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Buspirone free base

Anna E. Koziol, Zdzislaw Chilmonczyk, Jacek Cybulski
2006 Acta Crystallographica Section E  
Although the crystal structures of the stable form of (I) and several analogues of (I) have been determined (Chilmonczyk et al., 1995 (Chilmonczyk et al., , 1997)) , crystals of the free base, (I), have  ...  Theoretical calculations for the gas phase indicated that the fully extended conformations of the free base and cation of (I) are those of minimum energy (Chilmonczyk et al., 1995) .  ... 
doi:10.1107/s1600536806047726 fatcat:2fztwv6r4bhyneiudrkpyfpjxm

Proapoptotic effects of novel pentabromobenzylisothioureas in human leukemia cell lines

Mirosława Koronkiewicz, Zdzisław Chilmonczyk, Zygmunt Kazimierczuk
2011 Medicinal Chemistry Research  
A series of new pentabromobenzylisothioureas [ZKK-1-ZKK-5; (ZKKs)] carrying additional substituents on nitrogen atoms has been synthesized. The ZKKs were found to induce apoptosis in HL-60 (human promyleocytic leukemia) and K-562 (human chronic erythromyeloblastoid leukemia) cell lines in a concentration-dependent manner at low micromolar concentrations. ZKK-3 [(N,N 0dimethyl-S-2,3,4,5,6-pentabromobenzyl)isothiouronium bromide] showed the highest proapoptotic activity in HL-60 cells, whereas
more » ... -2 [N-methyl-S-(2,3,4,5,6pentabromobenzyl)isothiouronium bromide] was most effective in this respect in K-562 cells. During the ZKKsinduced apoptosis, an 85 kDa fragment of cleaved PARP (caspase-3 and caspase-7 substrate) was detected in both cell lines tested. The studied compounds also decreased mitochondrial transmembrane potential in both these cell lines and caused the cells to accumulate in G 1 and at the G 1 /S border of the cell cycle in a concentration-dependent manner. These results show promise for their study as new compounds in the treatment of leukemia, after an appropriate preclinical toxicity profile.
doi:10.1007/s00044-011-9841-8 pmid:22942618 pmcid:PMC3427706 fatcat:nxbz7expkraotlopotjj5hbski

Functional Selectivity and Antidepressant Activity of Serotonin 1A Receptor Ligands

Zdzisław Chilmonczyk, Andrzej Bojarski, Andrzej Pilc, Ingebrigt Sylte
2015 International Journal of Molecular Sciences  
Serotonin (5-HT) is a monoamine neurotransmitter that plays an important role in physiological functions. 5-HT has been implicated in sleep, feeding, sexual behavior, temperature regulation, pain, and cognition as well as in pathological states including disorders connected to mood, anxiety, psychosis and pain. 5-HT1A receptors has for a long time been considered as an interesting target for the action of antidepressant drugs. It was postulated that postsynaptic 5-HT1A agonists could form a new
more » ... class of antidepressant drugs, and mixed 5-HT1A receptor ligands/serotonin transporter (SERT) inhibitors seem to possess an interesting pharmacological profile. It should, however, be noted that 5-HT1A receptors can activate several different biochemical pathways and signal through both G protein-dependent and G protein-independent pathways. The variables that affect the multiplicity of 5-HT1A receptor signaling pathways would thus result from the summation of effects specific to the host cell milieu. Moreover, receptor trafficking appears different at pre-and postsynaptic sites. It should also be noted that the 5-HT1A receptor cooperates with other signal transduction systems (like the 5-HT1B or 5-HT2A/2B/2C receptors, the GABAergic and the glutaminergic systems), which also contribute to its antidepressant and/or anxiolytic OPEN ACCESS Int. J. Mol. Sci. 2015, 16 18475 activity. Thus identifying brain specific molecular targets for 5-HT1A receptor ligands may result in a better targeting, raising a hope for more effective medicines for various pathologies. Postsynaptic 5-HT1A heteroreceptors are expressed in target areas receiving serotonergic innervation. They are located on non-serotonergic neurons, primarily in the limbic areas, such as on the dendrites and soma of glutamatergic pyramidal neurons, and axon terminals of GABAergic and cholinergic neurons. The heteroreceptors are particularly enriched in the hippocampus, where immunohistochemistry and radioligand binding have demonstrated high receptor levels in the stratum radiatum of CA1 and the granule cell layer of the dentate gyrus, and moderate levels in CA3. 5-HT1A heteroreceptors are also highly expressed in the entorhinal cortex, frontal cortex, and lateral septum, and moderately expressed in the amygdala, superior colliculus, piriform cortex, and interpeduncular nucleus, as well as in several hypothalamic and thalamic nuclei. Activation of 5-HT1A heteroreceptors on these distinct neurons mediates hyperpolarizing response to released serotonin and usually reduces neuronal excitability and firing [10, 14] . Aside from 5-HT1A, the 5-HT1B receptor is also thought to serve as an autoreceptor. 5-HT1B receptors are expressed in the central nervous system (CNS), concentrated in the basal ganglia, striatum, and frontal cortex. In addition, the receptor may also act as a terminal heteroreceptor controlling the release of other neurotransmitters, such as acetylcholine, glutamate, dopamine, noradrenaline, and γ-aminobutyric acid [15, 16] . The receptor is also found on cerebral arteries and other vascular tissues. The putative 5HT1B receptor agonist, anpirtoline, has analgesic and antidepressant-like properties in rodents, and 5-HT1B receptor KO mice were reported to be both highly aggressive and have an increased preference for alcohol [16, 17] . Studies investigating the relationship between 5-HT1A and 5-HT1B receptors showed that 5-HT1A/1B knockouts had increased extracellular serotonin in the hippocampus, suggesting that the pairing of SSRI with 5-HT1A/1B antagonist might prove to be a potent treatment for anxiety and depression [18] . 5-HT1B receptor agonist (CP 94253) and antagonist (SB 216641) have been shown to be effective in preclinical models of anxiety and CP 94253 was also effective in the model of depression (forced swim test-FST) [19] . It was also shown that the activation of 5-HT1B heteroceptors induces antidepressant-like effect in mice [20] . 5-HT 1A Receptor Regulated Transcription Pathways Adenylate Cyclase The primary coupling linkage of the 5-HT1A receptor (and of all 5-HT1 receptors) is to the inhibition of adenylate cyclase (AC) and decrease protein kinase A (PKA) activity. However, the 5-HT1A receptor couples to the broadest panel of second messengers of any of the 5-HT receptors [21] . This receptor has been reported to activate or inhibit various enzymes, channels, and kinases, and to stimulate or inhibit production of diverse soluble second messengers. The receptor has been found to inhibit and activate AC and phospholipase C (PLC), to stimulate nitric oxide synthase (NOS) and a nicotinamide adenine dinucleotide phosphate (NADP) oxidase-like enzyme, to activate K + channels and high conductance anion channels, to inhibit Ca 2+ conductance and inhibit or stimulate Ca 2+ mobilization, and regulate a number of channels and transporters (Figure 2 ) [22] [23] [24] . The 5-HT1A receptor can activate protein kinase C (PKC), Src kinase, and mitogen-activated protein kinases (MAPKs), activate or inhibit phosphatidyl
doi:10.3390/ijms160818474 pmid:26262615 pmcid:PMC4581256 fatcat:tu4z5y3hrfeynhzhpggdshyx4q

4-{3-[(4-Fluorobenzylidene)amino]-2-oxo-1,3-oxazolidin-5-ylmethyl}morpholine

Rafal Kruszynski, Tadeusz J. Bartczak, Zdzislaw Chilmonczyk, Jacek Cybulski
2001 Acta Crystallographica Section E  
Bartczak, Zdzislaw Chilmonczyk and Jacek Cybulski S1. Comment Acta Cryst. (2001). E57, o386-o388 Figure 1 1 Figure 1 Figure 2 2 Figure 2 Fit of inverted (I) and (II).  ...  In the Pharmaceutical Research Institute in Warsaw, a series of derivatives of 3-amino-2-oxazolidinone have been prepared (Chilmonczyk et al., 1997) .  ... 
doi:10.1107/s1600536801005487 fatcat:77pci5q6a5dkpi6ws4iakp3bje

4-[3-(4-Chlorobenzylideneamino)-2-oxooxazolidin-5-ylmethyl]morpholin-4-ium chloride monohydrate

Tadeusz J. Bartczak, Rafal Kruszynski, Zdzislaw Chilmonczyk, Jacek Cybulski
2001 Acta Crystallographica Section E  
Bartczak, Rafal Kruszynski, Zdzislaw Chilmonczyk and Jacek Cybulski S1.  ...  Experimental The title compound was prepared according to the method of Chilmonczyk et al. (1997) .  ... 
doi:10.1107/s1600536801004548 fatcat:baf5m2ekmvdcnpwfiiudcbybse

Mass-spectrometric studies of new 6-nitroquipazines—serotonin transporter inhibitors

Janina Witowska-Jarosz, Małgorzata Jarończyk, Aleksander P. Mazurek, Ingebrigt Sylte, Andrzej J. Bojarski, Zdzisław Chilmonczyk, Maciej Jarosz
2011 Analytical and Bioanalytical Chemistry  
Six synthesized 6-nitroquipazine derivatives were examined by electron ionization (EI) and electrospray ionization (ESI) mass spectrometry in positive and negative ion mode. The compounds exhibit high affinity for the serotonin transporter (SERT) and belong to a new class of SERT inhibitors. The EI mass spectra registered in negative ion mode showed prominent molecular ions for all the compounds studied. All EI mass spectra and all ESI mass spectra showed similar fragmentation pathways of
more » ... lar ions, but the pathways differed between EI and ESI. The differences were explained with the aid of theoretical evaluation of the stability of the respective radical ions (EI MS) and protonated ions (ESI MS).
doi:10.1007/s00216-011-5410-8 pmid:21960253 pmcid:PMC3249537 fatcat:m64qy6buzvbrvcj4ondq5csqhu

In Vitro Evaluation of Sulforaphane and a Natural Analog as Potent Inducers of 5-Fluorouracil Anticancer Activity

Małgorzata Milczarek, Lidia Mielczarek, Katarzyna Lubelska, Aleksandra Dąbrowska, Zdzisław Chilmonczyk, Dariusz Matosiuk, Katarzyna Wiktorska
2018 Molecules  
Isothiocyanates (R-NCS) are sulphur-containing phytochemicals. The main source are plants of the Brassicaceae family. The best known plant-derived isothiocyanate is sulforaphane that has exhibited anticancer activity in both in vivo and in vitro studies. Recent attempts to expand their use in cancer therapy involve combining them with standard chemotherapeutics in order to increase their therapeutic efficacy. The aim of this paper is to determine the impact of sulforaphane and its natural
more » ... alyssin on the anticancer activity of the well-known anticancer drug 5-fluorouracil. The type of drug-drug interactions was determined in prostate and colon cancer cell lines. Confocal microscopy, western blot and flow cytometry methods were employed to determine the mechanism of cytotoxic and cytostatic action of the combinations. The study revealed that additive or synergistic interactions were observed between 5-fluorouracil and both isothiocyanates, which enhanced the anticancer activity of 5-fluorouracil, particularly in colon cancer cell lines. An increased cytostatic effect was observed in case of alyssin while for sulforaphane the synergistic interaction with 5-fluorouracil involved an intensification of apoptotic cell death.
doi:10.3390/molecules23113040 pmid:30469330 pmcid:PMC6278648 fatcat:p63stc7xhvanzctls2nayiiv3q

Fullerene Derivatives of Nucleoside HIV Reverse Transcriptase Inhibitors—In Silico Activity Prediction

Aleksandra Dąbrowska, Tomasz Pieńko, Przemysław Taciak, Katarzyna Wiktorska, Zdzisław Chilmonczyk, Aleksander Mazurek, Adam Stasiulewicz
2018 International Journal of Molecular Sciences  
Here we present new derivatives of nucleoside reverse transcriptase inhibitors with a C20 fullerene. The computational chemistry methods used in this study evaluate affinity of designed compounds towards the HIV-1 reverse transcriptase (RT) binding site and select the most active ones. The best of the designed compounds have superior or similar affinity to RT active site in comparison to most active test compounds, including drugs used in anti-HIV therapy.
doi:10.3390/ijms19103231 fatcat:7gfut7rxgrf6zirrubtalukggm

Oncotoxic Properties of Serotonin Transporter Inhibitors and 5-HT1A Receptor Ligands

Jarosław Walory, Lidia Mielczarek, Małgorzata Jarończyk, Mirosława Koronkiewicz, Jerzy Kossakowski, Ryszard Bugno, Andrzej Bojarski, Zdzisław Chilmonczyk
2018 International Journal of Molecular Sciences  
The cytotoxic activity of several serotonin transporter (SERT) inhibitors and subtype of serotonin receptor 1A (5-HT1A receptor) ligands have been examined in androgen-insensitive human PC-3 prostate and neuroblastoma SH-SY5Y cancer cells. Almost all of the studied compounds (except 5-HT1A receptor agonist (2R)-(+)-8-Hydroxy-2-(di-n-propylamino)tetralin hydrobromide (8-OH-DPAT)) exhibited absolute cytotoxic activity against the examined cancer cells. The compound
more » ... phthalenyl)-1-piperazinyl]ethyl]benzamide hydrochloride (S14506) that showed highest activity against neuroblastoma tumors was the 5-HT1A receptor agonist (although not alike other 5-HT1A receptor agonists). On the other hand, the compound 6-nitro-2-(4-undecylpiperazin-1-yl)quinoline hydrochloride (AZ07) that had the highest activity against PC-3 prostate cancer cells was a compound exhibiting antagonistic activity against the 5-HT1A receptor. Thus, compounds of oncotoxic properties S14506 and AZ07 should be evaluated further for their potential use in the prevention and treatment of cancer. Most of the 15 compounds tested exhibited either agonistic or antagonistic activity for both the cyclic adenosine monophosphate (cAMP) and extracellular signal-regulated kinase 1 and 2 (ERK1/2) pathways in human embryonic kidney 293 (HEK293) cells that overexpress the 5HT1AR gene. However, compounds paroxetine, N-Ac-paroxetine and 2-[4-(cyclobutylmethyl)piperazin-1-yl]-6-nitroquinoline hydrochloride (AB22) simultaneously exhibited antagonistic activity on the cAMP pathway and agonistic activity on the ERK1/2 pathway. Fluoxetine relative to compound AZ07 had almost three times lower cytotoxic activity against PC-3 prostate cancer cells. However, the proapoptotic activity of fluoxetine compared to compound AZ07 is almost two times higher which would suggest that the cytotoxic activity of both compounds may be dependent on different cell death mechanisms. Compound S14506 was found to be an antagonist of the serine-threonine protein kinase B (Akt) pathway. Prosurvival Akt activity may be reversed by Akt antagonists. Therefore, the antagonistic activity of S14506 on the Akt pathway may evoke caspase-3 expression and cytotoxicity. It appears that one should not expect a straightforward relationship between the activation of particular serotonergic pathways by selective serotonin reuptake inhibitors (SSRIs) and 5-HT1A receptor ligands and their cytotoxic or cytoprotective activity. Additionally, nuclear transcription factor κB (NF-κB), which may be involved in 5-HT-dependent biochemical pathways by coordinating different subunits in the formation of a dimer, may regulate the transcription of different transduction pathways. Therefore, it can be suggested that the mechanism of the cytotoxic activity of certain compounds (serotonergic against nonserotonergic) may depend on the compound and cancer type being examined. Docking studies showed that S14506, buspirone and spiperone bind in similar ways in the 5-HT1A receptor model and interacted with similar 5-HT1A receptor residues. S14506 and spiperone were found to be located closer to both phenylalanines in TM6 than buspirone, thus exhibiting more antagonist binding modes.
doi:10.3390/ijms19103260 fatcat:ofonkkdesrhpbpuokxzec2vgmy

COMBINATIONS OF ISOTHIOCYANATES WITH DRUGS – A CHANCE OR THREAT TO CHEMOPREVENTION AND CANCER TREATMENT?

Lidia Mielczarek, Zdzisław Chilmonczyk, Piotr Suchocki, Piotr Wroczyński, Katarzyna Wiktorska
2018 Acta Poloniae Pharmaceutica - Drug Research  
Isothiocyanates (ITCs) are a group of compounds of natural origin which exhibit anticancer properties. In addition to the cytotoxic impact on cancer cells, confirmed in the multiple cell lines and the in vivo models, ITCs exhibit the cytoprotective effect in normal cells by regulating the activity of enzymes involved in xenobiotic metabolism. These properties of ITCs have led to a continuing increase in the number of studies which have shown that ITCs can sensitize cancer cells to cytostatic
more » ... gs used as standard in cancer therapies. On the other hand, these compounds may decrease the effectiveness of drugs by deregulating the metabolizing system of the cell. This paper discusses the results of a preclinical study on ITCs applications in combination therapy as well as their role in drug metabolism.
doi:10.32383/appdr/78769 fatcat:y2bczaezazbd3i52quyqpx4ule

4-[3-(4-Fluorobenzylideneamino)-2-oxo-1,3-oxazolidin-5-ylmethyl]morpholin-4-ium chloride monohydrate

Rafal Kruszynski, Tadeusz J. Bartczak, Zdzislaw Chilmonczyk, Jacek Cybulski
2001 Acta Crystallographica Section E  
Experimental The title compound was prepared according to the method of Chilmonczyk et al. (1997) . S3.  ...  In the Pharmaceutical Research Institute in Warsaw, a series of derivatives of 3-amino-2-oxazolidinone have been prepared (Chilmonczyk et al., 1997) .  ... 
doi:10.1107/s1600536801006742 fatcat:rl6f4yrfyrec7e7wrf77ylmw4u

Molecular mechanism of serotonin transporter inhibition elucidated by a new flexible docking protocol

Mari Gabrielsen, Rafał Kurczab, Aina W. Ravna, Irina Kufareva, Ruben Abagyan, Zdzisław Chilmonczyk, Andrzej J. Bojarski, Ingebrigt Sylte
2012 European Journal of Medicinal Chemistry  
Keywords: SERT homology models SERT inhibitors 4D flexible docking Structure-based pharmacophore models Virtual screening ICM a b s t r a c t The two main groups of antidepressant drugs, the tricyclic antidepressants (TCAs) and the selective serotonin reuptake inhibitors (SSRIs), as well as several other compounds, act by inhibiting the serotonin transporter (SERT). However, the binding mode and molecular mechanism of inhibition in SERT are not fully understood. In this study, five classes of
more » ... RT inhibitors were docked into an outward-facing SERT homology model using a new 4D ensemble docking protocol. Unlike other docking protocols, where protein flexibility is not considered or is highly dependent on the ligand structure, flexibility was here obtained by side chain sampling of the amino acids of the binding pocket using biased probability Monte Carlo (BPMC) prior to docking. This resulted in the generation of multiple binding pocket conformations that the ligands were docked into. The docking results showed that the inhibitors were stacked between the aromatic amino acids of the extracellular gate (Y176, F335) presumably preventing its closure. The inhibitors interacted with amino acids in both the putative substrate binding site and more extracellular regions of the protein. A general structureedocking-based pharmacophore model was generated to explain binding of all studied classes of SERT inhibitors. Docking of a test set of actives and decoys furthermore showed that the outwardfacing ensemble SERT homology model consistently and selectively scored the majority of active compounds above decoys, which indicates its usefulness in virtual screening.
doi:10.1016/j.ejmech.2011.09.056 pmid:22071255 pmcid:PMC3357065 fatcat:iueh3h7izrelli22zwrp4xigmy

Does molecular docking reveal alternative chemopreventive mechanism of activation of oxidoreductase by sulforaphane isothiocyanates?

Pawel Mazur, Tomasz Magdziarz, Andrzej Bak, Zdzislaw Chilmonczyk, Teresa Kasprzycka-Guttman, Irena Misiewicz-Krzemińska, Katarzyna Skupińska, Jaroslaw Polanski
2009 Journal of Molecular Modeling  
et al.. Does molecular docking reveal alternative chemopreventive mechanism of activation of oxidoreductase by sulforaphane isothiocyanates?. Abstract: Isothiocyanates (ITC) are well-known chemopreventive agents extracted from vegetables. This activity results from the activation of human oxidoreductase. In this letter, the uncompetitive activatory mechanism of ITC was investigated using docking and molecular dynamics simulations. This indicates that NAD(P)H:quinone oxidoreductase can
more » ... y improve enzyme-substrate recognition within the catalytic site if the ITC activator supports the interaction in the uncompetitive binding site. Response to Reviewers: Answers for reviews: Reviewer #2: The authors have explored uncompetitive activatory mechanism of isothiocyanates using docking and molecular dynamics simulations. Though the methodology adopted appears to be sound and discussion is convincing, how can they validate their hypothesis (either theoretical or experimental)? Considering the suggestions we performed additional analyses to validate the hypothesis. It seems that the movements of several amino acids in the USB are constrained by the presence of the activator resulting in some changes of the CBS. Further testing of this hypothesis has been performed using the reduced dynamic approach (RedMD), which is based on the coarse-grained model of the protein. Thus, two representations of NAD(P)H:quinine oxidoreductase have been prepared by changing each amino acid with artificial alpha carbon atoms having appropriate properties to fake replaced amino acids. The first representation mimics the original 1D4A crystal structure, whereas the second one modifies the properties of ASN45÷47 of chain B and D, respectively. Generally, it reflects the constrains caused by the presence of activators in the UBS. Moreover, the applied modifications resulted in the increase of masses and harmonic constant. Afterward, both representations have been used in Langevin's dynamics simulations repeated 10 times with different random seeds. The whole trajectory time was 20 ns with temperature of 300K. The atomic coordinates of the system were sampled every 20fs. Then, the entire set of the obtained frames has been superimposed by ASN45÷47 of chain B and D, respectively. Fig. 5 in the revised version represents the resulting histograms of RMSD of alpha carbon (CA) of TYR128 calculated against its reference position taken from the original 1D4A file for the whole trajectory. The red color indicates TYR128 CA of the 1D4A reference structure, while the blue one Abstract graphic Click here to download Abstract graphic: graphical.doc Abstract Isothiocyanates (ITC) are well-known chemopreventive agents extracted from vegetables. This activity results from the activation of human oxidoreductase. In this letter, the uncompetitive activatory mechanism of ITC was investigated using docking and molecular dynamics simulations. This indicates that NAD(P)H:quinone oxidoreductase can efficiently improve enzyme-substrate recognition within the catalytic site if the ITC activator supports the interaction in the uncompetitive binding site.
doi:10.1007/s00894-009-0628-5 pmid:20024690 fatcat:7n55v7zaerdx3ixam2tow47xjy

Identification of Novel Serotonin Transporter Compounds by Virtual Screening

Mari Gabrielsen, Rafał Kurczab, Agata Siwek, Małgorzata Wolak, Aina W. Ravna, Kurt Kristiansen, Irina Kufareva, Ruben Abagyan, Gabriel Nowak, Zdzisław Chilmonczyk, Ingebrigt Sylte, Andrzej J. Bojarski
2014 Journal of Chemical Information and Modeling  
The serotonin (5-hydroxytryptamine, 5-HT) transporter (SERT) plays an essential role in the termination of serotonergic neurotransmission by removing 5-HT from the synaptic cleft into the presynaptic neuron. It is also of pharmacological importance being targeted by antidepressants and psychostimulant drugs. Here, five commercial databases containing approximately 3.24 million drug-like compounds have been screened using a combination of two-dimensional (2D) fingerprint-based and
more » ... al (3D) pharmacophore-based screening and flexible docking into multiple conformations of the binding pocket detected in an outward-open SERT homology model. Following virtual screening (VS), selected compounds were evaluated using in vitro screening and full binding assays and an in silico hit-to-lead (H2L) screening was performed to obtain analogues of the identified compounds. Using this multistep VS/H2L approach, 74 active compounds, 46 of which had K(i) values of ≤1000 nM, belonging to 16 structural classes, have been identified, and multiple compounds share no structural resemblance with known SERT binders.
doi:10.1021/ci400742s pmid:24521202 pmcid:PMC3982395 fatcat:pbxb47lzsfbnzbgdhvlsticfzy

The Comparison of MTT and CVS Assays for the Assessment of Anticancer Agent Interactions

Lidia Śliwka, Katarzyna Wiktorska, Piotr Suchocki, Małgorzata Milczarek, Szymon Mielczarek, Katarzyna Lubelska, Tomasz Cierpiał, Piotr Łyżwa, Piotr Kiełbasiński, Anna Jaromin, Anna Flis, Zdzisław Chilmonczyk (+1 others)
2016 PLoS ONE  
Multiple in vitro tests are widely applied to assess the anticancer activity of new compounds, including their combinations and interactions with other drugs. The MTT (3-(4,5dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay is one of the most commonly used assays to assess the efficacy and interactions of anticancer agents. However, it can be significantly influenced by compounds that modify cell metabolism and reaction conditions. Therefore, several assays are sometimes used to
more » ... en for potential anticancer drugs. However, the majority of drug interactions are evaluated only with this single method. The aim of our studies was to verify whether the choice of an assay has an impact on determining the type of interaction and to identify the source of discrepancies. We compared the accuracy of MTT and CVS (crystal violet staining) assays in the interaction of two compounds characterized by similar anticancer activity: isothiocyanates (ITCs) and Selol. Confocal microscopy studies were carried out to assess the influence of these compounds on the reactive oxygen species (ROS) level, mitochondrial membrane potential, dead-to-live cell ratio and MTT-tetrazolium salt reduction rate. The MTT assay was less reliable than CVS. The MTT test of Selol and 2-oxoheptyl ITC, which affected the ROS level and MTT reduction rate, gave false negative (2-oxoheptyl ITC) or false positive (Selol) results. As a consequence, the MTT assay identified an antagonistic interaction between Selol and ITC, while the metabolism-independent CVS test identified an additive or synergistic interaction. In this paper, we show for the first time that the test assay may change the interpretation of the compound interaction. Therefore, the test method should be chosen with caution, considering the mechanism of action of the compound.
doi:10.1371/journal.pone.0155772 pmid:27196402 pmcid:PMC4873276 fatcat:ia74bfkc6bf3talhdqe776h3oq
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