Analysis of Selected Salivary Adipokines and Cytokines in Patients with Obesity-A Pilot Study.

Obesity is a chronic, progressive and relapsing disease that produces many adverse health, social and economic effects. The aim of the study was to analyse the concentrations of selected proinflammatory parameters in the saliva of obese and normal body weight individuals. The study included 116 people divided into two groups: the study group (n = 75, subjects with obesity) and the control group (n = 41, individuals with normal body weight). Bioelectrical impedance analysis was performed, and saliva samples were collected from all study participants to determine the concentrations of selected proinflammatory adipokines and cytokines. Statistically significantly higher concentrations of MMP-2, MMP-9 and IL-1ß were found in the saliva of obese women compared to women with normal body weight. Furthermore, statistically significantly higher concentrations of MMP-9, IL-6 and resistin were observed in the saliva of obese men compared to men with normal body weight. Higher concentrations of selected proinflammatory cytokines and adipokines were found in the saliva of obese individuals compared to individuals with normal body weight. It is likely that higher concentrations of MMP-2, MMP-9 and IL-1ß can be detected in the saliva of obese women compared to non-obese women, while higher concentrations of MMP-9, IL-6 and resistin can be found in the saliva of obese men compared to non-obese men, which suggests that further research to confirm our observations and determine the mechanisms of development of metabolic complications associated with obesity depending on gender is needed.

Pro-Inflammatory Adipokine and Cytokine Profiles in the Saliva of Obese Patients with Non-Alcoholic Fatty Liver Disease (NAFLD)-A Pilot Study.

Undiagnosed and untreated non-alcoholic fatty liver disease (NAFLD) can lead to the development of many complications, such as cirrhosis, hepatocellular carcinoma, or cardiovascular diseases. Obese people are at increased risk of developing NAFLD. Due to the current lack of routine diagnostics, it is extremely important to look for new diagnostic methods and markers for this disease. The aim of this study was to assess the concentration of selected pro-inflammatory adipokines and cytokines in the unstimulated saliva of obese people with fatty liver disease in various stages (with or without slight fibrosis) and to analyze them for possible use as early markers of NAFLD diagnosis. The study involved 96 people who were divided into 5 groups based on the criterion of body mass index (BMI) and the degree of fatty liver (liver elastography). There were statistically significant differences between the groups in the concentrations of MMP-9 (matrix metalloproteinase 9), resistin, and IL-1ß (interleukin 1ß) in saliva. Statistically significant, positive correlations between hepatic steatosis and the concentration of MMP-2 (matrix metalloproteinase 2), resistin, and IL-1ß in saliva were also found. Statistically significant positive correlations were also found between the concentration of resistin in saliva and the concentration of ALT (alanine aminotransferase) and GGTP (gamma-glutamyl transpeptidase) in serum. MMP-2, IL-1ß, and resistin may be potential markers of NAFLD development, assessed in saliva. However, further research is needed because this is the first study to evaluate the concentrations of the selected pro-inflammatory parameters in the saliva of patients with NAFLD.

Synthesis, Biological Activity, ADME and Molecular Docking Studies of Novel Ursolic Acid Derivatives as Potent Anticancer Agents.

A series of new ursolic acid (UA) derivatives substituted with various amino acids (AAs) or dipeptides (DP) at the C-3 position of the steroid skeleton was designed and synthesized. The compounds were obtained by the esterification of UA with the corresponding AAs. The cytotoxic activity of the synthesized conjugates was determined using the hormone-dependent breast cancer cell line MCF-7 and the triple-negative breast cancer cell line MDA. Three derivatives (l-seryloxy-, l-prolyloxy- and l-alanyl-l-isoleucyloxy-) showed micromolar IC50 values and reduced the concentrations of matrix metalloproteinases 2 and 9. Further studies revealed that for two compounds (l-seryloxy- and l-alanyl-l-isoleucyloxy-), a possible mechanism of their antiproliferative action is the activation of caspase-7 and the proapoptotic Bax protein in the apoptotic pathway. The third compound (l-prolyloxy- derivative) showed a different mechanism of action as it induced autophagy as measured by an increase in the concentrations of three autophagy markers: LC3A, LC3B, and beclin-1. This derivative also showed statistically significant inhibition of the proinflammatory cytokines TNF-α and IL-6. Finally, for all synthesized compounds, we computationally predicted their ADME properties as well as performed molecular docking to the estrogen receptor to assess their potential for further development as anticancer agents.

Multi-Targeting Anticancer Activity of a New 4-Thiazolidinone Derivative with Anti-HER2 Antibodies in Human AGS Gastric Cancer Cells.

Combining chemotherapy with immunotherapy still remains a regimen in anticancer therapy. Novel 4-thiazolidinone-bearing hybrid molecules possess well-documented anticancer activity, and together with anti-HER2 antibodies, may represent a promising strategy in treating patients with gastric cancer with confirmed human epidermal growth factor receptor 2 (HER2) expression. The aim of the study was to synthesize a new 4-thiazolidinone derivative (Les-4367) and investigate its molecular mechanism of action in combination with trastuzumab or pertuzumab in human AGS gastric cancer cells. AGS cell viability and antiproliferative potential were examined. The effect of the tested combinations as well as monotherapy on apoptosis and autophagy was also determined. Metalloproteinase-2 (MMP-2), intercellular adhesion molecule 1 (ICAM-1), pro-inflammatory and anti-inflammatory cytokine concentrations were also demonstrated by the ELISA technique. We proved that pertuzumab and trastuzumab were very effective in increasing the sensitivity of AGS gastric cancer cells to novel Les-4367. The molecular mechanism of action of the tested combination is connected with the induction of apoptosis. Additionally, the anticancer activity is not associated with the autophagy process. Decreased concentrations of pro-inflammatory cytokines, MMP-2 and ICAM-1-were observed. The novel combination of drugs based on anti-HER2 antibodies with Les-4367 is a promising strategy against AGS gastric cancer cells.

LC-PDA-MS and GC-MS Analysis of Scorzonera hispanica Seeds and Their Effects on Human Breast Cancer Cell Lines.

Scorzonera hispanica is an herbaceous perennial cultivated in Central and Southern Europe. This study aimed to qualitatively and quantitatively evaluate the composition of oil, extracts, and fractions (SH1-SH12) obtained from S. hispanica seeds. Furthermore, an evaluation of biological activities in breast cancer cell lines was also performed. GC-MS analysis revealed that the primary components of the seed oil (SH12) were fatty acids and ß-sitosterol. In the evaluation of extracts (SH1-SH3, SH8-SH10) and fractions (SH4-SH7, SH11) composition, the presence of apigenin, derivatives of p-coumaric and caffeic acids, was reported. In the biological assays, methanolic extract (SH1), diethyl ether (SH4), and chloroform (SH11) fractions exhibited cytotoxicity toward cells. The highest activity was observed for fatty acids- and 3,4-dimethoxycinnamate-rich SH11 (IC50: 399.18 µg/mL for MCF-7, 781.26 µg/mL for MDA-MB-231). SH11 was also observed to induce apoptosis in MCF-7 cells (52.4%). SH1, SH4, and SH11 attenuate signaling pathways and affect the expression of apoptosis-, autophagy-, and inflammation-related proteins. SH12 was non-toxic toward either cancer or normal cell lines in concentrations up to 1 mg/mL. The results suggest that S. hispanica seeds exhibit a wide range of potential uses as a source of oil and bioactive compounds for complementary therapy of breast cancer.

2-{5-[(Z,2Z)-2-Chloro-3-(4-nitrophenyl)-2-propenylidene]-4-oxo-2-thioxothiazolidin-3-yl}-3-methylbutanoic Acid as a Potential Anti-Breast Cancer Molecule.

It was established that the synthesis of hybrid molecules containing a thiazolidinone and a (2Z)-2-chloro-3-(4-nitrophenyl)prop-2-ene structural fragments is an effective approach for the design of potential anticancer agents. Given the results of the previous SAR-analysis, the aim of the study was to synthesize a novel 4-thiazolidinone derivative Les-3331 and investigate its molecular mechanism of action in MCF-7 and MDA-MB-231 breast cancer cells. The cytotoxic properties and antiproliferative potential of Les-3331 were determined. The effect of the tested compound on apoptosis induction and mitochondrial membrane potential was checked by flow cytometry. ELISA was used to determine caspase-8 and caspase-9, LC3A, LC3B, Beclin-1, and topoisomerase II concentration. Additionally, PAMPA, in silico or in vitro prediction of metabolism, CYP3A4/2D6 inhibition, and an Ames test were performed. Les-3331 possesses high cytotoxic and antiproliferative activity in MCF-7 and MDA-MB-231 breast cancer cells. Its molecular mechanism of action is associated with apoptosis induction, decreased mitochondrial membrane potential, and increased caspase-9 and caspase-8 concentrations. Les-3331 decreased LC3A, LC3B, and Beclin-1 concentration in tested cell lines. Topoisomerase II concentration was also lowered. The most probable metabolic pathways and no DDIs risk of Les-3331 were confirmed in in vitro assays. Our studies confirmed that a novel 4-thiazolidinone derivative represents promising anti-breast cancer activity.

Synthesis and Anticancer Activity of 1,3,4-Thiadiazoles with 3-Methoxyphenyl Substituent.

Based on the results of previous work, we designed and synthesized 1,3,4-thiadiazole derivatives. The cytotoxic activity of the obtained compounds was then determined in biological studies using MCF-7 and MDA-MB-231 breast cancer cells and a normal cell line (fibroblasts). The results showed that all compounds displayed weak anticancer activity towards two breast cancer lines: an estrogen-dependent cell line (MCF-7) and an estrogen-independent cell line (MDA-MB-231). The compound most active towards MCF-7 breast cancer cells was SCT-4, which decreased DNA biosynthesis to 70% ± 3 at 100 µM. The mechanism of the anticancer action of 1,3,4-thiadiazole was also investigated. We choose a set of the most investigated proteins, which are attractive anticancer targets. In silico studies demonstrated a possible multitarget mode of action for the synthesized compounds but the most likely mechanism of action for the new compounds is connected with the activity of caspase 8.

New 1,3,4-Thiadiazole Derivatives with Anticancer Activity.

We designed and synthesized the 1,3,4-thiadiazole derivatives differing in the structure of the substituents in C2 and C5 positions. The cytotoxic activity of the obtained compounds was then determined in biological studies using MCF-7 and MDA-MB-231 breast cancer cells and normal cell line (fibroblasts). The results showed that in both breast cancer cell lines, the strongest anti-proliferative activity was exerted by 2-(2-trifluorometylophenylamino)-5-(3-methoxyphenyl)-1,3,4-thiadiazole. The IC50 values of this compound against MCF-7 and MDA-MB-231 breast cancer cells were 49.6 µM and 53.4 µM, respectively. Importantly, all new compounds had weaker cytotoxic activity on normal cell line than on breast cancer cell lines. In silico studies demonstrated a possible multitarget mode of action for the synthesized compounds. The most likely mechanism of action for the new compounds is connected with the activities of Caspase 3 and Caspase 8 and activation of BAX proteins.

Phytochemical Composition and Biological Activities of Scorzonera Species.

The genus Scorzonera comprises nearly 200 species, naturally occurring in Europe, Asia, and northern parts of Africa. Plants belonging to the Scorzonera genus have been a significant part of folk medicine in Asia, especially China, Mongolia, and Turkey for centuries. Therefore, they have become the subject of research regarding their phytochemical composition and biological activity. The aim of this review is to present and assess the phytochemical composition, and bioactive potential of species within the genus Scorzonera. Studies have shown the presence of many bioactive compounds like triterpenoids, sesquiterpenoids, flavonoids, or caffeic acid and quinic acid derivatives in extracts obtained from aerial and subaerial parts of the plants. The antioxidant and cytotoxic properties have been evaluated, together with the mechanism of anti-inflammatory, analgesic, and hepatoprotective activity. Scorzonera species have also been investigated for their activity against several bacteria and fungi strains. Despite mild cytotoxicity against cancer cell lines in vitro, the bioactive properties in wound healing therapy and the treatment of microbial infections might, in perspective, be the starting point for the research on Scorzonera species as active agents in medical products designed for miscellaneous skin conditions.

Autophagy Modulators in Cancer Therapy.

Autophagy is a process of self-degradation that plays an important role in removing damaged proteins, organelles or cellular fragments from the cell. Under stressful conditions such as hypoxia, nutrient deficiency or chemotherapy, this process can also become the strategy for cell survival. Autophagy can be nonselective or selective in removing specific organelles, ribosomes, and protein aggregates, although the complete mechanisms that regulate aspects of selective autophagy are not fully understood. This review summarizes the most recent research into understanding the different types and mechanisms of autophagy. The relationship between apoptosis and autophagy on the level of molecular regulation of the expression of selected proteins such as p53, Bcl-2/Beclin 1, p62, Atg proteins, and caspases was discussed. Intensive studies have revealed a whole range of novel compounds with an anticancer activity that inhibit or activate regulatory pathways involved in autophagy. We focused on the presentation of compounds strongly affecting the autophagy process, with particular emphasis on those that are undergoing clinical and preclinical cancer research. Moreover, the target points, adverse effects and therapeutic schemes of autophagy inhibitors and activators are presented.

Mucin 1 as a Molecular Target of a Novel Diisoquinoline Derivative Combined with Anti-MUC1 Antibody in AGS Gastric Cancer Cells.

BACKGROUND: The aim of the study was to examine the molecular mechanism of the anticancer action of a monoclonal antibody against MUC1 and a diisoquinoline derivative (OM-86II) in human gastric cancer cells. METHODS: The cell viability was measured by the MTT assay. The disruption of mitochondrial membrane potential and activity of caspase-8 and caspase-9 was performed by flow cytometry. Fluorescent microscopy was used to confirm the proapoptotic effect of compounds. LC3A, LC3B and Beclin-1 concentrations were analyzed to check the influence of the compounds on induction of autophagy. ELISA assessments were performed to measure the concentration of mTOR, sICAM1, MMP-2, MMP-9 and pro-apoptotic Bax. RESULTS: The anti-MUC1 antibody with the diisoquinoline derivative (OM-86II) significantly reduced gastric cancer cells' viability. This was accompanied by an increase in caspase-8 and caspase-9 activity as well as high concentrations of pro-apoptotic Bax. We also proved that the anti-MUC1 antibody with OM-86II decreased the concentrations of MMP-9, sICAM1 and mTOR in gastric cancer cells. After 48 h of incubation with such a combination, we observed higher levels of the crucial component of autophagosomes (LC3) and Beclin-1. CONCLUSIONS: Our study proved that the anti-MUC1 antibody sensitizes human gastric cancer cells to the novel diisoquinoline derivative (OM-86II) via induction of apoptosis and autophagy, and inhibition of selected proteins such as mTOR, sICAM1 and MMP-9.

The Anticancer Action of a Novel 1,2,4-Triazine Sulfonamide Derivative in Colon Cancer Cells.

Cancer therapy is one of the most important challenges of modern medical and chemical sciences. Among the many methods of combating cancer, chemotherapy plays a special role. Imperfect modern chemotherapy justifies continuing the search for new, more effective, and safe drugs. Sulfonamides are the classic group of chemotherapeutic drugs with a broad spectrum of pharmacological activity. Recent literature reports show that sulfonamide derivatives have anti-tumor activity in vitro and in vivo. The aim of the study was to synthesize a novel 1,2,4-triazine sulfonamide derivative and check its anticancer potential in DLD-1 and HT-29 colon cancer cells. The biological studies included MTT assay, DNA biosynthesis, cell cycle analysis, Annexin V binding assay, ethidium bromide/acridine orange staining, and caspase-8, -9, and -3/7 activity. The concentrations of important molecules (sICAM-1, mTOR, Beclin-1, cathepsin B) involved in the pathogenesis and poor prognosis of colorectal cancer were also evaluated by ELISA. We demonstrated that the novel compound was able to induce apoptosis through intrinsic and extrinsic pathways and was capable of decreasing sICAM-1, mTOR, cathepsin B concentrations, whereas increased Beclin-1 concentration was detected in both colon cancer cell lines. The novel compound represents promising multi-targeted potential in colorectal cancer, but further in vivo examinations are needed to confirm the claim.

DNA topoisomerases as molecular targets for anticancer drugs.

The significant role of topoisomerases in the control of DNA chain topology has been confirmed in numerous research conducted worldwide. The prevalence of these enzymes, as well as the key importance of topoisomerase in the proper functioning of cells, have made them the target of many scientific studies conducted all over the world. This article is a comprehensive review of knowledge about topoisomerases and their inhibitors collected over the years. Studies on the structure-activity relationship and molecular docking are one of the key elements driving drug development. In addition to information on molecular targets, this article contains details on the structure-activity relationship of described classes of compounds. Moreover, the work also includes details about the structure of the compounds that drive the mode of action of topoisomerase inhibitors. Finally, selected topoisomerases inhibitors at the stage of clinical trials and their potential application in the chemotherapy of various cancers are described.

The Effect of Novel 7-methyl-5-phenyl-pyrazolo[4,3-e]tetrazolo[4,5-b][1,2,4]triazine Sulfonamide Derivatives on Apoptosis and Autophagy in DLD-1 and HT-29 Colon Cancer Cells.

The discovery of cytotoxic drugs is focused on designing a compound structure that directly affects cancer cells without an impact on normal cells. The mechanism of anticancer activity is mainly related with activation of apoptosis. However, recent scientific reports show that autophagy also plays a crucial role in cancer cell progression. Thus, the objective of this study was to synthesize 7-methyl-5-phenyl-pyrazolo[4,3-e]tetrazolo[4,5-b][1,2,4]triazine utilizing nucleophilic substitution reaction at the position N1. The biological activity of tested compounds was assessed in DLD-1 and HT-29 cell lines. The induction of apoptosis was confirmed by Annexin V binding assay and acridine orange/ethidium bromide staining. The loss of mitochondrial membrane potential and caspase-8 activity was estimated using cytometer flow analysis. The concentration of p53, LC3A, LC3B and beclin-1 was measured using the ELISA technique. Our study revealed that anticancer activity of 7-methyl-5-phenyl-pyrazolo[4,3-e]tetrazolo[4,5-b][1,2,4]triazine derivatives is related with initiation of apoptosis occur on the intrinsic pathway with mitochondrial membrane decrease and extrinsic with increase of activity of caspase-8. Moreover, a decrease in beclin-1, LC3A, and LC3B were observed in two cell lines after treatment with novel compounds. This study showed that novel 7-methyl-5-phenyl-pyrazolo[4,3-e]tetrazolo[4,5-b][1,2,4]triazine derivatives might be a potential strategy in colon cancer treatment.

The molecular mechanism of anticancer action of novel octahydropyrazino[2,1-a:5,4-a']diisoquinoline derivatives in human gastric cancer cells.

Objective The aim of the current study was to examine the anticancer activity and the detailed mechanism of novel diisoquinoline derivatives in human gastric cancer cells (AGS). Methods The viability of AGS cells was measured by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Cell cycle analysis and apoptosis assay were performed by standard flow cytometric method. Confocal microscopy bioimaging was used to demonstrate the expression of pivotal proteins engaged in apoptosis (caspase-8, caspase-3, p53) and cell signaling (AKT, ERK1/2). Results All compounds decreased the number of viable cells in a dose-dependent manner after 24 and 48 h of incubation, although compound 2 was a more cytotoxic agent, with IC50 values of 21 ± 2 and 6 ± 2 µM, compared to 80 ± 2 and 45 ± 2 µM for etoposide. The cytotoxic and antiproliferative effects of novel compounds were associated with the induction of apoptosis. The highest percentage of early and late apoptotic cells was observed after 48 h of incubation with compound 2 (89.9%). The value was higher compared to compound 1 (20.4%) and etoposide (24.1%). The novel diisoquinoline derivatives decreased the expression of AKT and ERK1/2. Their mechanism was associated with p53-mediated apoptosis, accumulation of cells in the G2/M phase of cell cycle and inhibition of topoisomerase II. Conclusion These data strongly support compound 2 as a promising molecule for treatment of gastric cancer.

A novel series of pyrazole-platinum(II) complexes as potential anti-cancer agents that induce cell cycle arrest and apoptosis in breast cancer cells.

Six novel compounds of platinum(II) with pyrazole derivatives PtPz1-PtPz6 were synthesised and characterised (PtPz1 - [Pt2N-hydroksymethyl-3,5-dimethylpyrazole4(berenil)2]Cl4; PtPz2 - [Pt23,5-dimethylpyrazole4(berenil)2]Cl4; PtPz3 - [Pt23,4-dimethylpyrazole4(berenil)2]Cl4; PtPz4 - [Pt2pyrazole4(berenil)2]Cl4; PtPz5- [Pt25-methylpyrazole4(berenil)2]Cl4; PtPz6 - [Pt2N-ethylpyrazole4(berenil)2]Cl4). The cytotoxic activity of these complexes against MCF-7 and MDA-MB-231 breast cancer cell lines was determined using the MTT assay. Evaluation of apoptosis induction was done with the Annexin V-fluorescein isothiocyanate/propidium iodide assay. In addition, using a flow cytometer, we determined the influence of test compounds on the cell cycle and caspase-3, -8, and -9 activity. The obtained results of caspase activity were confirmed by cell imaging. Moreover, using the flow cytometer, the effects of the test compounds on mitochondrial potential change were assessed. The test results showed that novel pyrazole-platinum(II) complexes exhibited stronger anti-proliferative activity against two breast cancer cell lines than reference cisplatin. Compounds PtPz1, PtPz2, and PtPz3 with methyl substituents at the pyrazole ring showed stronger activity than pyrazole or ethylpyrazole containing complexes. Studies have shown that inhibition of cell survival occurs by arresting the G1 cell cycle and inducing apoptosis. Our analysis associated with the response of MCF-7 and MDA-MB-231 cells to treatment with PtPz1-PtPz6 showed that it leads the cells through the external and intrinsic (mitochondrial) apoptotic pathway via indirect DNA damage.

Biological evaluation of octahydropyrazin[2,1-a:5,4-a']diisoquinoline derivatives as potent anticancer agents.

In this study, we evaluated the cytotoxic activity and antiproliferative potency of novel octahydropyrazin[2,1-a:5,4-a']diisoquinoline derivatives (1-7) in MCF-7 and MDA-MB-231 breast cancer cell lines. Annexin V binding assay and disruption of the mitochondrial potential were performed to determine apoptosis. The activity of caspases 3, 8, 9, and 10 was measured after 24 h of incubation with tested compounds to explain detailed molecular mechanism of induction of apoptosis. The results from experiments were compared with effects obtained after incubation in the presence of camptothecin and etoposide. Our study demonstrated that the most active compounds in both analyzed breast cancer cell lines were compounds 3 and 4. We also observed that all compounds induced apoptosis. We demonstrated the higher activity of caspases 3, 8, 9, and 10, which confirmed that induction of apoptosis is associated with external and internal cell death pathway. Our study revealed that the novel compounds in group of diisoquinoline derivatives are promising candidates in anticancer treatment by activation of both extrinsic and intrinsic apoptotic pathways.

Biological evaluation of dimethylpyridine-platinum complexes with potent antiproliferative activity.

This study investigates the effect of three new platinum complexes: Pt2(2,4-dimethylpyridine)4(berenil)2 (Pt14), Pt2(3,4-dimethylpyridine)4(berenil)2 (Pt15) and Pt2(3,5-dimethylpyridine)4(berenil)2 (Pt16) on growth and viability of breast cancer cells and their putative mechanism(s) of cytotoxicity. Cytotoxicity was measured with MTT assay and inhibition of [3H]thymidine incorporation into DNA in both breast cancer cells. Results revealed that Pt14-Pt16 exhibit substantially greater cytotoxicity than cisplatin against MCF-7 and MDA-MB-231 breast cancer cells. In the case of human skin fibroblast cell, cytotoxicity assays demonstrated that these compounds are less toxic to normal cells than cisplatin. In addition, the effects of Pt14-Pt16 are investigated using the flow cytometry assessment of annexin V binding, analysis of mitochondrial potential, markers of apoptosis such as caspase-3, caspase-8, caspase-9, caspase-10 and defragmentation of DNA by TUNEL assay. These results indicate that Pt14-Pt16 induce apoptosis by the mitochondrial and external pathway.

The combined treatment with novel platinum(II) complex and anti-MUC1 increases apoptotic response in MDA-MB-231 breast cancer cells.

New strategy of cancer's targeting treatment is combining monoclonal antibodies with chemotherapeutic agents. An important goal of targeted therapy appears to be a transmembrane glycoprotein type I-mucin 1 (MUC1), which is overexpressed in tumors of epithelial origin, especially in breast cancer. The goal of the study was to check the effect of monoclonal antibody against MUC1 with novel platinum(II) complex (Pt12) on selected aspects of apoptosis in human MDA-MB-231 breast cancer cells. The number of apoptotic and necrotic cells was measured using annexin V binding assay. The decrease of mitochondrial membrane potential (MMP) and DNA fragmentation was analyzed. Finally, the influence of novel platinum(II) complex (Pt12) used with anti-MUC1 on the concentration of selected markers of apoptosis such as Bax, caspase-8, -9, and caspase-3 was performed using ELISA. The results from combined treatment were compared with those obtained using monotherapy. In our study, we proved that anti-MUC1 used in combination with Pt12 strongly induced apoptosis in MDA-MB-231 breast cancer cell line. The effect was stronger than treatment with Pt12, cisplatin, anti-MUC1, and anti-MUC1 used with cisplatin. We also observed the highest decrease of MMP and the strongest DNA fragmentation after such a combined treatment. The results obtained from ELISA showed increased concentration of Bax, caspases-8, -9, -3 compared to monotherapy. Our study proved that Pt12 together with anti-MUC1 strongly induced apoptosis in estrogen-negative breast cancer cell line (MDA-MB-231). The apoptosis may go through extrinsic pathway associated with caspase-8 as well as intrinsic pathway connected with caspase-9.

Cytotoxic efficacy of a novel dinuclear platinum(II) complex used with anti-MUC1 in human breast cancer cells.

Mucin 1 (MUC1) is overexpressed in various cancer cells especially in breast cancer cells. There are known research works on the use of anti-MUC1 antibody with docetaxel in ovarian cancer, but there are no data about combined therapy platinum compounds with anti-MUC1 in breast cancer. The aim of the study was to evaluate the antiproliferative properties of a new dinuclear platinum(II) complex (Pt12) used with anti-MUC1 in human breast cancer cells. The dinuclear platinum(II) complex (Pt12) has been synthesized, and its cytotoxicity with anti-MUC1 has been tested in both MCF-7 and MDA-MB-231 breast cancer cells. In this study, the effects of Pt12 with anti-MUC1 on collagen and DNA biosynthesis in human breast cancer cells were compared to those evoked by cisplatin and cisplatin with anti-MUC1. The mechanism of action of Pt12 with anti-MUC1 was studied employing flow cytometry assessment of annexin V binding assay. It was found that Pt12 with anti-MUC1 was more active inhibitor of DNA and collagen synthesis as well more cytotoxic agent than Pt12 alone and cisplatin with anti-MUC1. Cytotoxicity of Pt12 with anti-MUC1 against breast cancer cells is due to apoptotic cell death as well as necrotic cell death. These results indicate that the use of Pt12 with anti-MUC1 may constitute a novel strategy in the chemotherapy of breast cancer tumors.