Antitumor Effect of Iscador on Breast Cancer Cell Lines with Different Metastatic Potential.

Studies were performed for the first time on the effect of Iscador Qu and Iscador M on phototoxicity, cytotoxicity, antiproliferative activity, changes in ξ-potential of cells, membrane lipid order, actin cytoskeleton organization and migration on three breast cancer lines with different metastatic potential: MCF10A (control), MCF-7 (low metastatic) and MDA-MB231 (high metastatic) cells. The tested Iscador Qu and M did not show any phototoxicity. The antiproliferative effect of Iscador species appeared to be dose-dependent and was related to the metastatic potential of the tested cell lines. A higher selectivity index was obtained for Iscador Qu and M towards the low metastatic MCF-7 cell line compared to the high metastatic MDA-MB-231. Iscador Qu demonstrated higher selectivity for both cancer cell lines compared to Iscador M. The malignant cell lines exhibited a decrease in fibril number and thickness regardless of the type of Iscador used. The strongest effect on migration potential was observed for the low metastatic cancer cell line MCF-7 after Iscador treatment. Both Iscador species induced a slight increase in the percentage of cells in early apoptosis for the low and high metastatic cell lines, MCF-7 and MDA-MB-231, unlike control cells. Changes in the zeta potential and membrane lipid order were observed for the low metastatic MCF-7 cell line in contrast to the high metastatic MDA-MB-231 cells. The presented results reveal a higher potential of Iscador as an antitumor agent for the low metastatic cancer cell line MCF-7 compared to the high metastatic one. Iscador Qu appears to be more potent compared to Iscador M, but at this point, the exact mechanism of action is still unclear and needs further investigations.

Impact of Truncated Oxidized Phosphatidylcholines on Phospholipase A2 Activity in Mono- and Polyunsaturated Biomimetic Vesicles.

The interplay between inflammatory and redox processes is a ubiquitous and critical phenomenon in cell biology that involves numerous biological factors. Among them, secretory phospholipases A2 (sPLA2) that catalyze the hydrolysis of the sn-2 ester bond of phospholipids are key players. They can interact or be modulated by the presence of truncated oxidized phosphatidylcholines (OxPCs) produced under oxidative stress from phosphatidylcholine (PC) species. The present study examined this important, but rarely considered, sPLA2 modulation induced by the changes in biophysical properties of PC vesicles comprising various OxPC ratios in mono- or poly-unsaturated PCs. Being the most physiologically active OxPCs, 1-palmitoyl-2-(5'-oxo-valeroyl)-sn-glycero-3-phosphocholine (POVPC) and 1-palmitoyl-2-glutaryl-sn-glycero-3-phosphocholine (PGPC) have been selected for our study. Using fluorescence spectroscopy methods, we compared the effect of OxPCs on the lipid order as well as sPLA2 activity in large unilamellar vesicles (LUVs) made of the heteroacid PC, either monounsaturated [1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)], or polyunsaturated [1-palmitoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine (PDPC)] at a physiological temperature. The effect of OxPCs on vesicle size was also assessed in both the mono- and polyunsaturated PC matrices. Results: OxPCs decrease the membrane lipid order of POPC and PDPC mixtures with PGPC inducing a much larger decrease in comparison with POVPC, indicative that the difference takes place at the glycerol level. Compared with POPC, PDPC was able to inhibit sPLA2 activity showing a protective effect of PDPC against enzyme hydrolysis. Furthermore, sPLA2 activity on its PC substrates was modulated by the OxPC membrane content. POVPC down-regulated sPLA2 activity, suggesting anti-inflammatory properties of this truncated oxidized lipid. Interestingly, PGPC had a dual and opposite effect, either inhibitory or enhancing on sPLA2 activity, depending on the protocol of lipid mixing. This difference may result from the chemical properties of the shortened sn-2-acyl chain residues (aldehyde group for POVPC, and carboxyl for PGPC), being, respectively, zwitterionic or anionic under hydration at physiological conditions.

Redox-Cycling "Mitocans" as Effective New Developments in Anticancer Therapy.

Our study proposes a pharmacological strategy to target cancerous mitochondria via redox-cycling "mitocans" such as quinone/ascorbate (Q/A) redox-pairs, which makes cancer cells fragile and sensitive without adverse effects on normal cells and tissues. Eleven Q/A redox-pairs were tested on cultured cells and cancer-bearing mice. The following parameters were analyzed: cell proliferation/viability, mitochondrial superoxide, steady-state ATP, tissue redox-state, tumor-associated NADH oxidase (tNOX) expression, tumor growth, and survival. Q/A redox-pairs containing unprenylated quinones exhibited strong dose-dependent antiproliferative and cytotoxic effects on cancer cells, accompanied by overproduction of mitochondrial superoxide and accelerated ATP depletion. In normal cells, the same redox-pairs did not significantly affect the viability and energy homeostasis, but induced mild mitochondrial oxidative stress, which is well tolerated. Benzoquinone/ascorbate redox-pairs were more effective than naphthoquinone/ascorbate, with coenzyme Q0/ascorbate exhibiting the most pronounced anticancer effects in vitro and in vivo. Targeted anticancer effects of Q/A redox-pairs and their tolerance to normal cells and tissues are attributed to: (i) downregulation of quinone prenylation in cancer, leading to increased mitochondrial production of semiquinone and, consequently, superoxide; (ii) specific and accelerated redox-cycling of unprenylated quinones and ascorbate mainly in the impaired cancerous mitochondria due to their redox imbalance; and (iii) downregulation of tNOX.

2-Alkyl-Substituted-4-Amino-Thieno[2,3-d]Pyrimidines: Anti-Proliferative Properties to In Vitro Breast Cancer Models.

Thienopyrimidines are structural analogs of quinazolines, and the creation of new 2-alkyl derivatives of ethyl 4-aminothienopyrimidine-6-carboxylates for the study of their anti-proliferative properties is of great pharmacological interest. Some 2-alkyl-4-amino-thieno[2,3-d]pyrimidines 2-5 were synthesized, and their cyto- and phototoxicity against BALB 3T3 cells were established by an in vitro 3T3 NRU test. The obtained results indicate that the tested compounds are not cytotoxic or phototoxic, and that they are appropriate to be studied for their anti-proliferative and anti-tumor properties. The anti-proliferative potential of the compounds was investigated on MCF-7 and MDA-MB-231 cancer cells, as well as a MCF-10A cell line (normal human mammary epithelial cells). The most toxic to MCF-7 was thienopyrimidine 3 with IC50 13.42 µg/mL (IC50 0.045 µM), followed by compound 4 (IC50 28.89 µg/mL or IC50 0.11 µM). The thienopyrimidine 4 revealed higher selectivity to MCF-7 and lower activity (IC50 367 µg/mL i.e., 1.4 µM) than compound 3 with MCF-10A cells. With respect to MDA-MB-231 cells, ester 2 manifested the highest effect with IC50 52.56 µg/mL (IC50 0.16 µM), and 2-ethyl derivative 4 revealed IC50 62.86 µg/mL (IC50 0.24 µM). It was estimated that the effect of the substances on the cell cycle progression was due to cell cycle arrest in the G2 stage for MDA-MB-231, while arrest in G1 was detected for the estrogen (ER)-positive MCF-7 cell line. The tested compound's effects on the change of the zeta potential in the tumorigenic cells utilized in this study were determined. The calculation which we performed of the physicochemical properties and pharmacokinetic parameters influencing the biological activity suggested high intestinal absorption, as well as drug-likeness.

Myconoside interacts with the plasma membranes and the actin cytoskeleton and provokes cytotoxicity in human lung adenocarcinoma A549 cells.

Studies have been carried out on the effects of the phenyl glycoside myconoside, extracted from the relict, Balkan endemic resurrection plant Haberlea rhodopensis on the plasma membrane structural organization and the actin cytoskeleton. Because the plasma membrane is the first target of exogenous bioactive compounds, we focused our attention on the influence of myconoside on the membrane lipid order and actin cytoskeleton in human lung adenocarcinoma A549 cells, using fluorescent spectroscopy and microscopy techniques. We found that low myconoside concentration (5 µg/ml) did not change cell viability but was able to increase plasma membrane lipid order of the treated cells. Higher myconoside concentration (20 µg/ml) inhibited cell viability by decreasing plasma membrane lipid order and impairing actin cytoskeleton. We hypothesize that the observed changes in the plasma membrane structural organization and the actin cytoskeleton are functionally connected to cell viability. Biomimetic membranes were used to demonstrate that myconoside is able to reorganize the membrane lipids by changing the fraction of sphingomyelin-cholesterol enriched domains. Thus, we propose a putative mechanism of action of myconoside on A549 cells plasma membrane lipids as well as on actin filaments in order to explain its cytotoxic effect at high myconoside concentration.

Resveratrol Affects Sphingolipid Metabolism in A549 Lung Adenocarcinoma Cells.

Resveratrol is a naturally occurring polyphenol which has various beneficial effects, such as anti-inflammatory, anti-tumor, anti-aging, antioxidant, and neuroprotective effects, among others. The anti-cancer activity of resveratrol has been related to alterations in sphingolipid metabolism. We analyzed the effect of resveratrol on the enzymes responsible for accumulation of the two sphingolipids with highest functional activity-apoptosis promoting ceramide (CER) and proliferation-stimulating sphingosine-1-phosphate (S1P)-in human lung adenocarcinoma A549 cells. Resveratrol treatment induced an increase in CER and sphingosine (SPH) and a decrease in sphingomyelin (SM) and S1P. Our results showed that the most common mode of CER accumulation, through sphingomyelinase-induced hydrolysis of SM, was not responsible for a CER increase despite the reduction in SM in A549 plasma membranes. However, both the activity and the expression of CER synthase 6 were upregulated in resveratrol-treated cells, implying that CER was accumulated as a result of stimulated de novo synthesis. Furthermore, the enzyme responsible for CER hydrolysis, alkaline ceramidase, was not altered, suggesting that it was not related to changes in the CER level. The enzyme maintaining the balance between apoptosis and proliferation, sphingosine kinase 1 (SK1), was downregulated, and its expression was reduced, resulting in a decrease in S1P levels in resveratrol-treated lung adenocarcinoma cells. In addition, incubation of resveratrol-treated A549 cells with the SK1 inhibitors DMS and fingolimod additionally downregulated SK1 without affecting its expression. The present studies provide information concerning the biochemical processes underlying the influence of resveratrol on sphingolipid metabolism in A549 lung cancer cells and reveal possibilities for combined use of polyphenols with specific anti-proliferative agents that could serve as the basis for the development of complex therapeutic strategies.

Design, Cytotoxicity and Antiproliferative Activity of 4-Amino-5-methyl-thieno[2,3-d]pyrimidine-6-carboxylates against MFC-7 and MDA-MB-231 Breast Cancer Cell Lines.

Novel 4-amino-thieno[2,3-d]pyrimidine-6-carboxylates substituted at the second position were prepared by cyclocondensation of 2-amino-3-cyano-thiophene and aryl nitriles in an acidic medium. The design of the target compounds was based on structural optimization. The derivatives thus obtained were tested in vitro against human and mouse cell lines. The examination of the compound effects on BLAB 3T3 and MFC-10A cells showed that they are safe, making them suitable for subsequent experiments to establish their antitumor activity. The photoirritancy factor of the compounds was calculated. Using the MTT test, the antiproliferative activity to MCF-10A, MCF-7 and MDA-MB-231 cell lines was estimated. The best antiproliferative effect in respect to the MCF-7 cell line revealed compound 2 with IC50 4.3 ± 0.11 µg/mL (0.013 µM). The highest selective index with respect to MCF-7 cells was shown by compound 3 (SI = 19.3), and to MDA-MB-231 cells by compound 2 (SI = 3.7). Based on energy analysis, the most stable conformers were selected and optimized by means of density functional theory (DFT). Ligand efficiency, ligand lipophilicity efficiency and the physicochemical parameters of the target 4-amino-thienopyrimidines were determined. The data obtained indicated that the lead compound among the tested substances is compound 2.

Passive and electro-assisted delivery of hydrogel nanoparticles in solid tumors, visualized by optical and magnetic resonance imaging in vivo.

The present study describes a development of nanohydrogel, loaded with QD(705) and manganese (QD(705)@Nanogel and QD(705)@Mn@Nanogel), and its passive and electro-assisted delivery in solid tumors, visualized by fluorescence imaging and magnetic resonance imaging (MRI) on colon cancer-grafted mice as a model. QD(705)@Nanogel was delivered passively predominantly into the tumor, which was visualized in vivo and ex vivo using fluorescent imaging. The fluorescence intensity increased gradually within 30 min after injection, reached a plateau between 30 min and 2 h, and decreased gradually to the baseline within 24 h. The fluorescence intensity in the tumor area was about 2.5 times higher than the background fluorescence. A very weak fluorescent signal was detected in the liver area, but not in the areas of the kidneys or bladder. This result was in contrast with our previous study, indicating that FITC@Mn@Nanogel did not enter into the tumor and was detected rapidly in the kidney and bladder after i.v. injection [J. Mater. Chem. B 2013, 1, 4932-4938]. We found that the embedding of a hard material (as QD) in nanohydrogel changes the physical properties of the soft material (decreases the size and negative charge and changes the shape) and alters its pharmacodynamics. Electroporation facilitated the delivery of the nanohydrogel in the tumor tissue, visualized by fluorescent imaging and MRI. Strong signal intensity was recorded in the tumor area shortly after the combined treatment (QD@Mn@Nanogel + electroporation), and it was observed even 48 h after the electroporation. The data demonstrate more effective penetration of the nanoparticles in the tumor due to the increased permeability of blood vessels at the electroporated area. There was no rupture of blood vessels after electroporation, and there were no artifacts in the images due to a bleeding.

Lymph node mapping using quantum dot-labeled polymersomes.

The present study was designed to investigate whether poly-ion complex hollow vesicles (polymersomes), based on chemically-modified chitosan, are appropriate for lymph node mapping in the context of their application in the development of theranostic nanosized drug delivery systems (nano-DDS). The experiments were performed on Balb/c nude mice (colon cancer-grafted). The mice were subjected to anesthesia and quantum dot (QD(705))-labeled polymersomes (d-120 nm) were injected intravenously via the tail vein. The optical imaging was carried out on Maestro EX Imaging System (excitation filter: 435-480 nm; emission filter: 700 nm). A strong fluorescent signal, corresponding to QD(705) fluorescence, was detected in the lymph nodes, as well as in the tumor. A very weak fluorescent signal was found in the liver area. The half-life of QD(705)-labelled polymersomes was 6 ± 2 hours in the bloodstream and 11 ± 3 hours in the lymph nodes. The data suggest that polymersomes are very promising carriers for lymph node mapping using QD as a contrast agent. They are useful matrix for development of nano-formulations with theranostic capabilities.

Delivery of size-controlled long-circulating polymersomes in solid tumours, visualized by quantum dots and optical imaging in vivo.

The present study was designed to investigate whether poly-ion complex hollow vesicles (polymersomes), based on chemically modified chitosan, are appropriate for passive tumour targeting in the context of their application as drug carriers. The experiments were performed on colon cancer-grafted mice. The mice were subjected to anaesthesia and injected intravenously with water-soluble nanoparticles: (1) QD705-labelled polymersomes (average size ∼120 nm; size distribution ∼10%) or (2) native QD705. The optical imaging was carried out on Maestro EX 2.10 In Vivo Imaging System (excitation filter 435-480 nm; emission filter 700 nm, longpass). In the case of QD705, the fluorescence appeared in the tumour area within 1 min after injection and disappeared completely within 60 min. A strong fluorescent signal was detected in the liver on the 30th minute. The visualization of tumour using QD705 was based only on angiogenesis. In the case of QD705-labelled polymersomes, the fluorescence appeared in the tumour area immediately after injection with excellent visualization of blood vessels in the whole body. A strong fluorescent signal was detected in the tumour area within 16 hours. This indicated that QD705-labelled polymersomes were delivered predominantly into the tumour due to their long circulation in the bloodstream and enhanced permeability and retention effect. A very weak fluorescent signal was found in the liver area. The data suggest that size-controlled long-circulating polymersomes are very promising carriers for drug delivery in solid tumours, including delivery of small nanoparticles and contrast substances.

Fluorescent imaging for assessment of the effect of combined application of electroporation and rifampicin on HaCaT cells as a new therapeutic approach for psoriasis.

The study aimed to clarify the role of electric pulses in combination with chemotherapy on the viability of keratinocyte cell line HaCaT, in the context of its application as a new therapeutic approach for psoriasis. The data show that electroporation of HaCaT cells in combination with rifampicin induces cytoskeleton disruption and increases permeability of cell monolayer due to cell-cell junctions' interruption, visualized by fluorescent imaging of E-cadherin and actin integrity. This was accompanied with synergistic reduction of cell viability. The study proposes a new opportunity for more effective skin treatment than chemotherapy. The future application of this electrochemotherapeutic approach for combined local treatment of psoriasis may have serous benefits because of a high possibility to avoid side-effects of conventional chemotherapy.

Docosahexaenoic Acid Potentiates the Anticancer Effect of the Menadione/Ascorbate Redox Couple by Increasing Mitochondrial Superoxide and Accelerating ATP Depletion.

BACKGROUND/AIM: Mitochondria-targeted anticancer drugs ("mitocans") of natural origin are attractive candidates as adjuvants in cancer therapy. The redox couple menadione/ascorbate (M/A), which belongs to the "mitocans" family, induces selective oxidative stress in cancerous mitochondria and cells, respectively. DHA has also been found to regulate the mevalonate pathway, which is closely related to the prenylation of the cytotoxic menadione to the non-cytotoxic menaquinone. The aim of this study was to elucidate the ability of docosahexaenoic acid (DHA) to potentiate the anticancer effect of M/A by increasing ROS production, as well as affecting steady-state ATP levels in cancer cells. MATERIALS AND METHODS: The experiments were performed on leukemic lymphocyte Jurkat. Cells were treated with DHA, M/A, and their combination (M/A/DHA) and four parameters were examined using the following assays: cell viability and proliferation, steady-state ATP, mitochondrial superoxide, intracellular hydroperoxides. Three independent experiments with two or six parallel measurements were performed for each parameter. RESULTS: The triple combination M/A/DHA was characterized by much higher antiproliferative activity and cytotoxicity than M/A and DHA administered alone. DHA significantly accelerated M/A-induced ATP depletion in cells, which was accompanied by an additional increase in mitochondrial superoxide compared to cells treated with M/A or DHA alone. CONCLUSION: DHA significantly enhanced M/A-induced cytotoxicity in leukemic lymphocytes by inducing severe mitochondrial oxidative stress and accelerated ATP depletion. Selective DHA-mediated suppression of cholesterol synthesis in cancer cells (involved in the prenylation of cytotoxic menadione to the less cytotoxic phylloquinone), as well as DHA-mediated inhibition of superoxide dismutase are suggested to underlie the potentiation of the anticancer effect of M/A.

Redox-mediated Anticancer Activity of Anti-parasitic Drug Fenbendazole in Triple-negative Breast Cancer Cells.

BACKGROUND/AIM: An increasing number of studies are reporting anticancer activity of widely used antiparasitic drugs and particularly benzimidazoles. Fenbendazole is considered safe and tolerable in most animal species at the effective doses as an anthelmintic. Little is known about the redox-modulating properties of fenbendazole and the molecular mechanisms of its antiproliferative effects. Our study aimed to investigate the possibility of selective redox-mediated treatment of triple-negative breast cancer cells by fenbendazole without affecting the viability and redox status of normal breast epithelial cells. MATERIALS AND METHODS: The experiments were performed on three cell lines: normal breast epithelial cells (MCF-10A) and cancer breast epithelial cells (MCF7 - luminal adenocarcinoma, low metastatic; MDA-MB-231 - triple-negative adenocarcinoma, highly metastatic). Cells were treated with fenbendazole for 48-h and three parameters were analyzed using conventional assays: cell viability and proliferation, level of intracellular superoxide, and level of hydroperoxides. RESULTS: The data demonstrated that MDA-MB-231 cells were more vulnerable to fenbendazole-induced oxidative stress than MCF-7 cells. In normal breast epithelial cells MCF-10A, fenbendazole significantly suppressed oxidative stress compared to untreated controls. These data correlate with the effect of fenbendazole on cell viability and the IC50 values, which is indirect evidence of the potential targeting anticancer effect of the drug, especially in MDA-MB-231 cells. CONCLUSION: The difference in the levels of oxidative stress induced by fenbendazole in MDA-MB-231 and MCF-7 indicates that the two types of breast cancer respond to the drug through different redox-related mechanisms.

Newly Synthesized Lignin Microparticles as Bioinspired Oral Drug-Delivery Vehicles: Flavonoid-Carrier Potential and In Vitro Radical-Scavenging Activity.

The aim of the present study was to synthesize lignin microparticles, to evaluate their physicochemical, spectral, morphological and structural characteristics, to examine their encapsulation and in vitro release potential and behaviour towards the flavonoid morin in simulated physiological medium and to assess the in vitro radical-scavenging potential of the morin-loaded lignin microcarrier systems. The physicochemical, structural and morphological characteristics of alkali lignin, lignin particles (LP) and morin-encapsulated lignin microparticles (LMP) were determined based on particle size distribution, SEM, UV/Vis spectrophotometric, FTIR and potentiometric titration analyses. The encapsulation efficiency of LMP was 98.1%. The FTIR analyses proved that morin was successfully encapsulated in the LP without unexpected chemical reactions between the flavonoid and the heteropolymer. The in vitro release performance of the microcarrier system was successfully mathematically described by Korsmeyer-Peppas and the sigmoidal models outlining the general role of diffusion during the initial stages of the in vitro release process in simulated gastric fluid (SGF), and the predominant contribution of biopolymer relaxation and erosion was determined in simulated intestinal medium (SIF). The higher radical-scavenging potential of LMP, as compared to that of LP, was proven via DPPH and ABTS assays. The synthesis of lignin microcarriers not only provides a facile approach for the utilization of the heteropolymer but also determines its potential for the design of drug-delivery matrices.

Electroporation, electrochemotherapy and electro-assisted drug delivery in cancer. A state-of-the-art review.

This review focuses on electrochemotherapy that consists in the delivery of anti-cancer drugs using high-voltage electrical pulses. Technical issues, choice of drugs, and protocol of drug delivery are still under investigation and no consensus has been achieved yet. The different aspects of electrochemotherapy are discussed in the present paper. It includes interrogations about the choice of the preferred anti-cancer drug and dose to be delivered on the solid tumors. Another promising area is related to the electro-assisted release of nanoparticles (quantum dots) in xenografted solid tumors. Molecular mechanisms of enhanced drug delivery are discussed in terms of high cholesterol level and large fraction of lipid rafts in cancer cells. Electrochemotherapy is a paradigmatic example of cooperation between physicists, biophysicists, chemists, technicians, manufacturers, biologists, clinicians, and patients to improve a very promising treatment delivery in line with the conception of personalized medicine.

Thieno[2,3-d]pyrimidin-4(3H)-one Derivatives of Benzimidazole as Potential Anti- Breast Cancer (MDA-MB-231, MCF-7) Agents.

AIMS: The purpose of this study was the synthesis of some new thienopyrimidine derivatives of 1,3-disubstituted benzimidazoles and the evaluation of their cytotoxicity against MDA-MB-231, MCF-7, and 3T3 cells lines. BACKGROUND: An overexpression or mutational activation of TK receptors EGFR and HER2/neu is characteristic of tumors. It has been found that some thieno[2,3-d]pyrimidines exhibited better inhibitory activity against Epidermal Growth Factor Receptor (EGFR/ErbB-2) tyrosine kinase in comparison to aminoquinazolines. Breast cancer activity towards MDA-MB-231 and MCF-7 cell lines by inhibiting EGFR was revealed by a novel 2-arylbenzimidazole. This motivated the synthesis of new thienopyrimidines possessing benzimidazole fragments in order to evaluate their cytotoxicity to the above-mentioned cell lines. OBJECTIVE: The objectives of the study were to design and synthesize a novel series of thieno[2,3-d]pyrimidines bearing biologically active moieties, such as 1,3-disubstituted-benzimidazole heterocycle, structurally similar to diaryl ureas in order to evaluate their cytotoxicity against MDA-MB-231, and MCF-7 breast cancer cell lines. METHODS: N,N-disubstituted benzimidazole-2-one carbonitriles were synthesized by Aza-Michael addition and used as precursors to generate some of the new thieno[2,3-d]pyrimidines in acidic medium The interaction of chloroethyl-2-thienopyrimidines, 2-amino-benzimidazole and benzimidazol-2-one nitriles under solid-liquid transfer catalysis conditions led to new thienopyrimidines. MTT assay for cell survival was performed in order to evaluate the cytotoxicity of the tested compounds. A fluorescence study was conducted to elucidate some aspects of the mechanism of action. RESULTS: The effects of nine synthesized compounds were investigated towards MDA-MB-231, MCF-7 and 3T3 cell lines. Thieno[2,3-d]pyirimidine-4-one 16 (IC50 - 0.058µM) and 21 (IC50 - 0.029µM) possess high cytotoxicity against MDA-MB-231 cells after 24h. The most cytotoxic compounds against breast cancer MCF-7 cells was compound 21 (IC50 - 0.074µM), revealing lower cytotoxicity against mouse fibroblast 3T3 cells with IC50 - 0.20µM. SAR analysis was performed. Fluorescence study of the treatment of MDA-MB cells with compound 21 was carried out in order to clarify some aspects of the mechanism of action. CONCLUSION: The relationship between cytotoxicity of compounds 14 and 20 against MCF-7 and 3T3 cells can suggest a similar mechanism of action. The antitumor potential of the tested compounds proves the necessity for further investigation to estimate the exact inhibition pathway in the cellular processes. The fluorescence study of the treatment of MDA-MB cells with compound 21 showed a rapid process of apoptosis.

Electrodelivery of drugs into cancer cells in the presence of poloxamer 188.

In the present study it is shown that poloxamer 188, added before or immediately after an electrical pulse used for electroporation, decreases the number of dead cells and at the same time does not reduce the number of reversible electropores through which small molecules (cisplatin, bleomycin, or propidium iodide) can pass/diffuse. It was suggested that hydrophobic sections of poloxamer 188 molecules are incorporated into the edges of pores and that their hydrophilic parts act as brushy pore structures. The formation of brushy pores may reduce the expansion of pores and delay the irreversible electropermeability. Tumors were implanted subcutaneously in both flanks of nude mice using HeLa cells, transfected with genes for red fluorescent protein and luciferase. The volume of tumors stopped to grow after electrochemotherapy and the use of poloxamer 188 reduced the edema near the electrode and around the subcutaneously growing tumors.

Redox-related Molecular Mechanism of Sensitizing Colon Cancer Cells to Camptothecin Analog SN38.

BACKGROUND/AIM: The aim of this study was to elucidate the possibility of sensitizing colon cancer cells to the chemotherapeutic drug SN38 and investigate its mechanism of action after combined treatment with electroporation (EP). MATERIALS AND METHODS: Cells were treated with SN38, EP and their combination for 24/48 h. The cell viability, actin cytoskeleton integrity, mitochondrial superoxide, hydroperoxides, total glutathione, phosphatidyl serine expression, DNA damages and expression of membrane ABC transporters were analyzed using conventional analytical tests. RESULTS: The combination of EP and SN38 affected cell viability and cytoskeleton integrity. This effect was accompanied by: (i) high production of intracellular superoxide and hydroperoxides and depletion of glutathione; (ii) increased DNA damage and apoptotic/ferroptotic cell death; (iii) changes in the expression of membrane ABC transporters - up-regulation of SLCO1B1 and retention of SN38 in the cells. CONCLUSION: The anticancer effect of the combined treatment of SN38 and EP is related to changes in the redox-homeostasis of cancer cells, leading to cell death via apoptosis and/or ferroptosis. Thus, electroporation has a potential to increase the sensitivity of cancer cells to conventional anticancer therapy with SN38.

Bacterial Natural Disaccharide (Trehalose Tetraester): Molecular Modeling and in Vitro Study of Anticancer Activity on Breast Cancer Cells.

Isolation and characterization of new biologically active substances affecting cancer cells is an important issue of fundamental research in biomedicine. Trehalose lipid was isolated from Rhodococcus wratislaviensis strain and purified by liquid chromatography. The effect of trehalose lipid on cell viability and migration, together with colony forming assays, were performed on two breast cancer (MCF7-low metastatic; MDA-MB231-high metastatic) and one "normal" (MCF10A) cell lines. Molecular modeling that details the structure of the neutral and anionic form (more stable at physiological pH) of the tetraester was carried out. The tentative sizes of the hydrophilic (7.5 Å) and hydrophobic (12.5 Å) portions of the molecule were also determined. Thus, the used trehalose lipid is supposed to interact as a single molecule. The changes in morphology, adhesion, viability, migration, and the possibility of forming colonies in cancer cell lines induced after treatment with trehalose lipid were found to be dose and time dependent. Based on the theoretical calculations, a possible mechanism of action and membrane asymmetry between outer and inner monolayers of the bilayer resulting in endosome formation were suggested. Initial data suggest a mechanism of antitumor activity of the purified trehalose lipid and its potential for biomedical application.

Characterization and potential antitumor effect of a heteropolysaccharide produced by the red alga Porphyridium sordidum.

Taking into account the rising trend of the incidence of cancers of various organs, effective therapies are urgently needed to control human malignancies. However, almost all chemotherapy drugs currently on the market cause serious side effects. Fortunately, several studies have shown that some non-toxic biological macromolecules, including algal polysaccharides, possess anti-cancer activities or can increase the efficacy of conventional chemotherapy drugs. Polysaccharides are characteristic secondary metabolites of many algae. The efficacy of polysaccharides on the normal and cancer cells is not well investigated, but our investigations proved a cell specific effect of a newly isolated extracellular polysaccharide from the red microalga Porphyridium sordidum. The investigated substance was composed of xylose:glucose and galactose:manose:rhamnose in a molar ratio of 1:0.52:0.44:0.31. Reversible electroporation has been exploited to increase the transport through the plasma membrane into the tested breast cancer tumor cells MCF-7 and MDA-MB231. Application of 75 µg/mL polysaccharide in combination with 200 V/cm electroporation induced 40% decrease in viability of MDA-MB231 cells and changes in cell morphology while control cells (MCF10A) remained with normal morphology and kept vitality.