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Serotonin 5HT1a receptors belongs to a class G-protein coupled receptors and it is widely recognized as one of the targets for treating neurological disorders such depression and schizophrenia. N-arylpiperazine structural motif is present in many compounds with pronounced 5HT1a activity including recently approved aripiprazole for the treatment of the major depressive disorder.

Alzheimer's disease (AD) is a serious, neurodegenerative disease, characterized with a progressive loss of cognitive and behavioral functions leading to fatal outcome. Tacrine belongs to a class of cholinesterase inhibitors(ChEls) and it is the first ChEI marketed for the treatment of AD symptoms.

Alzheimer's disease (AD) is a progressive, neurodegenerative disorder characterized by deterioration of cognitive skills and it is the most abundant form of dementia according to the World Health Organization (WHO).1 Tacrine belongs to a class of acetylcholinesterase inhibitors (AChEIs) and it was formerly used in the treatment of AD symptoms. Albeit no longer in clinical use due to its hepatotoxicity, tacrine ring system is extensively investigated as pharmacophore in novel AChEIs, many of which showed improved pharmacological profile in the preclinical studies.2 Here we present optimized synthetic pathway and pharmacological evaluation of three novel tacrine derivatives (5, 6 and 7), possessing functionalized piperidine connected with tacrine via five methylene groups alkyl chain

Alzheimer’s disease (AD) is a neurodegenerative disease which affects over 50 milion people worldwide.1) AD mainly affects people ages 65 and above. Symptoms are memory loss, decline in learning capacity, and language problems. One of currently available treatments for Alzheimer's disease consists of Cholinesterase (ChEIs) inhibitors, which raise acetylcholine levels in the brain. The drugs currently approved by the FDA for this purpose are donepezil, rivastigmine and galantamine. As a part of our research we designed and synthesized seven novel compounds (6a-g) as potential acetylcholinesterase inhibitors (AChEIs). The compounds contain N-benzyl piperidine moiety, which is common in many inhibitors including donepezil,2) and N-aryl piperazine moieties.

Herein, we report on the synthesis and pharmacological evaluation of ten novel fluorinated cinnamylpiperazines as potential monoamine oxidase B (MAO-B) ligands. The designed derivatives consist of either cinnamyl or 2-fluorocinnamyl moieties connected to 2-fluoropyridylpiperazines. The three-step synthesis starting from commercially available piperazine afforded the final products in overall yields between 9% and 29%. An in vitro competitive binding assay using l-[3H]Deprenyl as radioligand was developed and the MAO-B binding affinities of the synthesized derivatives were assessed. Docking studies revealed that the compounds 8–17 were stabilized in both MAO-B entrance and substrate cavities, thus resembling the binding pose of l-Deprenyl. Although our results revealed that the novel fluorinated cinnamylpiperazines 8–17 do not possess sufficient MAO-B binding affinity to be eligible as positron emission tomography (PET) agents, the herein developed binding assay and the insights gained within our docking studies will certainly pave the way for further development of MAO-B ligands.

The aim of this study was to develop highly specific radiofluorinated ligands for quantitative positron emission tomography (PET) imaging of monoamine oxidase-B (MAO-B) in brain. A series of 8 fluoro derivatives of 1-cinnamyl-4-arylpiperazine were synthesized by standard methods of organic synthesis. The affinity of the compounds was determined in a competitive binding assay using L-[3H] deprenyl as radioligand on rat brain homogenates. The KD of the radioligand was determined by homologous competition. An efficient, three-step procedure for the synthesis of the potential MAO-B ligands was developed. A competitive binding assay was established, using L-[3H]deprenyl as the radioligand, and rat brain membrane homogenate. The compounds were screened (three concentrations 10-9, 10-7 and 10-5) for their MAO-B affinity. We successfully synthesized a series of fluorinated MAO-B ligands. Unfortunately, their affinities toward MAO-B have proved to be rather low. To increase the affinity further modifications are needed.

Serotonin 5HT1a receptor belongs to a class of G-protein coupled receptors. It serves as a potential target for neurological disorders such as depression, anxiety etc. It is a well-known fact that N-arylpiperazine moiety is present in compounds with pronounced 5HT1a activity. Taking into account previously published results1 novel structures of N-{4-[2-(4- arylpiperazin-1-yl)ethyl]phenyl}arylamides (Figure 1.) were designed for target synthesis. Proposed modifications include: different position of hydroxyl group in aryl amide part of molecule and addition of methoxy and chloro substituents to the phenyl ring of parent compounds, since their introduction in the molecule leads to increased receptor affinity. New compounds were synthesized by acylation of N-arylpiperazines using 4- nitrophenylacetic acid. Obtained amides were converted in 1-(4-nitrophenethyl)-4- arylpiperazines using diborane in THF. Reduction of nitro compounds by Ra/Ni provided 1- (4-aminophenethyl)-4-arylpiperazines. Target arylamides were obtained by condensation 1- (4-aminophenethyl)-4-arylpiperazines with corresponding aryl acids in presence of propylphosphoric acid anhydride (PPAA) in DMF. All newly synthesized compounds were evaluated for their activity toward 5HT1a receptors by in vitro competitive displacement assay of [3H] 8-OH-DPAT. HEK cell line were used as a source of 5HT1a receptors. Introduction of 2-methoxy and 2,3-dichloro groups,as well as meta and para hydroxyl group in molecule resulted in increment of affinity toward 5HT1a receptors comparing to the parent compounds.

A total of 14 novel arylpiperazines were synthesized, and pharmacologically evaluated by measuring their affinities towards the D2 dopamine receptor (D2DR) in a [3H]spiperone competition assay. All herein described compounds consist of a benzimidazole moiety connected to the N-(2-methoxyphenyl)piperazine via linkers of various lengths. Molecular docking analysis and molecular dynamics simulations were performed on D2DR-arylpiperazine complexes with an objective to explore the receptor-ligand interactions and properties of the receptor binding site. Crystal structure of D2DR that has been published recently was used throughout this study. The major finding is that high affinity arylpiperazines must interact with both the orthosteric binding site and the extended binding pocket of D2DR and thereforeshould contain a linker of 5 or 6 methylene groups long. This is the peer-reviewed version of the article: [https://doi.org/10.2298/JSC181029104P] [http://cer.ihtm.bg.ac.rs/handle/123456789/3097]

A scalable, cost-efficient and simple synthetic pathway towards potential bivalent opioid/dopamine receptor ligands was developed and optimized. Three novel compounds that contain both opioid and dopamine pharmacophores linked by the four methylene group chain were synthesized in 33, 35 and 39 % overall yield after a four-step synthetic route starting from three commercially available N-aryl piperazines. The anilino piperidine precursor was easily prepared in three steps, as previously published, starting from 4- piperidone. The synthesis presented in this paper could be of interest for heterocyclic and general organic chemistry. The newly designed compounds possessing two pharmacophores, opioid and D2/D3, are potentially useful pharmacological probes. Of particular interest would be the simultaneous binding to both opioid and D2/D3 receptors, and the resulting pharmacological responses may be useful for the further understanding of tolerance and dependence phenomena in opioid clinical use and/or abuse. Три нова једињења, која садрже опиоидну и допаминску фармакофору, спојене преко четири метиленске групе, синтетисанa су у четири фазе у укупном приносу од приближно 35 %, полазећи од три комерцијално доступна N-арилпиперазина. Анилино- пиперидински прекурсор је добијен у три једноставне реакционе фазе, полазећи од 4-пиперидона, према познатој литературној методи. Синтеза приказана у овом раду може бити од значаја за хетероцикличну и органску хемију у целини. Новосинтетисана једињења која поседују две фармакофоре, опиоидну и Д2/Д3, су потенцијално корисни супстрати за фармаколошка испитивања. Од посебног интереса може бити истовремено везивање за опиоидне и Д2/Д3 рецепторе, а резултујући фармаколошки одговор био би користан у даљем разумевању толеранције и зависности, као феномена везаних за кли- ничку употребу и/или злоупотребу опијата.

The ratio of affinities toward the dopamine D-2 and the 5-hydroxytryptamine 5-HT1A receptors is one of the important parameters that determine the efficiency of antipsychotic drugs. Here, we present the synthesis of ortho-, meta-, and para-N-{[2-(4-phenyl-piperazin-1-yl)-ethyl]-phenyl}-arylamides and their structure-activity relationship studies on dopamine D-2 and 5-hydroxytryptamine 5-HT1A receptors. It was shown that the biological activity of the described ligands strongly depends on their topology as well as on the nature of the heteroaryl group in the head of the molecules. Docking simulations together with conformational analysis revealed a rational explanation for the ligands' behavior. The molecular model of receptor-ligand interactions described herein provided us with a tool for the rational design of new compounds with a favorable D-2/5-HT1A profile. Supplementary material: [http://cherry.chem.bg.ac.rs/handle/123456789/3511]

Second extracellular loop (ecl2) of the dopamine (DA) D2 receptor is an essential part of the binding pocket of dopaminergic ligands. To form a part of the ligand-binding surface, it has to fold down into the transmembrane domain of the DA receptor. The current study describes the modeling of the D2 DA receptor ecl2 and its interactions with arylpiperazine ligands. In order to model the D2 DA receptor ecl2, several arylpiperazine ligands were used to propose a pharmacophore model. D2 DA receptor ecl2 model was built using Accelrys Discovery Studio. To test the proposed model, docking analysis was performed and key amino acid residues were determined. The proposed receptor-ligand interactions were rationalized and compared with measured binding affinities. It is shown that D2 DA receptor ecl2 significantly participates in the formation of the receptor-ligand complex through aromatic, hydrophobic and polar interaction. Considering them would benefit molecular modeling of G- protein-coupled receptors (GPCRs) and facilitate the design of novel active compounds. Druga ekstracelularna petlja dopaminskog D2 receptora je esencijalni deo vezivnog mesta receptora. Da bi se definisala gornja strana vezivnog mesta, ona mora da se savije nadole, i orijentiše ka transmembranskom domenu receptora. U ovom radu opisan je proces modelovanja druge ekstracelularne petlje dopaminskog D2 receptora i njene interakcije sa arilpiperazinskim ligandima. Za modelovanje je korišćen Accelrys Discovery Studio paket programa. Predloženi model je testiran doking analizom literaturno dostupnih liganada i poređenjem dobijenih rezultata sa njihovim afinitetom vezivanja za D2 receptor. Određeni su amino-kiselinski ostaci koji stupaju u interakcije sa ligandima. Ključne interakcije su definisane i upoređene sa afinitetima liganada prema receptoru kako bi se predloženim modelom objasnile razlike u eksperimentalnim rezultatima. Naša istraživanja su pokazala da druga ekstracelularna petlja dopaminskog D2 receptora može stupati u različite interakcije sa arilpiperazinskim ligandima koje između ostalih uključuju hidrofobne, aromatične interakcije ali i vodnonične veze. Ova saznanja, u kombinaciji sa predloženim modelom D2 receptora, koji uključuje ekstracelularne petlje, mogu biti od velike koristi prilikom budućeg dizajna novih dopaminergičkih liganada.

Molecular modelling studies were undertaken in order to identify key interactions of selected ligands with α1A adrenergic receptor, responsible for their binding and presumably receptor activation. The previously made model of α1A adrenergic receptor was optimized by molecular dynamics and different arylpiperazine ligands were docked. The results show a high correlation to the experimentally determined binding affinities. Ligand orientations and its interactions with specific amino acid residues in the binding site explain trends in its structure-activity relationship. The key interactions for those trends are mainly aromatic, which are suggested by the calculation of their ESP surfaces.

Second extracellular loop (ec12) of the dopamine (DA) D2 receptor is an essential part of the binding pocket of dopaminergic ligands. To form a part of the ligand-binding surface, it has to fold down into the transmembrane domain of the DA receptor. The current study describes the modeling of the D2 DA receptor ec12 and its interactions with arylpiperazine ligands. In order to model the D2 DA receptor ec12, several arylpiperazine ligands were used to propose a pharmacophore model. D2 DA receptor ec12 model was built using Accelrys Discovery Studio. To test the proposed model, docking analysis was performed and key amino acid residues were determined. The proposed receptor ligand interactions were rationalized and compared with measured binding affinities. It is shown that D2 DA receptor ec12 significantly participates in the formation of the receptor ligand complex through aromatic, hydrophobic and polar interaction. Considering them would benefit molecular modeling of G-protein-coupled receptors (GPCRs) and facilitate the design of novel active compounds. Druga ekstracelularna petlja dopaminskog D2 receptora je esencijalni deo vezivnog mesta receptora. Da bi se definisala gornja strana vezivnog mesta, ona mora da se savije nadole, i orijentiše ka transmembranskom domenu receptora. U ovom radu opisan je proces modelovanja druge ekstracelularne petlje dopaminskog D2 receptora i njene interakcije sa arilpiperazinskim ligandima. Za modelovanje je korišćen Accelrys Discovery Studio paket programa. Predloženi model je testiran doking analizom literaturno dostupnih liganada i poređenjem dobijenih rezultata sa njihovim afinitetom vezivanja za D2 receptor. Određeni su amino-kiselinski ostaci koji stupaju u interakcije sa ligandima. Ključne interakcije su definisane i upoređene sa afinitetima liganada prema receptoru kako bi se predloženim modelom objasnile razlike u eksperimentalnim rezultatima. Naša istraživanja su pokazala da druga ekstracelularna petlja dopaminskog D2 receptora može stupati u različite interakcije sa arilpiperazinskim ligandima koje između ostalih uključuju hidrofobne, aromatične interakcije ali i vodnonične veze. Ova saznanja, u kombinaciji sa predloženim modelom D2 receptora, koji uključuje ekstracelularne petlje, mogu biti od velike koristi prilikom budućeg dizajna novih dopaminergičkih liganada.

Knowledge of three-dimensional protein structure is of the paramount importance for determination of receptor and enzyme action mechanism in interaction with other molecules. In spite of growing number of solved 3D protein structures, number of determined protein sequences exceeds by far the number of defined spatial protein structures. Besides, sequence databases grow more rapidly then databases of 3D protein structures. When crystal protein structures or data obtained by NMR are not available, we have to reach for next 'best' possible solution - modeling of the protein spatial structure. One of the aspects that can be applied to obtain usable protein model is homology modeling. This method is based on the design of the three-dimensional structure of the unknown protein based on layout of the homolog protein of known 3D structure. To get a big picture of this method, in this text, we explained few aspects of homology modeling.

Our previously described research on docking analysis of a series of isosteric N4-arylpiperazines on a model of 5-HT1A receptor was used earlier to investigate interactions of different ligands with the receptor binding site. Due to the limitations of molecular mechanics (MM) methods, docking analysis failed to give precise results about interactions that influence binding affinity of the ligands, but we presumed that aromatic-aromatic interactions, or edge-to-face, to be more precise, play an important role in the binding process. In order to further elaborate on this hypothesis, ab initio approach was used to calculate possible edge-to-face interactions on a model system and correlate them to ligand affinity. Obtained results indicate that those dispersive interactions can show notable influence on the binding of the ligands to 5-HT1A receptor. Stabilization energies of modeled receptor-ligand complex, calculated using Becke's "half-and-half" hybrid DFT method showed strong correlation with the affinity of investigated ligands towards 5-HT1A receptor.

The present study identifies xanthones gentiakochianin and gentiacaulein as the active principles responsible for the in vitro antiglioma action of ether and methanolic extracts of the plant Gentiana kochiana. Gentiakochianin and gentiacaulein induced cell cycle arrest in G(2)/M and G(0)/G(1) phases, respectively, in both C6 rat glioma and U251 human glioma cell lines. The more efficient antiproliferative action of gentiakochianin was associated with its ability to induce microtubule stabilization in a cell-free assay. Both the xanthones reduced mitochondrial membrane potential and increased the production of reactive oxygen species in glioma cells, but only the effects of gentiakochianin were pronounced enough to cause caspase activation and subsequent apoptotic cell death. The assessment of structure-activity relationship in a series of structurally related xanthones from G. kochiana and Gentianella austriaca revealed dihydroxylation at positions 7, 8 of the xanthonic nucleus as the key structural feature responsible for the ability of gentiakochianin to induce microtubule-associated G(2)/M cell block and apoptotic cell death in glioma cells. (C) 2008 Elsevier Ltd. All rights reserved. null

Eight new compounds with halogen atom introduced into the benzimidazole- 2-thione dopaminergic pharmacophore of 5-[2-(4-arylpiperazin-1-yl)ethyl]-1,3-dihydro- 2H-benzimidazole-2-thiones with the arylpiperazine part of the molecule being selected according to known structure-affinity requirements, have been synthesized. All the new compounds were evaluated for the in vitro binding affinity at the dopamine (DA) D1 and D2 and serotonin 5-HT1A receptors by the competitive radioassays, performed on synaptosomal membranes prepared from fresh bovine caudate nuclei and hippocampi. All the new compounds were strong competitors for the binding of the radioligands to the D2 and 5-HT1A receptors, with the most active of them having 34 and 170 time higher affinity than non-halogenated congeners in the D2 DA receptor radioassays (compounds 9.1b and 9.2b, respectively). Divergently, these compounds were without significant affinities for the D1 DA receptors. . Sintetisano je osam novih jedinjenja kod kojih je atom halogena uveden u benzimidazol-2-tionsku dopaminergičku farmakoforu 5-[2-(4-arilpiperazin-1-il)etil]-1,3-dihidro-2N-benzimidazol-2-tiona sa arilpiperazinskim delom molekula izabranim shodno pozna- tim zahtevima o odnosu strukture i reaktivnosti. Za sva novosintetisana jedinjenja je određen afinitet vezivanja za dopaminske (D1 i D2) i 5-NT1A receptore u in vitro eksperimentima kompeticije sa radioligandima. Kao izvor dopaminskih i 5-NT1A receptora su korištene sinaptozomalne membrane izolovane iz goveđeg nukleusa kaudatusa i hipokampusa. Sva novosintetisana jedinjenja pokazala su se kao jaki kompetitori [3H]spiperona i [3H]8-OH-DPAT, od kojih najaktivnija (9.1b i 9.2b) poseduju 34 i 170 puta veći afinitet ka D2 DA receptorima od polaznih, nehalogenovanih jedinjenja. Sa druge strane, ova jedinjenja ne poseduju značajan afinitet ka D1 dopaminskim receptorima. . null

Scrotonin receptors (5-HTRs), especially the 5-HT1A subtype, have been the subject of intensive research for the past decade, due to their function in human physiology. Several structurally different classes of ligands are known to bind to the 5-HT1A receptor, but arylpiperazine derivatives are among the most important ligands. In the work, docking analyses were used to explain the binding affinities of a series of ligands with different N-1 substituent. All ligands had ill common the arylpiperazine structure, while the N-1 substituent was modified to investigate the influence of ligand structure on its binding affinity. The shape and size, as well as the rigidity of the substituents were altered to investigate the possible effects on the formation of the receptor - ligand complex. Serotoninski receptori su, a naročito 5-HT 1A pod tip, zbog značajne uloge u fiziologiji ljudskog organizma, predmet intenzivnog izučavanja tokom protekle decenije. Poznato je da se za 5-HT 1A receptor vezuje nekoliko strukturno različitih klasa liganada, ali su arilpiperazinski derivati među najznačajnijim. Da bi objasnili vezivanje serije liganada sa različitim N-1 supstituentima za receptor koristili smo analizu vezivanja (docking analizu). Svi ligandi su imali zajedničku arilpiperazinsku strukturu dok su im N-1 supstituenti modifikovani tako što je menjan oblik, veličina kao i krutost supstituenta da bi se istražio njihov eventualni uticaj na formiranje kompleksa receptor - ligand.

In order to find the essential structural motif of the D2L dopamine receptor necessary for the interaction with a-subunit of Gi1 protein, four fragments of the third cytoplasmic loop (CPL3) of this receptor were cloned, expressed in E. coli and purified. After that, fusion proteins with glutathione-S-transferase (GST) were prepared and the interactions quantified by a colorimetric assay for GST activity determination. The presence of D2L-CPL3 fragment-Gia1 complexes was detected by SDS-polyacrylamide gel electrophoresis (PAGE). Kd values for the interaction of the three fragments with Gia1 were similar and in nmol/L range of concentrations, while the peptide representing the insert in the long form of the dopamine D2 receptor expressed about 10-fold lower binding affinity. These results could serve to design new therapeutic agents that might act at the level of receptor/G protein interaction rather than at the level of ligand-receptor binding. U cilju pronalaženja bitnih strukturnih motiva potrebnih za interakciju sa a podjedinicom Gi1 proteina klonirana su, eksprimirana i prečišćena 4 fragmenta treće citoplazmatične petlje (CPL3) dopaminskog D2L receptora koji su dalje pripremljeni kao fuzioni proteini sa glutation-S-transferazom (GST). Interakcije su kvantifikovane bojenom reakcijom za određivanje aktivnosti GST. Postojanje kompleksa D2L-CPL3 fragment- Gia1 je dokazano elektroforetskom analizom na SDS-poliakrilamidnom gelu (PAGE). Kd vrednosti za tri fragmenta su bile vrlo slične i u nmol/L opsegu koncentracija, dok je peptid koji predstavlja insert u dugom obliku dopaminskog D2 receptora posedovao oko 10 puta manji afinitet vezivanja za Gia1. Ovi rezultati mogu biti osnova za sintezu novih terapeutskih agenasa koji bi delovali na nivou interakcije receptora i G proteina umesto na nivou vezivanja liganda za receptor.

1-(2-Heteroarylalkyl)-4-phenylpiperazines containing methyl group in either the alpha- or the beta-position of the side alkyl chain were synthesized as racemic mixtures. They were evaluated for in vitro binding affinity at the D-1 and D-2 dopamine and 5-HT1A serotonin receptors using synaptosomal membranes of the bovine caudate nucleus and hippocampus, respectively, as a source of the corresponding receptors. Tritiated SCH 23390 (D-1 receptor-selective), spiperone (D-2 receptor-selective), and 8-OH-DPAT (5-HT1A receptor-selective) were employed as the radioligands. None of the new compounds expressed significant affinity for the D-1 receptor. Introduction of the methyl group into the beta-position of the parent molecules increased the affinity for the D-2 receptor (10b-13b), and decreased the affinity for the 5-HT1A receptor with the exception of imidazole (11b) which was a rather efficient displacer of 8-OH-DPAT. Most potent of the newly synthesized compounds in [H-3]spiperone assay were compounds (+/-)6-[1-methyl-2-(4-phenylpiperazin-n-1-yl)-ethyl]-1, 4-dihydroquinoxaline-2,3-dione (10b), K-d = 6.0 nM and (+/-)5-[1-methyl-2-(4-phenylpiperazin-1-yl)-ethyl]-1,3-dihydrobenzoirnidazol-2-thione (13b), K-d = 5.3 nM. However, compounds containing methyl group in alpha-position (10a-13a) of the parent molecules expressed a decreased affinity for the D-2 receptor, while the affinity for the 5-HT1A receptor remained in the same range of concentrations as that of closely related achiral parent compounds (14-17) run in the same binding assays as references.

Several 2-methylthiomethylbenzimidazoles (3a-e) and the corresponding sulphinyl derivatives 4a-e were synthesized and evaluated measuring their in vitro binding affinity at the D 1 and D 2 dopamine (source: synaptosomal membranes of the bovine nucleus caudatus) and 5-HT(1A) serotonin (source: synaptosomal membranes of the bovine hippocampus) receptors. [ 3 H]SCH 23390, [ 3 H]spiperone, and [ 3 H]-8-OH-DPAT were employed as specific radioligands for the D 1 , D 2 and 5-HT(1A) receptors, respectively. None of the compounds except for 3b acting as a moderate [ 3 H]SCH 23390, competitor, expressed binding affinity at the D 1 receptor. Compounds 4a and 4e were inactive displacers of both [ 3 H]spiperone and [ 3 H]-8-OH-DPAT. Ligands 4b, 3d and 4d acted as weak to moderate [ 3 H]spiperone competitors and 3a was a weak [ 3 H]- 8-OH-DPAT displacer. The remaining ligands expressed binding affinity at the corresponding receptors in a nanomolar concentration range. Among them, compound 3b with K(i) of 14.2 nM and 8.4 nM in [ 3 H]spiperone and [ 3 H]-8- OH-DPAT binding assay, respectively, was the most potent mixed dopaminergic/serotonergic ligand. Although sterically similar, the two classes of ligands differ with regard to electronic properties of substituents in position 2 of the benzimidazole ring. Oxidation of 2- (methylthiomethyl)benzimidazoles afforded ligands devoid of binding affinity at the 5-HT(1A) receptor and significantly reduced binding affinity at the D 2 receptor. This points to the importance of electronic properties of substituents in position 2 of benzimidazole ring for the D 2 /5-HT(1A) affinity ratio of this type of ligands.

With an aim of creating new, high affinity dopaminergic ligands, six different 3- and 4-substituted 1-{2-[5-(1H-benzimidazole-2-thione)]ethyl}piperidines and nine related heterocyclic congeners were synthesized and evaluated for in vitro binding affinity at D 1 and D 2 dopamine receptors. Synaptosomal membranes prepared from fresh bovine caudate nuclei were used as a source of the dopamine receptors. Only 4-[bis-(4-fluorophenyl)methylene]-piperidines, compounds 9e, 10d, and 11d, expressed moderate affinity for the D 1 receptors, while all other compounds were inactive competitors of [ 3H]SCH 23390. Compounds 9c, 9d, 10c, 11a, and 11c were inactive in the D 2 receptor binding assay, as well. Derivatives of 4-phenylpiperidine (9-11b) and 3-phenyl-piperidine (10a) expressed a moderate to low affinity for the D 2 receptors. However, racemic (±)-1-{2-[5-(1H-benzimidazole-2-thione)]ethyl}-3-phenylpiperidine 9a and its enantiomer (+)-9a behaved as selective, high affinity D 2 receptor ligands, the latter being some four times more active than the racemate.

Cilj našeg rada bio je ispitivanje biološke aktivnosti metilamino- i metoksiderivata avarona i razjašnjavanje mehanizma njihovog biološkog dejstva. Za sintezu su izabrani derivati za koje se očekivalo da će imati negativniji polutalasni potencijal od avarona i samim tim pokazivati povećanu aktivnost. Sintetisani su 4'-(metilamino)-avaron, 3'-(metilamino)-avaron i 3'-metoksi-avaron. Antibakterijska aktivnost dobijenih derivata je ispitivana prema gram-pozitivnim bakterijama: Bacillus subtilis, Clostridium sporogenes, Streptosporangium longisporum, Micrococcus flavus, Sarcina lutea i Staphylococcus aureus, prema gramnegativnim bakterijama: Klebsiella pneumoniae, Proteus vulgaris, Pseudomonas aeruginosa, Salmonella enteritidis, Escherichia coli i prema kulturama gljivica: Aspergillus niger, Candida albicans i Saccharomyces cerevisiae disk-difuzionom metodom. Takođe, ispitivana je i toksičnost derivata na račiće Artemia salina. Potencijalna antioksidativna aktivnost avarona i dobijenih derivata ispitivana je testom sa DPPH. Antitumorska aktivnost ispitivana je za sve derivate na osam vrsta ćelija raka (ćelijske linije raka grlića materice, melanoma i leukemije, rak dojke pozitivan na estrogeni receptor, rak dojke negativan na estrogeni receptor, rak pluća, leukemija T-ćelija i promijelocitna leukemija) pri čemu je ispitivana i njihova citotoksičnost na limfocite. Svim hinonskim derivatima hemijski je modifikovan model-enzim lizozim. Dobijenim modifikatima lizozima je nakon modifikacije određena enzimska aktivnost. Sama modifikacija je praćena UV/Vis spektrofotometrijom, SDS elektroforezom i masenom spektrometrijom. Mesto vezivanja hinonskih derivata za molekul lizozima određeno je tehnikom MALDI TOF posle tripsinske digestije modifikata. S obzirom na uočeno vezivanje avaronskih derivata za ε-amino grupu lizina (Lys-97) u lizozimu, sintetisano je jedinjenje 4'-((5-tert-butoksikarbonil)amino)-5-karboksipentil)amino)- avarona, koje je poslužilo kao model jedinjenje za dalja ispitivanja biološke aktivnosti lizozim-hinonskog adukta... aim of our work has been investigation of biological activity of methylamino- and metoxy- derivatives of avarone, their bioconjugates of lysozyme and study of the mechanism of their biological action. For synthesis were chosen derivatives for which it was expected to have more negative half-wave potential than avarone and therefore a higher activity. The selected compounds are 3’-methylamino, 4’-methylamino- and 3’-methoxyavarone. Antimicrobial activity of the synthesized derivatives was investigated towards Gram positive bacteria: Bacilus subtilis, Clostridium sporogenes, Sreptosporangium longisporum., Micrococcus flavus, Sarcina lutea and Staphylococcus aureus, Gram negative bacteria: Klebsiella pneumoniae, Proteus vulgaris, Pseudomonas aeruginosa, Salmonella enteritidis and Escherichia coli and fungi cultures: Aspergilus niger, Candida albicans and Sacharomyces cerevisiae, all by disc diffusion method. Toxicity against Artemia salina nauplii was surveyed as well. Antioxidant activity of avarone and its derivatives was assessed by DPPH assay. Antitumor activity was determined for all derivatives towards eight lines of tumor cells (myelogenous leukemia (K562), cervix carcinoma (HeLa), human malignant melanoma cells (Fem-X), Jurkat T cell leukemia, estrogen receptor negative breast carcinoma (MDA-MB-231), estrogen receptor positive breast carcinoma (MCF7), human fetal lung fibroblast (MRC-5) and human promyelocytic leukemia (HL-60)). For all derivatives toxicity towards lymphocytes was determined. Model enzyme lysozyme was modified with the synthesized quinones and for all the obtained bioconjugates MIC value towards Gram positive and Gram negative bacteria were determined. Modification reaction was monitored by UV/VIS spectrophotometry, SDS electrophoresis and mass spectrometry. Binding position of quinone derivatives on lysozyme was determined by MALDI TOF spectrometry after trypsin digestion. Since the avarone derivatives were found to bind to lysine (Lys-97) in lysozyme, 4’-((5-((tert-butoxycarbonyl)amino)-5-carboxypentyl)-amino)avarone has been synthesized as a model compound for further investigation of the biological activity of lysozyme―quinone aduct..