Single-walled carbon nanotubes loaded hydroxyapatite-alginate beads with enhanced mechanical properties and sustained drug release ability.

Single-walled carbon nanotubes (SWCNTs) containing biomaterial with enhanced mechanical properties for the potential orthopedic application were synthesized and investigated. X-ray diffraction and X-ray fluorescence analysis were indications of the formation of calcium-deficient (Ca/P = 1.65) hydroxyapatite (HA) with a small carbonate content under influence of microwave irradiation. The investigated mechanical properties (maximal relative deformation, compressive strength and Young's modulus) of SWCNT loaded HA-alginate composites confirm their dependence on SWCNTs content. The compressive strength of HA-alginate-SWCNT and the HA-alginate control (202 and 159 MPa, respectively) lies within the values characteristic for the cortical bone. The addition of 0.5% SWCNT, in relation to the content of HA, increases the Young's modulus of the HA-alginate-SWCNT (645 MPa) compared to the SWCNT-free HA-alginate sample (563 MPa), and enhances the material shape stability in simulated physiological conditions. Structural modeling of HA-alginate-SWCNT system showed, that physical adsorption of SWCNT into HA-alginate occurs by forming triple complexes stabilized by solvophobic/van der Waals interactions and H-bonds. The high-performance liquid chromatography demonstrated the influence of SWCNTs on the sustained anaesthesinum drug (used as a model drug) release (456 h against 408 h for SWCNT-free sample). Cell culture assay confirmed biocompatibility and stimulation of osteoblast proliferation of 0.05% and 0.5% SWCNT-containing composites during a 3-day cultivation. All these facts may suggest the potential possibility of using the SWCNT-containing materials, based on HA and alginate, for bone tissue engineering.

A Novel Nanoconjugate of Landomycin A with C60 Fullerene for Cancer Targeted Therapy: In Vitro Studies.

INTRODUCTION: Landomycins are a subgroup of angucycline antibiotics that are produced by Streptomyces bacteria and possess strong antineoplastic potential. Literature data suggest that enhancement of the therapeutic activity of this drug may be achieved by means of creating specific drug delivery systems. Here we propose to adopt C60 fullerene as flexible and stable nanocarrier for landomycin delivery into tumor cells. METHODS: The methods of molecular modelling, dynamic light scattering and Fourier transform infrared spectroscopy were used to study the assembly of C60 fullerene and the anticancer drug Landomycin A (LA) in aqueous solution. Cytotoxic activity of this nanocomplex was studied in vitro towards two cancer cell lines in comparison to human mesenchymal stem cells (hMSCs) using 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) test and a live/dead assay. The morphology of the cells incubated with fullerene-drug nanoparticles and their uptake into target cells were studied by scanning electron microscopy and fluorescence light microscopy. RESULTS: The viability of primary cells (hMSCs, as a model for healthy cells) and cancer cell lines (human osteosarcoma cells, MG-63, and mouse mammary cells, 4T1, as models for cancer cells) was studied after incubation with water-soluble C60 fullerenes, LA and the mixture C60 + LA. The C60 + LA nanocomplex in contrast to LA alone showed higher toxicity towards cancer cells and lower toxicity towards normal cells, whereas the water-soluble C60 fullerenes at the same concentration were not toxic for the cells. CONCLUSIONS: The obtained physico-chemical data indicate a complexation between the two compounds, leading to the formation of a C60 + LA nanocomposite. It was concluded that immobilization of LA on C60 fullerene enhances selectivity of action of this anticancer drug in vitro, indicating on possibility of further preclinical studies of novel C60 + LA nanocomposites on animal tumor models.

C60 fullerene loaded hydroxyapatite-chitosan beads as a promising system for prolonged drug release.

Bioactive composite material in the form of beads, based on natural polysaccharide Chitosan (CS), hydroxyapatite (HA), and C60 fullerene (C60), was synthesized under influence of microwave irradiation (MW). The bead is a 3D matrix consisting of CS macromolecules cross-linked with sodium tripolyphosphate with HA and C60 particles immobilized in its structure. XRD and FTIR data confirmed the formation of calcium deficient carbonate substituted HA with needle-shaped nanocrystallites of about 80 nm. MW does not influence CS structure. C60 enhances the beads shape stability. HPLC studies suggest the (up to 18 days) prolonged release of hydrophobic Anaesthesinum from C60-containing composites. Changing the C60 content allows adjusting the drug release time. The presence of CS and C60 provides the Anaestesinum release profile, according to the calculated correlation coefficient (r = 0.99), close to the zero order kinetic release profile. The inhibition zone test (ZOI) shows the antimicrobial activity of the composites containing 0.004 mg/ml C60 against S. aureus ATCC 25923 and E. coli ATCC 25922 (ZOI 16 ±â€¯1 and 10 ±â€¯2 mm, respectively). Cell viability test indicates no toxicity of 0.3 and 0.15 mg/ml C60-containing composites.

Antitumor effects and hematotoxicity of C60-Cis-Pt nanocomplex in mice with Lewis lung carcinoma.

BACKGROUND: Cisplatin (Cis-Pt) is a widely used anticancer drug but its therapeutic efficiency is limited by hemato-, cardio-, hepato-, nephro- and neurotoxicity. Complexation of Cis-Pt with C60 fullerene nanoparticle will allow to enhance the antitumor activity of the drug and to reduce its side toxic effects. AIM: To estimate the antitumor effects of С60-Cis-Pt nanocomplex in Lewis lung carcinoma (LLC) and analyze hematological toxicity in tumor-bearing mice. MATERIALS AND METHODS: Complexation of C60 fullerene and Cis-Pt molecule was studied by computer simulation. С60-Cis-Pt nanocomplex was i.p. injected to LLC-bearing mice in a total dose of 7.5 mg/kg (C60:Cis-Pt as 3.75:3.75 mg/kg). The survival of tumor-bearing mice and the relative reduction of tumor weight was recorded. Blood indices were determined using the Particle Counter PCE-210 automatic hematology analyzer. RESULTS: Computer simulation demonstrated the formation of С60-Cis-Pt nanocomplex in physiological medium and its stability due to the hydrophobic interactions. Treatment with C60-Cis-Pt nanocomplex increased survival time of LLC-bearing mice by 32%, normalized hemoglobin content (up to 100 g/l), erythrocyte and platelet count as compared to the untreated LLC-bearing mice. Tumor weight decreased by 35.5%; the mitotic index of tumor cells decreased by 78%, and apoptotic index increased by 75%. The revealed effects of the C60-Cis-Pt nanocomplex were more pronounced than the effects of Cis-Pt or C60 fullerene alone in equivalent dose. CONCLUSION: Treatment with C60-Cis-Pt nanocomplex prolonged the survival of LLC-bearing mice and reduced anemia in LLC-bearing mice.

Reconciling the controversial data on the effects of C60 fullerene at the organismal and molecular levels using as a model Drosophila melanogaster.

Fullerenes have attracted attention since their discovery as structural units of complex carbon nanostructures capable of transporting drugs and macromolecules. As such artificial nanomaterials are applied in biology and medicine, they are routinely scrutinized for their effects on living organisms. The results of such studies range from direct destabilizing effects on DNA molecules to amelioration of the toxic effects of known genotoxic agents. We tested the effect of buckminsterfullerene (C60) on Drosophila melanogaster at DNA, tissue and organism levels. The water-soluble pristine C60 fullerene at the concentration of 20 µg/ml and 40 µg/ml leads to the activation of the mus209 gene in D. melanogaster larvae salivary glands, which can indicate higher levels of DNA damage. However, the absence of effects at the cell and organismal level could be explained by the activation of repair systems or by active elimination of damaged cells.

Effect of С60 fullerenes on the intensity of colon damage and hematological signs of ulcerative colitis in rats.

The application of pristine С60 fullerene aqueous colloid solution (C60FAS; 0.5 mg/kg body weight) for rats experienced acute colitis, either intraperitoneally or intrarectally (1) restores the colonic mucosa healing and epithelial barrier integrity, evidenced by autopsies and histological findings and the normalization of phenolsufonphthalein dye daily excretion; (2) attenuates the consequences of hemorrhages, such as signs of anemia and increased platelet count; (3) improves the liver redox status suppressing the lipid peroxidation (malonic dialdehyde and protein carbonyl group levels tended down) and stimulating the antioxidant defense system (glutathione peroxidase activity grew up). In addition, C60FAS intrarectally increases the monocyte count and decreases content of neutrophil granulocytes, i.e. diminishes the rate of the inflammatory process and activates the processes of colon tissues restoring with monocytes involvement. Therefore, intrarectal administration of C60FAS may serve as an efficient tool of ulcerative colitis therapy.

Composite material based on hydroxyapatite and multi-walled carbon nanotubes filled by iron: Preparation, properties and drug release ability.

The novel bioactive composite material based on hydroxyapatite and multi-walled carbon nanotubes filled by iron was synthesized by the "wet chemistry" method and characterized in detail by various experimental techniques including the X-ray diffraction, Fourier transform infrared and energy-dispersive X-ray fluorescence spectroscopy, thermogravimetric and differential thermal analysis. The swelling behaviour was quantified by measuring the changes in sample weight as a function of sample immersion time in a phosphate buffered saline (PBS). Bioactivity test was carried out by soaking the samples in PBS. The material composition influence on the model drug release was studied using the high-performance liquid chromatography method. Finally, the mechanical properties (maximal relative deformation, strength and Young's modulus) of the samples under loading were investigated too. The findings clear demonstrate the possibility of application of the created composite material in bioengineering of bone tissue to fill bone defects of various geometries with the function of prolonged release of the drug. It is assumed that this composite material can be used in 3D modeling of areas of bone tissue that have to bear a mechanical load.

C60 Fullerene Effects on Diphenyl-N-(trichloroacetyl)-amidophosphate Interaction with DNA In Silico and Its Cytotoxic Activity Against Human Leukemic Cell Line In Vitro.

New representative of carbacylamidophosphates - diphenyl-N-(trichloroacetyl)-amidophosphate (HL), which contains two phenoxy substituents near the phosphoryl group, was synthesized, identified by elemental analysis and IR and NMR spectroscopy, and tested as a cytotoxic agent itself and in combination with C60 fullerene.According to molecular simulation results, C60 fullerene and HL could interact with DNA and form a rigid complex stabilized by stacking interactions of HL phenyl groups with C60 fullerene and DNA G nucleotide, as well as by interactions of HL CCl3 group by ion-π bonds with C60 molecule and by electrostatic bonds with DNA G nucleotide.With the use of MTT test, the cytotoxic activity of HL against human leukemic CCRF-CM cells with IC50 value detected at 10 µM concentration at 72 h of cells treatment was shown. Under combined action of 16 µM C60 fullerene and HL, the value of IC50 was detected at lower 5 µM HL concentration and at earlier 48 h period of incubation, besides the cytotoxic effect of HL was observed at a low 2.5 µM concentration at which HL by itself had no influence on cell viability. Binding of C60 fullerene and HL with minor DNA groove with formation of a stable complex is assumed to be one of the possible reasons of their synergistic inhibition of CCRF-CЕM cells proliferation.Application of C60 fullerene in combination with 2.5 µM HL was shown to have no harmful effect on structural stability of blood erythrocytes membrane. Thus, combined action of C60 fullerene and HL in a low concentration potentiated HL cytotoxic effect against human leukemic cells and was not followed by hemolytic effect.

C60 Fullerene as Promising Therapeutic Agent for the Prevention and Correction of Skeletal Muscle Functioning at Ischemic Injury.

The therapeutic effect of pristine C60 fullerene aqueous colloid solution (C60FAS) on the functioning of the rat soleus muscle at ischemic injury depending on the time of the general pathogenesis of muscular system and method of administration C60FAS in vivo was investigated. It was found that intravenous administration of C60FAS is the optimal for correction of speed macroparameters of contraction for ischemic muscle damage. At the same time, intramuscular administration of C60FAS shows pronounced protective effect in movements associated with the generation of maximum force responses or prolonged contractions, which increase the muscle fatigue level. Analysis of content concentration of creatine phosphokinase and lactate dehydrogenase enzymes in the blood of experimental animals indicates directly that C60FAS may be a promising therapeutic agent for the prevention and correction of ischemic-damaged skeletal muscle function.

Structure and Thermal Stability of Co- and Fe - Intercalated Double Silicene Layers.

The arrangement of Fe and Co atoms between two silicene planes was theoretically investigated. The research has shown that below 500 K Co atoms form stable lattices-"hexagonal" (with the lattice parameters 0.635 nm of AB configuration) and cubic (with the lattice parameter of 0.244 nm), whereas Fe atoms form cubic lattices only (with the lattice parameter of 0.281 nm). The system intercalated with Co atoms is stable enough at high temperatures up to ~625 K, while the Fe-silicene system is stable only at ~770 K. The silicene UV-spectrum depending on the intercalate concentration and association constant of the "silicene-intercalate" system was calculated.

Cytotoxic Effects of Dimorfolido-N-Trichloroacetylphosphorylamide and Dimorfolido-N-Benzoylphosphorylamide in Combination with C60 Fullerene on Leukemic Cells and Docking Study of Their Interaction with DNA.

Dimorfolido-N-trichloroacetylphosphorylamide (HL1) and dimorfolido-N-benzoylphosphorylamide (HL2) as representatives of carbacylamidophosphates were synthesized and identified by the methods of IR, 1H, and 31P NMR spectroscopy. In vitro HL1 and HL2 at 1 mM concentration caused cell specific and time-dependent decrease of leukemic cell viability. Compounds caused the similar gradual decrease of Jurkat cells viability at 72 h (by 35%). HL1 had earlier and more profound toxic effect as compared to HL2 regardless on leukemic cell line. Viability of Molt-16 and CCRF-CEM cells under the action of HL1 was decreased at 24 h (by 32 and 45%, respectively) with no substantial further reducing up to 72 h. Toxic effect of HL2 was detected only at 72 h of incubation of Jurkat and Molt-16 cells (cell viability was decreased by 40 and 45%, respectively).It was shown that C60 fullerene enhanced the toxic effect of HL2 on leukemic cells. Viability of Jurkat and CCRF-CEM cells at combined action of C60 fullerene and HL2 was decreased at 72 h (by 20 and 24%, respectively) in comparison with the effect of HL2 taken separately.In silico study showed that HL1 and HL2 can interact with DNA and form complexes with DNA both separately and in combination with C60 fullerene. More stable complexes are formed when DNA interacts with HL1 or C60 + HL2 structure. Strong stacking interactions can be formed between HL2 and C60 fullerene. Differences in the types of identified bonds and ways of binding can determine distinction in cytotoxic effects of studied compounds.

Comparative Analysis of the Antineoplastic Activity of C60 Fullerene with 5-Fluorouracil and Pyrrole Derivative In Vivo.

The antitumor activity of pristine C60 fullerene aqueous solution (C60FAS) compared to 5-fluorouracil (5-FU) and pyrrole derivative 1-(4-Cl-benzyl)-3-Cl-4-(CF3-fenylamino)-1H-pyrrol-2.5-dione (MI-1) cytostatic drugs was investigated and analyzed in detail using the model of colorectal cancer induced by 1.2-dimethylhydrazine (DMH) in rats. The number, size, and location of the tumors were measured, and the pathology was examined. It was found that the number of tumors and total lesion area decreased significantly under the action of C60FAS and MI-1. Because these drugs have different mechanisms of action, their simultaneous administration can potentially increase the effectiveness and significantly reduce the side effects of antitumor therapy.

[MUSCLE FATIGUE: FACTORS OF DEVELOPMENT AND WAYS OF CORRECTION].

The data regarding the analysis of the physiological and biochemical mechanisms of muscle fatigue and ways to prevent it are summarized. The effect of the most common endogenous and exogenous antioxidants in the biochemical processes in muscle fatigue was analyzed. It is shown that biocompatible, non-toxic water-soluble C(60) fullerenes, which possess powerful antioxidative properties, promise great prospects in the correction of skeletal muscle fatigue caused by the destructive action of free radicals.

Thermodynamic Complexing of Monocyclopentadienylferrum (II) Intercalates with Double-Walled Carbon Nanotubes.

By employing the methods of molecular mechanics, semi-empirical quantum-chemical РМ3 and Monte-Carlo, the positioning of monocyclopentadienylferrum (II) molecules in double-walled (5,5)@(10,10) carbon nanotubes (CNT) depending on their concentration and temperature has been studied. The molecules have been found out to form stable bonds with CNT walls, with a tendency between intercalate stability and the CNT structure. The temperature growth (over ~500 K) causes gradual bond ruining followed by extrusion of interwall intercalate. Further temperature increase up to 600-700 K is characterised with intercalate external surface desorption, stabilising the whole system and keeping the interwall intercalate only. The CNT's UV-spectrum (5,5)@(10,10) depending on the intercalate concentration and association constant of the "double-walled CNT-intercalate" system have been calculated. A combination of unique optical, electrical and magnetic behaviour of cyclopentadienyl complexes with their ability to form high-stable intercalate with CNT opens a prospect of their applying in nanotechnology.

Interaction of C60 fullerene complexed to doxorubicin with model bilipid membranes and its uptake by HeLa cells.

With an aim to elucidate the effects of C60 fullerene complexed with antibiotic doxorubicin (Dox) on model bilipid membranes (BLM), the investigation of the electrical properties of BLM under the action of Dox and C60 fullerene, and of their complex, C60+Dox,was performed. The complex as well as its components exert a clearly detectable influence on BLM, which is concentration-dependent and also depends on phospholipid composition. The mechanism of this effect originates either from intermolecular interaction of the drug with fatty-acid residues of phospholipids, or from membranotropic effects of the drug-induced lipid peroxidation, or from the sum of these two effects. By fluorescence microscopy the entering of C60 + Dox complex into HeLa cells was directly shown.

PROSPECTS OF C60 FULLERENE APPLICATION AS A MEAN OF PREVENTION AND CORRECTION OF ISCHEMIC-REPERFUSION INJURY IN THE SKELETAL MUSCLE TISSUE.

lack of accurate diagnostic tests for the rapid detection of ischemic injury remains an urgent problem. In this context, a search for specific markers of ischemia as well as new therapeutic agents for prevention and treatment of ischemic injury continues. Water-soluble nano-sized C60 fullerenes, as powerful antioxidants, can act as promising means for correction of various muscle system states, in the base of which lie the destructive effects of free radicals.

Structural self-organization of C60 and cisplatin in physiological solution.

The specific features of structural self-organization of C60 fullerene and antitumor drug cisplatin (Cis) in physiological solution (0.9% NaCl) have been investigated by means of small-angle neutron scattering, scanning electron and atomic force microscopies, as well as isothermal titration calorimetry, dynamic light scattering and UV-Vis spectroscopy. The formation of C60 + Cis complexes, has been reported, unveiling the mechanism of medico-biological synergy observed during administration of the mixture of these drugs.

Impact of C 60 fullerene on the dynamics of force-speed changes in soleus muscle of rat at ischemia-reperfusion injury.

The effect of C60 fullerene nanoparticles (30-90 nm) on dynamics of force response development to stimulated soleus muscle of rat with ischemic pathology, existing in muscle during the first 5 hours and first 5 days after 2 hours of ischemia and further reperfusion, was investigated using the tensometric method. It was found that intravenous and intramuscular administration of C60 fullerene with a single dose of 1 mg/kg exert different therapeutic effects dependent on the investigated macroparameters of muscle contraction. The intravenous drug administration was shown to be the most optimal for correction of the velocity macroparameters of contraction due to muscle tissue ischemic damage. In contrast, the intramuscular administration displays protective action with respect to motions associated with generation of maximal force response or continuous contractions elevating the level of muscle fatigue. Hence, C60 fullerene, being a strong antioxidant, may be considered as a promising agent for effective therapy of pathological states of the muscle system caused by pathological action of free radical processes.

Application of C60 Fullerene-Doxorubicin Complex for Tumor Cell Treatment In Vitro and In Vivo.

Development of nanocarriers for effective drug delivery to molecular targets in tumor cells is a real problem in modern pharmaceutical chemistry. In the present work we used pristine C60 fullerene as a platform for delivery of anticancer drug doxorubicin (Dox) to its biological targets. The formation of a complex of C60 fullerene with Dox (C60 + Dox) is described and physico-chemical characteristics of such complex are presented. It was found that Dox conjugation with C60 fullerene leads to 1.5-2-fold increase in Dox toxicity towards various human tumor cell lines, compared with such effect when the drug is used alone. Cytotoxic activity of C60 + Dox complex is accompanied by an increased level of cell produced hydrogen peroxide at early time point (3 h) after its addition to cultured cells. At the same time, cellular production of superoxide radicals does not change in comparison with the effect of Dox alone. Cytomorphological studies have demonstrated that C60 + Dox complexes kill tumor cells by apoptosis induction. The results of in vivo experiments using Lewis lung carcinoma in mice confirmed the enhancement of the Dox toxicity towards tumor cells after drug complexation with C60 fullerene. The effect of such complex towards tumor-bearing mice was even more pronounced than that in the in vitro experiment with targeting human tumor cells. The tumor volume decreased by 2.5 times compared with the control, and an average life span of treated animals increased by 63% compared with control. The obtained results suggest a great perspective of application of C60 + Dox complexes for chemotherapy of malignant tumors.

[Application of the methods of molecular modeling to the search for new biologically active substances].

The searching for new chemical compounds possessing specific biological activity is a complex problem that needs the usage of modern methods of molecular modeling. In particular, for the prupose of searching for potentially active compounds for whole class of SH2 domains, a comparison of all available structures, their cluster analysis, molecu- lar docking, selection of all possible pharmacophore models and GTM prediction were done. Obtained results testify to the considerable variability of binding of SH2 domains.