Publications by authors named "Anna Makuch-Kocka"

11 Publications

  • Page 1 of 1

Effect of Chronic Administration of 5-(3-chlorophenyl)-4-Hexyl-2,4 -Dihydro-3-1,2,4-Triazole-3-Thione (TP-315)-A New Anticonvulsant Drug Candidate-On Living Organisms.

Int J Mol Sci 2021 Mar 25;22(7). Epub 2021 Mar 25.

Department of Pharmacology, Faculty of Health Sciences, Medical University of Lublin, 20-093 Lublin, Poland.

About 70 million people suffer from epilepsy-a chronic neurodegenerative disease. In most cases, the cause of the disease is unknown, but epilepsy can also develop as the result of a stroke, trauma to the brain, or the use of psychotropic substances. The treatment of epilepsy is mainly based on the administration of anticonvulsants, which the patient must most often use throughout their life. Despite significant progress in research on antiepileptic drugs, about 30% of patients still have drug-resistant epilepsy, which is insensitive to pharmacotherapy used so far. In our recent studies, we have shown that 4-alkyl-5-aryl-1,2,4-triazole-3-thiones act on the voltage-gated sodium channels and exhibit anticonvulsant activity in an MES (maximal electroshock-induced seizure) and 6Hz test in mice. Previous studies have shown their beneficial toxic and pharmacological profile, but their effect on a living organism during chronic use is still unknown. In the presented study, on the basis of the previously conducted tests and the PAMPA (parallel artificial membrane permeability assay) BBB (blood-brain barrier) test, we selected one 1,2,4-triazole-3-thione derivative-TP-315-for further studies aimed at assessing the impact of its chronic use on a living organism. After long-term administration of TP-315 to Albino Swiss mice, its effect on the functional parameters of internal organs was assessed by performing biochemical, morphological, and histopathological examinations. It was also determined whether the tested compound inhibits selected isoforms of the CYP450 enzyme system. On the basis of the conducted tests, it was found that TP-315 does not show nephrotoxic nor hepatotoxic effects and does not cause changes in hematological parameters. In vitro tests showed that TP-315 did not inhibit CYP2B6, CYP2D6, CYP3A4, or CYP3A5 enzymes at the concentration found in the serum of mice subjected to long-term exposure to this compound.
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http://dx.doi.org/10.3390/ijms22073358DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8037910PMC
March 2021

Determination of Cytotoxic Activity of Extracts against Human Melanoma Cells and Comparison of Their Cytotoxicity with Cytotoxicity of Some Anticancer Drugs.

Molecules 2021 Mar 20;26(6). Epub 2021 Mar 20.

Department of Environmental Chemistry and Bioanalytics, Faculty of Chemistry, Nicolaus Copernicus University, Gagarina 7, 87-100 Torun, Poland.

Melanoma is an enormous global health burden, and should be effectively addressed with better therapeutic strategies. Therefore, new therapeutic agents are needed for the management of this disease. The aim of this study was the investigation of cytotoxic activity of some isoquinoline alkaloid standards and extracts obtained from -collected before, during, and after flowering-against three different human melanoma cells (A375, G361, SK-MEL-3). The cytotoxicity of these extracts was not previously tested on these melanoma cell lines. Determination of alkaloid contents was performed by HPLC-DAD using Polar RP column and mobile phase containing acetonitrile, water, and 1-butyl-3-methylimidazolium tetrafluoroborate. The cytotoxicity of alkaloid standards was investigated by determination of cell viability and calculation of IC values. Significant differences were observed in the alkaloids content and cytotoxic activity of the extracts, depending on the season of collection of the plant material. In the extracts high contents of sanguinarine (from 4.8543 to 9.5899 mg/g of dry plant material) and chelerythrine (from 42.7224 to 6.8722 mg/g of dry plant material) were found. For both of these alkaloids, very high cytotoxic activity against the tested cell lines were observed. The IC values were in the range of 0.11-0.54 µg/mL for sanguinarine and 0.14 to 0.46 µg/mL for chelerythrine. IC values obtained for extracts against all tested cell lines were also very low (from 0.88 to 10.96 µg/mL). Cytotoxic activity of alkaloid standards and extracts were compared with the cytotoxicity of anticancer drugs-etoposide, cisplatin, and hydroxyurea. In all cases except the one obtained for cisplatin against A375, which was similar to that obtained after flowering against the same cell line, IC values obtained for anticancer drugs were higher than the IC values obtained for sanguinarine, chelerythrine, and extracts. Our results showed that extracts and isoquinoline alkaloids, especially sanguinarine and chelerythrine, could be recommended for further in vivo experiments in order to confirm the possibility of their application in the treatment of human melanomas.
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http://dx.doi.org/10.3390/molecules26061738DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8003779PMC
March 2021

The Family Genes Expression in Patients with Triple Negative Breast Cancer.

Int J Mol Sci 2021 Feb 12;22(4). Epub 2021 Feb 12.

Chair and Department of Biology and Genetics, Medical University of Lublin, 4a Chodźki St., 20-093 Lublin, Poland.

The (baculoviral IAP repeat-containing; BIRC) family genes encode for Inhibitor of Apoptosis (IAP) proteins. The dysregulation of the expression levels of the genes in question in cancer tissue as compared to normal tissue suggests that the apoptosis process in cancer cells was disturbed, which may be associated with the development and chemoresistance of triple negative breast cancer (TNBC). In our study, we determined the expression level of eight genes from the BIRC family using the Real-Time PCR method in patients with TNBC and compared the obtained results with clinical data. Additionally, using bioinformatics tools (Ualcan and The Breast Cancer Gene-Expression Miner v4.5 (bc-GenExMiner v4.5)), we compared our data with the data in the Cancer Genome Atlas (TCGA) database. We observed diverse expression pattern among the studied genes in breast cancer tissue. Comparing the expression level of the studied genes with the clinical data, we found that in patients diagnosed with breast cancer under the age of 50, the expression levels of all studied genes were higher compared to patients diagnosed after the age of 50. We observed that in patients with invasion of neoplastic cells into lymphatic vessels and fat tissue, the expression levels of family genes were lower compared to patients in whom these features were not noted. Statistically significant differences in gene expression were also noted in patients classified into three groups depending on the basis of the Scarff-Bloom and Richardson (SBR) Grading System.
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http://dx.doi.org/10.3390/ijms22041820DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7918547PMC
February 2021

The Correlation of Mutations and Expressions of Genes within the PI3K/Akt/mTOR Pathway in Breast Cancer-A Preliminary Study.

Int J Mol Sci 2021 Feb 19;22(4). Epub 2021 Feb 19.

Chair and Department of Biology and Genetics, Medical University of Lublin, 20-093 Lublin, Poland.

There is an urgent need to seek new molecular biomarkers helpful in diagnosing and treating breast cancer. In this elaboration, we performed a molecular analysis of mutations and expression of genes within the PI3K/Akt/mTOR pathway in patients with ductal breast cancer of various malignancy levels. We recognized significant correlations between the expression levels of the studied genes. We also performed a bioinformatics analysis of the data available on the international database TCGA and compared them with our own research. Studies on mutations and expression of genes were conducted using High-Resolution Melt PCR (HRM-PCR), Allele-Specific-quantitative PCR (ASP-qPCR), Real-Time PCR molecular methods in a group of women with ductal breast cancer. Bioinformatics analysis was carried out using web source Ualcan and bc-GenExMiner. In the studied group of women, it was observed that the prevalence of mutations in the studied and genes was 29.63%. It was stated that the average expression level of the , , genes in the group of breast cancer patients is lower in comparison to the control group, while the average expression level of the and genes in the studied group was higher in comparison to the control group. It was also indicated that in the group of patients with mutations in the area of the and genes, the gene expression level is statistically significantly lower than in the group without mutations. According to our knowledge, we demonstrate, for the first time, that there is a very strong positive correlation between the levels of and gene expression in the case of patients with mutations and without mutations.
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http://dx.doi.org/10.3390/ijms22042061DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7922286PMC
February 2021

Determination of Cytotoxic Activity of Selected Isoquinoline Alkaloids and Plant Extracts Obtained from Various Parts of Collected in Various Vegetation Seasons.

Molecules 2021 Feb 4;26(4). Epub 2021 Feb 4.

Department of Environmental Chemistry and Bioanalytics, Faculty of Chemistry, Nicolaus Copernicus University, Gagarina 7, PL-87-100 Torun, Poland.

Melanoma is a serious form of skin cancer that begins in cells known as melanocytes. While it is less common than the other forms of skin cancer, melanoma is more dangerous because of its ability to spread to other organs more rapidly if it is not treated at an early stage. The number of people diagnosed with melanoma has increased over the last few decades. The most widely used treatments include surgery, chemotherapy, and radiation therapy. The search for new drugs to treat various cancers is one of the most important challenges of modern scientific research. Some isoquinoline alkaloids found in different plant species have strong cytotoxic effects on various cancer cells. We tested the effect of isoquinoline alkaloids and extracts obtained from various parts of collected in various vegetation seasons on human melanoma cancer cells and our data indicated that investigated extract induced significant reduction in cell viability of Human malignant melanoma cells (A375), human Caucasian malignant melanoma cell line (G361), and human malignant melanoma cell line (SKMEL3 cancer cell lines in a dose- and time-dependent manner. Differences in cytotoxic activity were observed for extracts obtained from various parts of . Significant differences were also obtained in the alkaloids content and cytotoxic activity of the extracts depending on the season of collection of plant material. Our investigations exhibit that these plant extracts can be recommended for further in vivo experiments in order to confirm the possibility of their use in the treatment of human melanomas.
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http://dx.doi.org/10.3390/molecules26040816DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7915140PMC
February 2021

Effect of Tryptophan-Derived AhR Ligands, Kynurenine, Kynurenic Acid and FICZ, on Proliferation, Cell Cycle Regulation and Cell Death of Melanoma Cells-In Vitro Studies.

Int J Mol Sci 2020 Oct 26;21(21). Epub 2020 Oct 26.

Department of Pharmacology, Medical University of Lublin, Chodźki 4a, 20-093 Lublin, Poland.

Tryptophan metabolites: kynurenine (KYN), kynurenic acid (KYNA) and 6-formylindolo[3,2-b]carbazole (FICZ) are considered aryl hydrocarbon receptor (AhR) ligands. AhR is mainly expressed in barrier tissues, including skin, and is involved in various physiological and pathological processes in skin. We studied the effect of KYN, KYNA and FICZ on melanocyte and melanoma A375 and RPMI7951 cell toxicity, proliferation and cell death. KYN and FICZ inhibited DNA synthesis in both melanoma cell lines, but RPMI7951 cells were more resistant to pharmacological treatment. Tested compounds were toxic to melanoma cells but not to normal human adult melanocytes. Changes in the protein level of cyclin D1, CDK4 and retinoblastoma tumor suppressor protein (Rb) phosphorylation revealed different mechanisms of action of individual AhR ligands. Importantly, all tryptophan metabolites induced necrosis, but only KYNA and FICZ promoted apoptosis in melanoma A375 cells. This effect was not observed in RPMI7951 cells. KYN, KYNA and FICZ in higher concentrations inhibited the protein level of AhR but did not affect the gene expression. To conclude, despite belonging to the group of AhR ligands, KYN, KYNA and FICZ exerted different effects on proliferation, toxicity and induction of cell death in melanoma cells in vitro.
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http://dx.doi.org/10.3390/ijms21217946DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7663343PMC
October 2020

A Tryptophan Metabolite, 8-Hydroxyquinaldic Acid, Exerts Antiproliferative and Anti-Migratory Effects on Colorectal Cancer Cells.

Molecules 2020 Apr 3;25(7). Epub 2020 Apr 3.

Department of Pharmacology, Medical University of Lublin, Chodźki 4a, 20-093 Lublin, Poland.

8-Hydroxyquinaldic acid, the end-metabolite of tryptophan, is well-known metal chelator; however, its role in humans, especially in cancer promotion and progression, has not been fully revealed. Importantly, 8-hydroxyquinaldic acid is the analog of kynurenic acid with evidenced antiproliferative activity towards various cancer cells. In this study, we revealed that 8-hydroxyquinaldic acid inhibited not only proliferation and mitochondrial activity in colon cancer HT-29 and LS-180 cells, but it also decreased DNA synthesis up to 90.9% for HT-29 cells and 76.1% for LS-180 cells. 8-Hydroxyquinaldic acid induced changes in protein expression of cell cycle regulators (CDK4, CDK6, cyclin D1, cyclin E) and CDKs inhibitors (p21 Waf1/Cip1, p27 Kip1), but the effect was dependent on the tested cell line. Moreover, 8-hydroxyquinaldic acid inhibited migration of colon cancer HT-29 and LS-180 cells and increased the expression of β-catenin and E-cadherin. Importantly, antiproliferative and anti-migratory concentrations of 8-hydroxyquinaldic acid were non-toxic in vitro and in vivo. We reported for the first time antiproliferative and anti-migratory activity of 8-hydroxyquinaldic acid against colon cancer HT-29 and LS-180 cells.
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http://dx.doi.org/10.3390/molecules25071655DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7181169PMC
April 2020

Cytotoxic activity of methanolic fractions of different Marrubium spp. against melanoma cells is independent of antioxidant activity and total phenolic content.

FEBS Open Bio 2020 01 2;10(1):86-95. Epub 2019 Dec 2.

Department of Toxicology, Medical University of Lublin, Poland.

The Marrubium genus (horehound) has proved to be an abundant source of biologically active compounds, but there is little knowledge about its potential anticancer activity. Moreover, some Marrubium species have not been the subject of study in this regard. In this study, we performed comparative analysis of phenolic acid (PhA) content and total phenolic content in fractions obtained from methanolic extracts of Marrubium vulgare L. (common horehound), Marrubium cylleneum Boiss. & Heldr. and Marrubium friwaldskyanum Boiss herbs. We examined the cytotoxicity of these fractions against a human melanoma cancer cell line (A375) and normal human skin fibroblasts (BJ) using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide test, cell cycle analysis and real-time monitoring of cell viability. We detected caffeic, p-coumaric, ferulic and gentisic acids among the PhAs. Although the extracts obtained demonstrated low total phenolic content and did not show significant antioxidative properties, the nonhydrolyzed PhA fraction exhibited cytotoxic activity against a human melanoma cancer cell line, without affecting normal fibroblasts. Both acidic and alkaline hydrolysis abolished this activity, indicating that the esterified forms of phenolic compounds caused the observed cytotoxic effects. Further investigation of these compounds may facilitate the development of novel drugs for cancer treatment.
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http://dx.doi.org/10.1002/2211-5463.12755DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943230PMC
January 2020

Imaging Flow Cytometric Analysis of Stilbene-Dependent Apoptosis in Drug Resistant Human Leukemic Cell Lines.

Molecules 2019 May 17;24(10). Epub 2019 May 17.

Clinical Genetics Department, Medical University of Lublin, 11 Radziwillowska Str., 20-080 Lublin, Poland.

The natural compounds have been researched extensively as an alternative to the conventional chemotherapy and radiation. Stilbene derivatives appear as a group of therapeutics which deserves special attention. The present study was designed to analyze the effects of stilbene derivatives on drug resistant human leukemic cells. The aim of this work was to evaluate the apoptotic effect of stilbene derivatives in various concentrations on leukemic cells (LC) with and without resistant phenotype. Human acute promyelocytic leukemia (APL) cell lines (HL60, HL60/MX1, HL60/MX2) and acute lymphoblastic leukemia (ALL) cell lines (CEM/C1, CCRF-CEM) were studied. T-resveratrol, piceatannol, rhaponticin, deoxyrhaponticin, pterostilbene were used to stimulate apoptosis. Mitoxantrone (MIT) was applied to induce drug resistance. -Resveratrol (RES), deoxyrhaponticin (D-RHAP), rhaponticin (RHAP), pterostilbene (PTER), and piceatannol (PIC) influenced viability and induced apoptosis in all investigated cell lines. Our results confirmed that RES, PIC, RHAP, D-RHAP, and PTER are essential therapeutic compounds with anticancer activity exhibited by induction of apoptosis in leukemic cells with and without resistant phenotype. Stilbene-induced apoptosis in HL60/MX1, HL60/MX2, CEM/C1, and CCRF-CEM leukemia cell lines have been presented in very few studies so far and our research is an important contribution to the investigation of these substances.
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http://dx.doi.org/10.3390/molecules24101896DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6571880PMC
May 2019

The Effect of Furanocoumarin Derivatives on Induction of Apoptosis and Multidrug Resistance in Human Leukemic Cells.

Molecules 2019 May 12;24(9). Epub 2019 May 12.

Chair and Department of Biology and Genetics, Faculty of Pharmacy with Medical Analytics Division, Medical University of Lublin, 20-059 Lublin, Poland.

Background: The insensitivity of cancer cells to therapeutic agents is considered to be the main cause of failure of therapy and mortality of patients with cancer. A particularly important problem in these patients is the phenomenon of multidrug resistance, consisting of abnormal, elevated expression of transport proteins (ABC family). The aim of this research included determination of IC50 values of selected furanocoumarins in the presence and absence of mitoxantrone in leukemia cells and analysis of changes in apoptosis using anexinV/IP and Casp3/IP after 24 h exposure of cell lines to selected coumarins in the presence and absence of mitoxantrone in IC50 concentrations.

Methods: Research was conducted on 3 cell lines derived from the human hematopoietic system: HL-60, HL-60/MX1 and HL-60/MX2. After exposure to coumarin compounds, cells were subjected to cytometric analysis to determine the induction of apoptosis by two methods: the Annexin V test with propidium iodide and the PhiPhiLux-G1D2 reagent containing caspase 3 antibodies.

Results: All of the furanocoumarin derivatives studied were found to induce apoptosis in leukemia cell lines.

Conclusions: Our results clearly show that the furanocoumarin derivatives are therapeutic substances with antitumor activity inducing apoptosis in human leukemia cells with phenotypes of resistance.
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http://dx.doi.org/10.3390/molecules24091824DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6539621PMC
May 2019

Development of the 1,2,4-triazole-based anticonvulsant drug candidates acting on the voltage-gated sodium channels. Insights from in-vivo, in-vitro, and in-silico studies.

Eur J Pharm Sci 2019 Mar 28;129:42-57. Epub 2018 Dec 28.

Department of Pharmacology, Medical University of Lublin, Lublin, Poland. Electronic address:

The treatment of epilepsy remains difficult mostly since almost 30% of patients suffer from pharmacoresistant forms of the disease. Therefore, there is an urgent need to search for new antiepileptic drug candidates. Previously, it has been shown that 4-alkyl-5-substituted-1,2,4-triazole-3-thione derivativatives possessed strong anticonvulsant activity in a maximal electroshock-induced seizure model of epilepsy. In this work, we examined the effect of the chemical structure of the 1,2,4-triazole-3-thione-based molecules on the anticonvulsant activity and the binding to voltage-gated sodium channels (VGSCs) and GABA receptors. Docking simulations allowed us to determine the mode of interactions between the investigated compounds and binding cavity of the human VGSC. Selected compounds were also investigated in a panel of ADME-Tox assays, including parallel artificial membrane permeability assay (PAMPA), single cell gel electrophoresis (SCGE) and cytotoxicity evaluation in HepG2 cells. The obtained results indicated that unbranched alkyl chains, from butyl to hexyl, attached to 1,2,4-triazole core are essential both for good anticonvulsant activity and strong interactions with VGSCs. The combined in-vivo, in-vitro and in-silico studies emphasize 4-alkyl-5-substituted-1,2,4-triazole-3-thiones as promising agents in the development of new anticonvulsants.
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http://dx.doi.org/10.1016/j.ejps.2018.12.018DOI Listing
March 2019
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