New high-molecular-weight contrast agents based on polyamidoamine (PAMAM) dendrimers for targeted imaging of malignant tumors characterized by overexpression of human epidermal growth factor receptor (EGFR) and human alpha-fetoprotein receptor (RECAF) were designed. Conjugates of second (G2) and third (G3) generation polyamidoamine dendrimers with 1,4,7,10-tetraazocyclodecane-1,4,7,10-tetraacetic acid (DOTA) were obtained. The quantitative composition of the conjugates was determined by 1HNMR spectroscopy. It was shown that four out of the 16 terminal NH2 groups in G2-DOTA and nine out of the 32 groups in G3-DOTA were modified with DOTA. The morphology, size, and charge of the synthesized macromolecules were characterized by dynamic light scattering and electrophoresis. Gadolinium(III) was loaded into the conjugates and the Gd content was determined by atomic emission spectroscopy. For increasing the selectivity of accumulation in the tumor cells, two recombinant proteins able to bind selectively to EGFR and RECAF, namely, human recombinant epidermal growth factor (rEGF) and human recombinant 3rd domain of alpha-fetoprotein (3dAFPpG), were conjugated with G2 and G3 dendrimers. The conjugates containing vector molecules were mainly accumulated via clathrin-dependent endocytosis, whereas G2-DOTA and G3-DOTA were absorbed via caveolin-dependent endocytosis and macropinocytosis. The dendrimer conjugates with vector molecules were intensely accumulated in A549 cells characterized by high expression of EGFR (Herl) and RECAF, whereas the accumulation of conjugates in the control K562 cells (with low expression of Her1) and in the CD14− population of human unstimulated mononuclear white blood cells was insignificant. The 3dAFPpG-conjugated dendrimers were partly recycled. All synthesized conjugates had a rather low toxicity in the range of 350–450 µmol L−1 (IC50).

The polymeric form ofjosamycin was developed by loading of the antibiotic into spherical submicron particles made of poly(DL- lactide-co-glycolide) (PLGA 50/50) by double emulsion method. The average diameter and zeta-potential of particles synthesized were less than 150 nm and -35 mV respectively. Polymeric form showed antimicrobial activity against gram-positive, gram-negative, and atypical bacteria. The highest antibacterial activity compared to substance (more than 8-fold) was detected against Enterococcus faecalis. The results of in vivo studies of specific activity of drugs on the model of staphylococcal sepsis in mice showed an increase in the efficacy of the studied polymeric form by 1.5-fold compared with the josamycin substance.

Reactive oxygen species are generated by the redox reaction involving metalloporphyrin and ascorbic acid (AA) and lead to oxidative stress followed by cancer cell death. Polymer particles based on the copolymer of lactic and glycolic acid (PLGA) containing Fe III Cl-tetraphenylporphyrin (FeClTPP) were prepared and characterized. These particles in combination with AA exhibit cytotoxic activity against the K562 (human chronic myelogenous leukemia) and MCF7 (human breast adenocarcinoma) cell lines. Results in vitro indicated significant antitumor efficiency on mice inoculated with P388 leukemic cells and treated with FeClTPP/AA. The cytotoxic activity of the combined system is achieved due to the formation of reactive oxygen species. The application of this system to the study of anticancer efficiency in vivo on the model of mice that were hypodermically inoculated with P388 lymphocytic leukemia revealed a significant inhibition of tumor growth. The use of FeClTPP in combination with AA seems to be promising in cancer treatment.

Silybin (Slb) and ursodeoxycholic acid (UDCA) are hepatoprotectors used in the pathogenetic therapy of liver and biliary tract. However, low bioavailability of these drugs restricts their application. In order to solve this problem, Slb and UDCA were incorporated into polymer carriers based on polylactic acid and poly(lactic-co-glycolic acid) by nanoprecipitation. The polymeric forms obtained according to the developed and optimized method are nanoparticles with a size from 100 to 200 μm. In vitro experiments showed a 1.5–2-fold higher hepatoprotective activity of Slb- and UDCA-containing polymeric nanoparticles compared to free substances.

The aggregation behavior of iron, manganese, and magnesium phthalocyanine (Pc) with poly-N-vinylpyrrolidone and polyethylene glycol was considered in this chapter. Biocidal activity of iron and manganese Pc complexes with polymer in relation to nine cultures of microorganisms was determined.

Method for the synthesis of polymeric nanoparticles (NP) with encapsulated daunorubicin (DNR) was developed on the basis of double emulsion solvent evaporation technique using biodegradable poly(lactide-co-glycolide) (PLGA), which is aimed at customization of pharmacokinetic properties of the preparation, enhanced accumulation of DNR in tumor cells and prolongation of its action. The obtained polymer nanoparticles (DNR-PLGA) had average size ranging around 138±36 nm, with zeta-potential of –25.3 mV and the polydispersity index (PDI) of 0.072. The release kinetics of DNR from polymer nanoparticles at pH 7.4 and 5.0 has been studied. In vitro studies showed similar specific activity of DNR- PLGA in K562 and MCF-7 cancer cell lines together with an increase in activity in K562 Adr and MCF-7 Adr cell lines, which are anthracycline resistant, by 1.6 and 3.4 times. The study demonstrated the efficacy of the developed PLGA-based DNR delivery system in the improvement of antitumor effect of DNR, overcoming multidrug resistance in cancer cells, and also in the decrease in nonspecific toxicity of the preparation.

In order to study the possibility of using a metal complex of the porphyrin series, FeIIICl-tetraphenylporphyrin (FeClTPP), and its polymeric form as antitumor agents for binary catalytic therapy, the technology of preparation of polymer particles containing FeClTPP was developed for the first time. The experimental data obtained allow one to conclude that the developed form using a copolymer of lactic and glycolic acids (ratio of monomer units 50: 50), ultrasonic homogenization, D-mannitol (as a cryoprotectant), the active agent to polymer ratio 1: 10, and the organic to aqueous phase ratio 1: 10 and 1: 20, respectively, has the optimal physicochemical parameters. It was found that the free substance of FeClTPP and polymer particles with FeClTPP are active against MCF-7 (human breast adenocarcinoma), HeLa (human cervical carcinoma), and MG-63 (human osteosarcoma) cell lines. The polymer particles containing FeClTFP exhibit the highest cytotoxic effect against MCF-7 lines as compared to the free substance. The results of the study indicate that both the substance of tetraphenylporphyrin and its resulting polymeric form are promising for the treatment of tumor diseases in binary catalytic therapy.

Photochemical synthesis of silver nanoparticles was proposed. Nanoparticle water formulations and films can be used for disinfection treatment, preventing the multiplication of dangerous microorganisms in the premises, equipment of the food industry, and in medicine.

To investigate the patterns of uptake of PLGA-PEG nanoparticles by human cancer cells, FITC-labeled polymer nanoparticles (FPN) are prepared and their properties are characterized. The amount of internalized FPN is measured by flow cytometry after quenching the fluorescence of the membrane-bound nanoparticles with Trypan Blue. The total uptake of the FPN by COLO 320 HSR and SW837 human cancer cells is shown to be proportional to the nanoparticle concentration and incubation time over a period of 24 h. The highest intracellular accumulation of the FPN is observed at 1 or 4 h, depending on the cell type. The translocation of the biodegradable and biocompatible polymer nanoparticles into the cancer cells that we observe may be the mechanism for enhancing the efficiency of the anticancer drug by loading it into these nanocarriers owing to a prolonged increase in the intracellular concentration of the therapeutic agent and the facilitation of its interaction with intracellular targets.

A new pharmaceutical form of the antitumor drug docetaxel (Dtx) has been developed based on a biodegradable copolymer of lactic and glycolic acids (PLGA 50/50). This form was constituted of polymeric particles (250 to 300 nm in size) and was highly active against mouse mammary adenocarcinoma (Ca755) and human breast adenocarcinoma (MCF-7 Wt ) cell lines. The results of the in vivo studies carried out on the C 57 Bl/6 female mice line showed an increase in the life span of the animals treated with the polymeric particles. Furthermore, the polymeric drug at a dose of 10 mg/kg and the drug substance at a dose of 20 mg/kg manifested a similar effect. At the same time, the acute toxicity of the Dtx polymeric form was 2.3 times lower than that of the drug substance.

Изменение популяции стволовых клеток рака молочной железы линии MCF-7 при одиночном и комбинированном действии ионизирующего излучения и конъюгатов дендритных полимеров с доксорубициномМатчук О

Second-generation (G2) polyamidoamine (PAMAM) dendrimers are branched polymers containing 16 surface primary amine groups. Due to their structural properties, these polymers can be used as universal carriers in various drug delivery systems. Amine-terminated PAMAM dendrimers are characterized by a high positive surface charge, leading to effective but nonspecific interactions with negatively charged cell plasmatic membranes. To reduce the nonspecific internalization of PAMAM dendrimers, their primary amine groups are often modified by acetic or succinic anhydrides, polyethylene glycol derivatives and other compounds. In this work, the role of primary amine groups, which are localized on the surface of doxorubicin-conjugated (Dox) dendrimers, was studied with regard to their intracellular distribution and internalization rates using SKOV3 human ovarian adenocarcinoma cells. It was demonstrated that all Dox-labeled G2-derivatives containing different numbers of acetamide groups synthesized in this work show high rates of cellular uptake at 37°С. As expected, the conjugate carrying the maximum number of primary amine groups demonstrated the highest rates of binding and endocytosis. At the same time, the G2-Dox conjugate containing the maximum number of acetamide groups showed colocalization with LAMP2, a marker of lysosomes and late endosomes, as well as the highest level of cytotoxic activity against SKOV3 cells. We conclude that second-generation PAMAM dendrimers are characterized by varied pathways of internalization and intracellular distribution due to the number of primary amine groups on their surface and, as a consequence, a different surface charge.

The dendritic polymers (dendrimers) are perspective nanocontainers for targeted transport of anticancer drugs to tumor cells. We used polyamidoamine dendrimers of the second generation (G2) covalently conjugated with doxorubicin (Dox) and vector protein - recombinant third domain (3D) of alpha-fetoprotein. The objects of the study were MCF-7/MDR1 breast cancer cells, which demonstrated resistance to traditional anticancer agents due to high expression of P-glycoprotein. Effects of free Dox, G2 dendrimers loaded with Dox (G2-Dox), or conjugates of dendrimers with the vector protein and Dox (3D-G2-Dox) were assessed by the criteria of surviving cell number and clonogenic activity 24 hours and 11 days after treatment with the agents at Dox concentration of 2.5 mu M, correspondingly. Flow cytometry was used to evaluate accumulation of Dox immediately after the treatment with the agents and removal of Dox during 24 hours of incubation in agent-free medium following by the treatment. Intracellular localization of Dox was studied using laser scanning microscopy. 3D-G2-Dox demonstrated the highest accumulation and the weakest removal from the cells in comparison with all other agents. The use of free Dox, G2-Dox, or 3D-G2-Dox resulted in a significant decrease in number of surviving cells by approximately 25-30% compared to the control (p <= 0.01). However, the most pronounced decrease in the clonogenic ability of cells was observed in the 3D-G2-Dox group (to 19% compared to the control, p < 0.01). Taking into account the previously obtained data on the extremely low 3D-G2-Dox accumulation in normal cells, it can be concluded that further development of 3D-G2-Dox as a possible anticancer drug is a promising way to overcome multiple drug resistance with minimal impact on normal cells.

Polymer particles containing carboplatin (CPt) were developed by the inclusion of this antitumor agent in a copolymer of lactic and glycolic acids (PLGA-COOH 50/50). The polymer particles were found to have a spherical form with an average diameter not exceeding 200 nm, the ζ-potential is equal to–32.2±1 mV. The CPt-loaded polymer particles possess a cytotoxic activity against human small cell and non-small cell lung carcinoma (lines H69 and A549), as well as against mouse mammary adenocarcinoma (line Ca755). The results of the in vivo studies carried out on female mice of line C57Bl/6 with inoculated mouse melanoma of line B16 showed increasing of lifespan of the animals and inhibition of tumor growth for groups treated with the polymer particles, as compared to the animals treated with the drug substance CPt.

Temozolomide (TMZ) is a cytostatic drug used in treatment of patients with malignant gliomas and melanomas. The development of innovative formulations for delivery of TMZ into target cells in order to reduce its systemic toxicity and gain prolonged effect is a topical problem of modern biotechnology and pharmacology. The article presents experimental data on development of the TMZ polymer-based formulation. The TMZ formulation presents nanoparticles with average size of 300 nm exhibiting high in vitro cytotoxic activity against melanoma and glioma cells (cell lines C6, U377MG, B16, and Mel-10). Melanoma (B16 cell line) bearing mice (C57Bl/61) treated by TMZ polymer nanoparticles revealed inhibition of tumor growth and increase of the animal lifespan. Also, loading of TMZ into the polymer particles was shown to decrease acute toxicity of the drug compared to the intact drug.

A recombinant alpha-fetoprotein (rAFP) was obtained in the yeast P. pastoris system, and its functional activity was confirmed. A method for producing polymer particles loaded with dactinomycin was developed, and a conjugate of these nanoparticles with rAFP was synthesized. The efficiency of the obtained conjugate on the HeLa, SKOV3, and MG-63 tumor cells and the absence of toxicity on the normal cells was shown. Experiments in vivo demonstrated a significant increase in the antitumor efficacy of the conjugate at a lower general toxicity as compared to the commercially available dactinomycin.

Accumulation of doxorubicin (Dox), its conjugates with the second generation dendritic polymer (G2-Dox) and vector pro- tein (recombinant third domain of alpha-fetoprotein - 3D-G2- Dox) in normal and tumor cells was studied in vitro within the framework of the development of selective transport system of anticancer drugs to the target cells. The objects of the study were cells of peripheral blood mononuclear fraction of healthy donors and cells of breast adenocarcinoma lines MCF-7 and MCF-7/MDR1, differing in chemosensitivity. G2-Dox and 3D-G2-Dox accumulated in tumor cells of the both lines better than free Dox (p<0,05). However removal of these drugs out of cells MCF-7 and MCF-7/MDR1 was significantly different: in the latter case all free Dox was excluded from the cells for 24 hours while Dox, accumulated in composition with dendrimers, still remained in the cells. It was important that 3D-G2-Dox (unlike the G2-Dox) accumulated in normal cells worse than free Dox (p<0.01). Thus, the results indicate that the use of 3D-G2-Dox is the most promising because it accumulates in tumor cells better and in normal cells worse than free Dox. Furthermore it can be assumed that the use of 3D-G2-Dox would be especially useful in cases of multi-drug resistance associated with the high expression of P-glycoprotein.