novel urea and bis-urea primaquine derivatives with hydroxyphenyl or halogenphenyl substituents: Synthesis and biological evaluation.

A series of novel compounds 3a-j and 6a-j with primaquine and hydroxyl or halogen substituted benzene moieties bridged by urea or bis-urea functionalities were designed, synthesized and evaluated for biological activity. The title compounds were prepared using benzotriazole as the synthon, through several synthetic steps. 3-[3,5-Bis(trifluoromethyl)phenyl]-1-{4-[(6-methoxyquinolin-8-yl)amino]pentyl}urea (3j) was the most active urea and 1-[({4-[(6-methoxyquinolin-8-yl)amino]pentyl}carbamoyl)amino]-3-[3-(trifluoromethyl)phenyl]urea (6h) the most active bis-urea derivative in antiproliferative screening in vitro against eight tested cancer cell lines. Urea derivatives 3a-g with hydroxy group or one halogen atom showed moderate antiproliferative effects against all the tested cell lines, but stronger activity against breast carcinoma MCF-7 cell line, while trifluoromethyl derivatives 3h-j showed antiproliferative effects against all the tested cell lines in low micromolar range. Finally, bis-ureas with hydroxy and fluoro substituents 6a-d showed extreme selectivity and chloro or bromo derivatives 6e-g high selectivity against MCF-7 cells (IC50 0.1-2.6 µM). p-Fluoro derivative 6d, namely 3-(4-fluorophenyl)-1-[({4-[(6-methoxyquinolin-8-yl)amino]pentyl}carbamoyl)amino]urea, is the most promising compound. Further biological experiments showed that 6d affected cell cycle and induced cell death of MCF-7 cell line. Due to its high activity against MCF-7 cell line (IC50 0.31 µM), extreme selectivity and full agreement with the Lipinski's and Gelovani's rules for prospective small molecular drugs, 6d may be considered as a lead compound in development of breast carcinoma drugs. Urea 3b and almost all bis-ureas showed high antioxidant activity in DPPH assay, but urea derivatives were more active in lipid peroxidation test. Only few compounds exhibited weak inhibition of soybean lipoxygenase. Compound 3j exhibited the strongest antimicrobial activity in susceptibility assay in vitro (MIC = 1.6-12.5 µg ml-1).

Novel semicarbazides and ureas of primaquine with bulky aryl or hydroxyalkyl substituents: synthesis, cytostatic and antioxidative activity.

Novel primaquine semicarbazides 7a-l and ureas 9a-g with modified benzhydryl, trityl, phenyl or hydroxyalkyl substituents were prepared and evaluated for cytostatic and antioxidative activities. Two synthetic approaches for preparation of the title semicarbazides were applied, both having certain advantages. In the first approach, the products grew from the semicarbazide side and the primaquine residue entered the molecule the last. In the second approach, semicarbazide grew from the primaquine side. This method was more convenient for synthesis of a series of semicarbazides: various products could be obtained from the same precursor N-(4-((6-methoxyquinolin-8-yl)amino)pentyl)hydrazinecarbox-amide (10). Primaquine ureas 9a-f were prepared from primaquine benzotriazolide 8 and corresponding amines and urea 9g directly from primaquine and 4-chloro-3-(fluoromethyl)phenyl isocyanate. All primaquine semicarbazide derivatives showed either prominent cytostatic activity towards all the tested cell lines (benzhydryl or trityl derivatives 7a-e) or high selectivity towards MCF-7 cells (hydroxyalkyl derivatives 7h-l), with IC50 values in the low micromolar range. The highest selectivity exerted symmetric bisprimaquine derivative 7f, with an IC50 0.2 µM against MCF-7 cells and practically no activity against other seven tested cancer cell lines. Urea derivatives 9a-f were generally less active than their semicarbazide analogues, but still selective towards MCF-7 cells. Urea 9g with the similar structure to cytostatic drug sorafenib, was the most active urea derivative. Semicarbazides 7g and 10 showed the best antioxidative activity as measured by DPPH (64% at 20 min and 90% at 60 min), while urea derivatives 9a-g, especially 9d, and semicarbazides 7a-g with lipophilic substituents exerted better LP antioxidant activity. Both semicarbazides and ureas with methoxy or chloro benzhydryl substituents and high Clog P values showed significant LOX inhibition.

Novel NSAID 1-acyl-4-cycloalkyl/arylsemicarbazides and 1-acyl-5-benzyloxy/hydroxy carbamoylcarbazides as potential anticancer agents and antioxidants.

The novel 1-acyl-4-cycloalkyl/arylsemicarbazides (5a-y) and 1-acyl-5-benzyloxy/hydroxycarbamoylcarbazides (8a-f) derived from the nonsteroidal anti-inflammatory drugs ibuprofen, fenoprofen and reduced ketoprofen were prepared, fully chemically characterized and evaluated for their cytostatic, antiviral and antioxidant activities. Compounds 5 and 8 consist of a region rich in electronegative atoms (five to nine nitrogen and oxygen atoms) framed by aryl or cycloalkyl residues on one or both terminal ends. The synthetic pathways applied for the preparation of the title compounds involved a benzotriazole as a synthetic auxiliary in several steps. Three of the tested compounds, namely 4-benzhydryl-1-[2-(3-phenoxyphenyl)propanoyl]semicarbazide (5l), 4-benzhydryl-1-[2-(3-benzylphenyl)propanoyl]semicarbazide (5s), and 4-benzhydryl-1-[2-(4-isobutylphenyl)propanoyl]semicarbazide (5f) showed pronounced antiproliferative activity in vitro against six cancer cell lines (IC(50)=3-23 µM). The same compounds highly inhibited soybean lipoxygenase (IC(50)=60 and 51.5 µM) and lipid peroxidation as well (99, 88 and 74%, respectively). 4-Benzyloxy-1-[2-(4-isobutylphenyl)propanoyl]semicarbazide (5t) and 5-benzyloxycarbamoyl-1-[2-(3-benzylphenyl)propanoyl]carbazide (8c) exerted complete lipid peroxidation inhibition. Semicarbazides 5w-y and carbazides 8d-f bearing a hydroxamic acid/hydroxyurea moiety showed a modest antiradical activity in DPPH test, while the best radical scavenger was 1-(1-benzotriazolecarbonyl)-4-benzyloxysemicarbazide (7). None of the compounds were inhibitory to a broad panel of DNA and RNA viruses in the cell culture at subtoxic concentrations.

Urea and carbamate derivatives of primaquine: synthesis, cytostatic and antioxidant activities.

The novel urea primaquine derivatives 3 were prepared by aminolysis of primaquine benzotriazolide 2 with several hydroxyamines and ethylendiamine, while carbamates 4 were synthesized from the same precursor 2 and alcohols. All compounds are fully chemically characterized and evaluated for their cytostatic and antioxidant activities. The most prominent antiproliferative activity was obtained by compounds 3c, 3d, 3g, and 5b (IC(50)=9-40 microM). 1-(5-Hydroxypentyl)-3-[4-(6-methoxy-quinolin-8-ylamino)-pentyl]urea (3c) showed extreme selectivity toward SW 620 colon cancer cells (IC(50)=0.2 microM) and a bit less toward lung cancer cells H 460. Hydroxyurea 3h showed the highest interaction with DPPH. Primaquine twin drug 3g showed very significant inhibition on LOX soybean (IC(50)=62 microM). Almost all the tested derivatives highly inhibited lipid peroxidation, significantly stronger than primaquine phosphate.

The novel phosphoramidate derivatives of NSAID 3-hydroxypropylamides: synthesis, cytostatic and antiviral activity evaluations.

The target phosphoramidates 5a-e were prepared in one step from 3-hydroxypropyl derivatives 3a-e of nonsteroidal anti-inflammatory drugs (fenoprofen, ketoprofen, ibuprofen, indomethacin, diclofenac). The products 3a-e and 5a-e were evaluated for their cytostatic and antiviral activity against malignant tumour cell lines and normal human fibroblasts (WI 38). All phosphoramidate derivatives 5a-e possess significantly greater inhibitory activities than the corresponding 3-hydroxypropyl derivatives 3a-e, whereby compound 5a showed the most potent inhibitory activities against cervical, pancreatic and colon carcinoma cell lines (IC(50)=5-7 microM).

The novel primaquine derivatives of N-alkyl, cycloalkyl or aryl urea: synthesis, cytostatic and antiviral activity evaluations.

The novel urea primaquine derivatives 3a-i were prepared by aminolysis of benzotriazolide 2 with the corresponding amine in the presence or absence of triethylamine. Compound 2 was prepared by acylation of primaquine with 1-benzotriazole carboxylic acid chloride. Among all compounds evaluated, the pyridine derivative 3h exhibited the best cytostatic activities against colon carcinoma, human T-lymphocyte and murine leukemia. However, this compound showed also rather marked cytotoxicity towards human normal fibroblasts. The highest selectivity in the inhibitory effects on human malignant tumor cell lines vs. normal fibroblasts was found for ureas 3c, 3d and 3g. Results of broad antiviral evaluation showed that pyridine and phenethyl derivatives of urea 3h and 3g exhibited some selective inhibition against cytomegalovirus.

Synthesis and in vitro antitumor effect of diclofenac and fenoprofen thiolated and nonthiolated polyaspartamide-drug conjugates.

This paper reports the synthesis and antiproliferative effects of new thiomer-diclofenac and fenoprofen conjugates, hydrophilic, bioadhesive, polymeric prodrugs, as well as antiproliferative effects of diclofenac, fenoprofen and a series of previously described polymer-fenoprofen conjugates on five tumor cell lines. Thiolated and nonthiolated polyaspartamides were the chosen polymeric components. Drug-loading ranged from 5.6 to 22.4%, and the amount of SH groups ranged from 6.9 to 45.6micromol g(-1). Tensile studies demonstrated a clear correlation between the amount of thiol and the mucoadhesive properties of the conjugates. The growth-inhibitory activity of the tested polymer-drug conjugates demonstrates that polyaspartamide-type polymers, especially thiolated polymers, enable inhibition of tumor cell growth with significantly lower doses of the active substance.

Synthesis and X-ray crystal structure study of the hydroxyurea and hydantoin derivatives of L-valine.

The novel hydroxyurea 5 derivative of L-valine was prepared by aminolysis of N-(1-benzotriazolecarbonyl)-L-valine cyclohexanemethylamide 4 with hydroxylamine. The corresponding hydantoin derivative 6 was synthesized by base catalyzed cyclization of the amide 4. The exact stereostructure of hydantoin derivative 6 has been determined by X-ray crystal structure analysis. The chiral atom of the hydantoin ring in 6 has S configuration what is in agreement with its configuration in the starting L-valine. The molecules of 6 are joined into infinite chains by N-H...O intermolecular hydrogen bond. The infinite chains are additionally linked by two C-H...O hydrogen bonds, thus forming two-dimensional network. The hydantoin derivative of L-valine 6 and its L-leucine analogue LH have similar packing arrangements, so they are homostructural.

Synthesis and characterization of thiomers of polyaspartamide type.

Synthesis of poly[alpha,beta-(N-2-hydroxyethyl-DL-aspartamide)]-thioglycolic acid (PHEA-TGA) conjugate as a new polyaspartamide thiomer is described. The parent polymer PHEA is chemically modified by introducing sulphydryl-bearing compound thioglycolic acid. By varying the reaction conditions several batches of PHEA-TGA conjugates were prepared and analyzed. Tensile studies revealed that total work of adhesion of PHEA-TGA increased more than twice compared to the unmodified polymer. Microparticles prepared from the thiolated polymer preserved its bioadhesive properties.

Macromolecular prodrugs XI. Synthesis and characterization of polymer-estradiol conjugate.

Estradiol-3-benzoate (EB), an ester derivative of the main oestrogen hormone estradiol, was chemically modified and bound to poly(alpha,beta-(N-2-hydroxyethyl-DL-aspartamide))-poly(alpha,beta-(N-2-aminoethyl-DL-aspartamide)) copolymer (PAHA). EB was first converted to estradiol-3-benzoate-17-(benzotriazole-1-carboxylate), which readily reacted with amino groups in PAHA affording the polymer-drug conjugate PAHA-EB. In PAHA-EB estradiol moiety was covalently bound to the polymeric carrier by carbamate linkage, through non-toxic ethylenediamine spacer. The synthesized compound is a potential hydrosoluble estradiol prodrug.

Macromolecular prodrugs: X. Kinetics of fenoprofen release from PHEA-fenoprofen conjugate.

The kinetics of fenoprofen release from poly[alpha,beta-(N-2-hydroxyethyl-DL-aspartamide)]-fenoprofen conjugate (PHEA-Fen) in aqueous buffer solutions (pH 10 and 1.1), simulated gastric (SGF) and intestinal fluids (SIF) was studied. In borate buffer pH 10, the following rate constants were obtained: k=0.2659 (t=60 degrees C) and k=0.0177 h(-1) (t=37 degrees C) and in glycine buffer solution pH 1.1 k=0.0036 h(-1). In SGF and SIF fenoprofen release did not occur in significant extend within 12 h. The hydrolysis of the ester bond between the polymeric carrier and fenoprofen followed the pseudo first-order kinetics, with activation energy indicative for the breakage of a sigma bond (E(a)=100.6 kJ mol(-1)). The concentration of the released fenoprofen was determined by high performance liquid chromatography (HPLC).

Macromolecular prodrugs. IX. Synthesis of polymer-fenoprofen conjugates.

Synthesis of several polymer-fenoprofen conjugates is described. Fenoprofen was first chemically modified into benzotriazolide 2 and amino acid amide derivatives: glycine fenoprofenamide (3a) and beta-alanine fenoprofenamide (3b) and their benzotriazolides 6a and 6b. Compounds 2 and 6 readily reacted with polyhydroxy aspartamide-type polymers, i.e. poly[alpha,beta-(N-2-hydroxyethyl-DL-aspartamide)] (PHEA) and poly[alpha,beta-(N-3-hydroxypropyl-DL-aspartamide)] (PHPA) forming conjugates 5, 8a,b and 9a,b, respectively. Conjugate 11 was obtained by partial aminolysis of poly-DL-(2,5-dioxo-1,3-pyrrolidinediyl) (PSI) with 2-aminoethyl fenoprofenamide (3c), followed by total aminolysis with 2-hydroxyethylamine. The synthesised polymer-drug conjugates differed in type of covalent bounding, type and/or length of spacer and drug-loading.

Gemfibrozil ester and amide derivatives--synthesis, spectroscopic characterisation and QSPR.

The synthesis and spectroscopic characterisation of various gemfibrozil esters 3 and amides 4 are described. In the first step gemfibrozil was reacted with N-1-benzotriazolecarboxylic acid chloride (1) yielding gemfibrozil benzotriazolide (2). Compound 2 readily reacted with alcohols and amines to form the corresponding esters 3 and amides 4, potential prodrugs of the well known hypolipaemic drug gemfibrozil. The quantitative structure property relationship (QSPR) was studied in the series of gemfibrozil esters and amides. The following topological descriptors and physicochemical parameters were used: Wiener number (W), connectivity index (l chi v), relative molecular mass (M(r)), van der Waals volume (Vw) and parameters of lipophilicity (log P and RM).

Macromolecular prodrugs. VIII. Synthesis of polymer-gemfibrozil conjugates.

Gemfibrozil is covalently linked to two similar polymers: poly[alpha,beta-(N-2-hydroxyethyl-DL-aspartamide)] and poly[alpha,beta-(N-3-hydroxypropyl-DL-aspartamide)]. The synthesised polymer drug conjugates differ in average molecular mass, type of covalent bonding, length of spacer, drug-loading and solubility.

Inhibition of high affinity choline uptake in the rat brain by neurotoxins: effect of monosialoganglioside GM1.

Mustard derivatives of ethyl-choline and hemicholinium-3 have been suggested as possible specific cholinergic neurotoxins. In this study a structural analog of hemicholinium-3, a,a'-bis[di(2-chloroethyl)amino]-4,4'-2-biacetophenone (toxin 7), was added to synaptosomes prepared from the cortex, striatum or hippocampus of rat brain. Synaptosomal high affinity choline uptake (HACU) was significantly decreased in a dose-dependent manner by addition of toxin 7, while synaptosomal uptake of GABA or dopamine was not changed. Incubation of cortical synaptosomes with the monosialoganglioside GM1 prevented the decrease in HACU seen following administration of toxin 7. This preventative effect of GM1 was greater if GM1 was added prior to or concomitant with toxin 7, than if GM1 was added following toxin 7. Two newly synthesized hemicholinium-3 analogs, 4-[3'-di(2-chloroethyl)aminopropionyl]biphenyl (toxin 5) and 4-[3'-di(2-bromoethyl)aminopropionyl]biphenyl (toxin 6) caused a large decrease in HACU when added to cortical synaptosomes, this decrease was significantly greater than that seen with the same dose of toxin 7 or ethyl-choline aziridinium (AF64A). Ultrastructural changes in the synaptosomal membrane following incubation with toxin 7 or toxin 7 with GM1 were examined by electron microscopy. Development of a compound which is both a potent neurotoxin, and is specific for cholinergic neurons will allow new insights into the normal function of the cholinergic system in the CNS and provide animal models of disease states in which cholinergic degeneration is an important element.

Structural analogs of hemicholinium: effects on cholinergic and GABAergic markers in the CNS.

The synthesis and neurotoxic effects of several structural analogs of hemicholinium were studied. All compounds were injected unilaterally into the lateral ventricle (4 nmol) and the effects of the hemicholinium derivatives on choline acetyltransferase (ChAT) and glutamic acid decarboxylase (GAD) activity were compared with those of AF64A in an equimolar concentration. Structures of the newly synthesized compounds were confirmed by i.r., NMR and u.v. spectrometry and elemental analysis. The most specific cholinotoxic effects were observed with a,a-bis[di(2-chloroethyl)amino]4,4-biacetophenone (toxin 7). This compound causes specific decrease of ChAT activity in the brain structures (hippocampus and cortex) containing cholinergic terminals deriving from septum and the nucleus basalis magnocellularis (NBM), respectively. Large ChAT-positive magnocellular neurons in the NBM became paler and lost their processes following treatment with toxin 7 after 1 week.