Lipid-coated ruthenium dendrimer conjugated with doxorubicin in anti-cancer drug delivery: Introducing protocols.

One of the major limitations for the treatment of many diseases is an inability of drugs to cross the cell membrane barrier. Different kinds of carriers are being investigated to improve drug bioavailability. Among them, lipid or polymer-based systems are of special interest due to their biocompatibility. In our study, we combined dendritic and liposomal carriers and analysed the biochemical and biophysical properties of these formulations. Two preparation methods of Liposomal Locked-in Dendrimers (LLDs) systems have been established and compared. Carbosilane ruthenium metallodendrimer was complexed with an anti-cancer drug (doxorubicin) and locked in a liposomal structure, using both techniques. The LLDs systems formed by hydrophilic locking had more efficient transfection profiles and interacted with the erythrocyte membrane better than systems using the hydrophobic method. The results indicate these systems have improved transfection properties when compared to non-complexed components. The coating of dendrimers with lipids significantly reduced their hemotoxicity and cytotoxicity. The nanometric size, low polydispersity index and reduced positive zeta potential of such complexes made them attractive for future application in drug delivery. The formulations prepared by the hydrophobic locking protocol were not effective and will not be considered furthermore as prospective drug delivery systems. In contrast, the formulations formed by the hydrophilic loading method have shown promising results where the cytotoxicity of LLD systems with doxorubicin was more effective against cancer than normal cells.

The effect of novel tyrosine-modified polyethyleneimines on human albumin structure - Thermodynamic and spectroscopic study.

The interaction of proteins with nanoparticle components are crucial for the evaluation of nanoparticle function, toxicity and biodistribution. Polyethyleneimines (PEIs) with defined tyrosine modifications are a class of novel polymers designed for improved siRNA delivery. Their interactions with biomacromolecules are still poorly described. This paper analyzes the interaction of different tyrosine-modified PEIs with human serum albumin as the most abundant serum protein. The ability of tyrosine modified, linear or branched PEIs to bind human serum albumin (HSA) was analyzed and further characterized. The interaction with hydrophobic parts of protein were studied using 1- nilinonaphthalene-8-sulfonic acid (ANS) and changes in the HSA secondary structure were evaluated using circular dichroism (CD). Complex formation and sizes were studied by transmission electron microscopy (TEM) and dynamic light scattering methods (DLS). We demonstrate that tyrosine modified PEIs are able to bind human serum albumin. Based on thermodynamic studies, van der Waals interaction, H-bonding and hydrophobic interactions are determined as main molecular forces involved in complex formation. Analysis of secondary structures revealed that the polymers decreased α-helix content, while increasing levels of randomly folded structures. Complex formation was confirmed by TEM and DLS. These findings are crucial for understanding polymer-protein interactions and the properties of nanoparticles.

Carbosilane ruthenium metallodendrimer as alternative anti-cancer drug carrier in triple negative breast cancer mouse model: A preliminary study.

The carbosilane metallodendrimer G1-[[NCPh(o-N)Ru(η6- p-cymene)Cl]Cl]4 (CRD13), based on an arene Ru(II) complex coordinated to imino-pyridine surface groups, has been conjugated with anti-cancer drugs. Ruthenium in the positively-charged dendrimer structure allows this nanoparticle to be considered as an anticancer drug carrier, made more efficient because ruthenium has anticancer properties. The ability of CRD13 to form complexes with Doxorubicin (DOX), 5-Fluorouracil (5-Fu), and Methotrexate (MTX) has been evaluated using zeta potential measurement, transmission electron microscopy (TEM) and computer simulation. The results show that it forms stable nanocomplexes with all those drugs, enhancing their effectiveness against MDA-MB-231 cancer cells. In vivo tests indicate that the CRD13/DOX system caused a decrease of tumor weight in mice with triple negative breast cancer. However, the tumors were most visibly reduced when naked dendrimers were injected.

Unmodified and tyrosine-modified polyethylenimines as potential carriers for siRNA: Biophysical characterization and toxicity.

Polyethylenimines (PEIs) are being explored as efficient non-viral nanocarriers for nucleic acid delivery in vitro and in vivo. To address limitations regarding PEI efficacy and biocompatibility, modifications of the chemical structure of linear and branched PEIs have been introduced, including grafting with tyrosine. The aim has been to compare linear and branched polyethylenimines of a wider range of different molecular mass with their tyrosine-modified derivatives. To do so, physico-chemical and biological properties of the polymers were investigated. Even in the absence of a negatively charged nucleic acid counterpart, PEIs form particle structures with defined size and surface potential. Tyrosine modification of PEI led to significantly reduced toxicity, while simultaneously increasing interaction with cellular membranes. All the effects were also dependent on the PEI molecular weight and structure (i.e., linear vs. branched). Especially in the case of linear PEIs, the improved membrane interaction also translated into slightly enhanced hemolysis, whereas their genotoxic potential was essentially abolished. Due to the improvement of properties critical for nano-vector efficacy and biocompatibility, our data demonstrate that tyrosine-modified PEIs are very promising and safe nanocarriers for the delivery of small RNAs, like siRNAs and miRNAs.

Hippophae rhamnoides L. leaf and twig extracts as rich sources of nutrients and bioactive compounds with antioxidant activity.

Plants have served for centuries as sources of compounds useful for human health such as antioxidant, anti-diabetic and antitumor agents. They are also rich in nutrients that improve the human diet. Growing demands for these compounds make it important to seek new sources for them. Hippophae rhamnoides L. is known as a plant with health-promoting properties. In this study we investigated the chemical composition and biological properties of bioactive components of ethanol extracts from leaves and twigs of H. rhamnoides L. Chemical components such as the total content of phenolic compounds, vitamins and amino acids and the antioxidant activities of these compounds in cellular and cell-free systems were assessed. The results suggest that the studied extracts are rich in bioactive compounds with potent antioxidant properties. Cytotoxicity and hemotoxicity assays showed that the extracts had low toxicity on human cells over the range of concentrations tested. Interaction with human serum albumin was investigated and conformational changes were observed. Our results indicate that leaf and twig extracts of H. rhamnoides L. should be considered as a non-toxic source of bioactive compounds which may be of interest to the food, pharmaceutical and cosmetic industries.

Comparison of tyrosine-modified low molecular weight branched and linear polyethylenimines for siRNA delivery.

Polyethylenimines (PEIs) have been previously introduced for siRNA delivery. In particular, in the case of higher molecular weight PEIs, this is associated with toxicity, while low molecular weight PEIs are often insufficient for siRNA complexation. The tyrosine-modification of PEIs has been shown to enhance PEI efficacy and biocompatibility. This paper evaluates a set of tyrosine-modified low molecular weight linear or branched polyethylenimines as efficient carriers of siRNA. Complexation efficacies and biophysical complex properties were analyzed by zeta potential, dynamic light scattering and circular dichroism measurements as well as gel electrophoresis. Biological knockdown was studied in 2 D cell culture and 3 D ex vivo tissue slice air-liquid interface culture. The results demonstrate that siRNAs were able to form stable complexes with all tested polymers. Complexation was able to protect siRNA from degradation by RNase and to mediate target gene knockdown, as determined on the mRNA level and in PC3-Luc3/EGFP and HCT116-Luc3/EGFP expressing reporter cells on the protein level, using flow cytometry and confocal microscopy. The direct comparison of the studied polymers revealed differences in biological efficacies. Moreover, the tyrosine-modified PEIs showed high biocompatibility, as determined by LDH release and mitochondria integrity (J-aggregate assay) as well as caspase 3/7 (apoptosis) and H2O2 levels (ROS). In 3 D tissue slices, complexes based on LP10Y proved to be most efficient, by combining tissue penetration with efficient gene expression knockdown.

Molecular Targets of Natural Compounds with Anti-Cancer Properties.

Cancer is the second leading cause of death in humans. Despite rapid developments in diagnostic methods and therapies, metastasis and resistance to administrated drugs are the main obstacles to successful treatment. Therefore, the main challenge should be the diagnosis and design of optimal therapeutic strategies for patients to increase their chances of responding positively to treatment and increase their life expectancy. In many types of cancer, a deregulation of multiple pathways has been found. This includes disturbances in cellular metabolism, cell cycle, apoptosis, angiogenesis, or epigenetic modifications. Additionally, signals received from the microenvironment may significantly contribute to cancer development. Chemical agents obtained from natural sources seem to be very attractive alternatives to synthetic compounds. They can exhibit similar anti-cancer potential, usually with reduced side effects. It was reported that natural compounds obtained from fruits and vegetables, e.g., polyphenols, flavonoids, stilbenes, carotenoids and acetogenins, might be effective against cancer cells in vitro and in vivo. Several published results indicate the activity of natural compounds on protein expression by its influence on transcription factors. They could also be involved in alterations in cellular response, cell signaling and epigenetic modifications. Such natural components could be used in our diet for anti-cancer protection. In this review, the activities of natural compounds, including anti-cancer properties, are described. The influence of natural agents on cancer cell metabolism, proliferation, signal transduction and epigenetic modifications is highlighted.

Nanoparticles for local delivery of siRNA in lung therapy.

An overview of the application of natural and synthetic, non-viral vectors for oligonucleotide delivery into the lung is presented in this review, with a special focus on lung cancer. Due to the specificity of the respiratory tract, its structure and natural barriers, the administration of drugs (especially those based on nucleic acids) is a particular challenge. Among widely tested non-viral drug and oligonucleotides carriers, synthetic polymers seem to be most promising. Unique properties of these nanoparticles allow for essentially unlimited possibilities regarding their design and modification. This gives hope that optimal nanoparticles with ideal nucleic acid carrier properties for lung cancer therapy will eventually emanate.

Combined therapy of ruthenium dendrimers and anti-cancer drugs against human leukemic cells.

Carbosilane ruthenium(ii) dendrimers have been complexed with conventional anti-cancer drugs. Due to its features, the presence of ruthenium within a dendrimer structure improves the anti-cancer properties of nanocomplexes containing 5-flurouracyl, methotrexate and doxorubicin. These dendrimers could be promising carriers of anti-cancer medicines. Ruthenium dendrimers that are positively charged can also enhance the cytotoxicity to cancer cells; moreover, they can form stable complexes with drugs. Results indicate that ruthenium dendrimers combined with doxorubicin and methotrexate significantly reduced the viability of leukaemia 1301 and HL-60 cancer cells.

Tyrosine-modified linear PEIs for highly efficacious and biocompatible siRNA delivery in vitro and in vivo.

Therapeutic gene silencing by RNA interference relies on the safe and efficient in vivo delivery of small interfering RNAs (siRNAs). Polyethylenimines are among the most studied cationic polymers for gene delivery. For several reasons including superior tolerability, small linear PEIs would be preferable over branched PEIs, but they show poor siRNA complexation. Their chemical modification for siRNA formulation has not been extensively explored so far. We generated a set of small linear PEIs bearing tyrosine modifications (LPxY), leading to substantially enhanced siRNA delivery and knockdown efficacy in vitro in various cell lines, including hard-to-transfect cells. The tyrosine-modified linear 10 kDa PEI (LP10Y) is particularly powerful, associated with favorable physicochemical properties and very high biocompatibility. Systemically administered LP10Y/siRNA complexes reveal antitumor effects in mouse xenograft and patient-derived xenograft (PDX) models, and their direct application into the brain achieves therapeutic inhibition of orthotopic glioma xenografts. LP10Y is particularly interesting for therapeutic siRNA delivery.

Synthesis and Characterization of FITC Labelled Ruthenium Dendrimer as a Prospective Anticancer Drug.

Metallodendrimers-dendrimers with included metals-are widely investigated as biocompatible equivalents to metal nanoparticles. Applications can be expected in the fields of catalysis, as chemical sensors in molecular recognition and as anticancer drugs. Metallodendrimers can also mimic certain biomolecules, for example, haemoprotein in the case of using a dendrimer with a porphyrin core. In previous papers, we showed the promising anticancer effects of carbosilane ruthenium dendrimers. The present paper is devoted to studying biocompatibility and the cytotoxic effect on normal and cancer cells of carbosilane ruthenium dendrimers labelled with fluorescent probe fluorescein isothiocyanate (FITC). The addition of fluorescent probe allowed tracking the metallodendrimer in both normal and cancer cells. It was found that carbosilane ruthenium dendrimer labelled with FITC in concentration up to 10 µmol/L was more cytotoxic for cancer cells than for normal cells. Thus, FITC labelled carbosilane ruthenium dendrimer is a good candidate for diagnostic imaging and studying anticancer effects of metallodendrimers in cancer therapy.

In Vitro Anticancer Properties of Copper Metallodendrimers.

Newly synthesized carbosilane copper dendrimers (CCD) with chloride and nitrate surface groups seem to be good candidates to be used as gene and drug carriers in anti-cancer therapy, due to their properties such as size and surface charge. Copper attached to the nanoparticles is an important element of many biological processes and recently their anti-cancer properties have been widely examined. Zeta size and potential, transmission electron microscopy (TEM), circular dichroism (CD), analysis of haemolytic activity, and fluorescence anisotropy techniques were used to characterize copper dendrimers. Additionally, their cytotoxic properties toward normal (PBMC) and cancer (1301; HL-60) cells were examined. All tested dendrimers were more cytotoxic against cancer cells in comparison with normal cells.

Dose and drug changes in chronic lymphocytic leukemia cell response in vitro: A comparison of standard therapy regimens with two novel cyclin­dependent kinase inhibitors.

Chronic lymphocytic leukemia (CLL) treatment is improving; however, some patients do not respond to therapy. Due to the high heterogeneity in disease development, there is an urgent need for personalization of therapy. In the present study, the response of leukemic mononuclear cells to anticancer drugs used for CLL treatment (cladribine + mafosfamide; CM or CM combined with rituximab; RCM) was compared with the response to new cyclin­dependent kinase (CDK) inhibitors: BP14 and BP30. Viable apoptotic and necrotic cells were quantified by flow cytometry using propidium iodide and Yo­Pro stains. CDK inhibitors were studied in several doses to determine the reduction of necrosis and simultaneous increase of apoptosis in leukemic cell incubations with anticancer agents. The distinct cell response to applied doses/anticancer agents was observed. Results obtained in the current manuscript confirmed that modulation of doses is important. This was particularly indicated in results obtained at 24 h of cells incubation with anticancer agent. While an important time for analysis of anticancer response efficacy (monitoring of apoptosis induction potential) seems to be 48 h of cells exposition to anticancer agents. High variability in response to the drugs revealed that both the nature and the dose of the anticancer agents could be important in the final effect of the therapy. The present findings support the thesis that personalized medicine, before drug administration in the clinic, could be important to avoid the application of ineffective therapy.

[Evolution of monoclonal antibodies in cancer treatment]. / Ewolucja przeciwcial monoklonalnych w leczeniu chorób nowotworowych.

Since late 90s of last century the new age of directed therapy began using mainly biological constructs produced in rodents called monoclonal antibodies. The side effects of monoclonal antibodies were a challenge for pharmaceutical companies to improve the biological properties of these biological drugs. The humanization of monoclonal constructs was an idea to improve monoclonal antibodies next generation activity cancer cell reduction in humans. Moreover for some other patients sensitive for monoclonal antibodies therapy could also potentially induce immunological differences that might imply on human health. The new idea related to monoclonal antibodies was to design a small molecule constructs of nanoantibodies with ability to enter into cells. Such small molecules could find their targets inside human cells, even in nuclei leading to differences in cancer cells expression. The existing knowledge on monoclonal antibodies as well as directed activity of nanoantibodies could improve anticancer treatment efficancy of diseases.