Abstract
The aim of the work is synthesis and study on the properties of polyfunctional magnetosensitive nanocomposites (NC) and target-directed magnetic fluids (MF) based on physiological solution (PS), magnetite, gemcitabine (GEM) and HER2 antibodies (AB), promising for use in targeted antitumor therapy against MDA-MB-231 aggressive tumor cells of triple-negative human breast cancer (BC) with high proliferative and metastatic activity. The specific surface area (Ssp) of samples was determined by the method of thermal desorption of nitrogen using a device KELVIN 1042 of “COSTECH Instruments”. The size of nanoparticles (NP) has been estimated by the formula DBET = 6/(ρSBET), where ρ is the density of NC particle, SBET is the value of the specific surface area calculated by the polymolecular adsorption theory of Brunauer, Emmett and Teller (BET). The surface condition of nanodispersed samples was studied by IR spectroscopy (“Perkin Elmer” Fourier spectrometer, a model 1720X). To calculate the concentration of hydroxyl groups on the surface of nanostructures, the method of differential thermal analysis was used in combination with differential thermogravimetric analysis. The thermograms were recorded using a derivatograph Q-1500D of MOM firm (Hungary) in the temperature range of 20–1000 °C at a heating rate of 10 deg/min. X-ray phase analysis of nanostructures was performed using a diffractometer DRON-4-07 (CuKα radiation with a nickel filter in a reflected beam, the Bragg-Brentano focusing). The size and shape of NP were determined by electron microscopy (a transmission electron microscope (TEM) JEM-2100F (Japan)). The hysteresis loops of the magnetic moment of the samples were measured using a laboratory vibration magnetometer of Foner type at room temperature. Measurement of optical density, absorption spectra and GEM concentration in solutions was performed by spectrophotometric analysis (Spectrometer Lambda 35 UV/Vis Perkin Elmer Instruments). The amount of adsorbed substance on the surface of magnetite was determined using a spectrophotometer at λ = 268 nm from a calibration graph. The thickness of the adsorbed layer of GEM in the composition of Fe3O4@GEM NC was determined by magnetic granulometry. To study the direct cytotoxic/cytostatic effect of a series of experimental samples of MF based on PS, Fe3O4 NP, GEM, HER2 AB, as well as MF components in mono- or complex use, onto MDA-MB-231 cells in vitro, IC50 index was determined. MF were synthesized on the basis of single-domain Fe3O4 and PS, stabilized with sodium oleate (Ol.Na) and polyethylene glycol (PEG), containing GEM and HER2 (Fe3O4@GEM/Ol.Na/PEG/HER2+PS). The cytotoxic/cytostatic activity of MF against MDA-MB-231 cells was studied. It was found that as a result of application of synthesized MF composed of Fe3O4@GEM/Ol.Na/PEG/HER2+PS at the concentration of magnetite of 0.05 mg/mL, GEM - 0.004 mg/mL and HER2 AB - 0.013 μg/mL, a synergistic effect arose, with reduction of the amount of viable BC cells to 51 %. It has been proved that when using MF based on targeted Fe3O4/GEM/HER2 complex, the increased antitumor efficacy is observed compared to traditional use of the drug GEM, with a significant reduction (by four times) of its dose. The high cytotoxic/cytostatic activity of Fe3O4/GEM/HER2 complexes is explained by the fact that endogenous iron metabolism disorders play a significant role in the mechanisms of realization of the apoptotic program under the influence of nanocomposite. Thus, when the nanocomposite system contains Fe3O4/GEM/HER2 complexes in MDA-MB-231 cells, a significant increase is observed in the level of “free iron”, which favours formation of reactive oxygen species and causes oxidative stress (Fenton reaction). The consequences of oxidative stress are induction of apoptosis, enhancement of lipid peroxidation processes, as well as structural and functional rearrangement of biological membranes. The prospects have been shown of further studies of Fe3O4@GEM/Ol.Na/PEG/HER2+PS MF in order to create on their basis a magnetically carried remedy for use in targeted antitumor therapy.
Highlights
One of the main problems of modern antitumor therapy with gemcitabine (GEM) is its toxic effects on the human body and the occurrence of severe adverse reactions in patients [1,2,3,4]
It was found that as a result of application of synthesized magnetic fluids (MF) composed of Fe3O4@GEM/Ol.Na/polyethylene glycol (PEG)/HER2+physiological solution (PS) at the concentration of magnetite of 0.05 mg/mL, GEM - 0.004 mg/mL and HER2 AB - 0.013 μg/mL, a synergistic effect arose, with reduction of the amount of viable breast cancer (BC) cells to 51 %
It has been proved that when using MF based on targeted Fe3O4/GEM/HER2 complex, the increased antitumor efficacy is observed compared to traditional use of the drug GEM, with a significant reduction of its dose
Summary
One of the main problems of modern antitumor therapy with gemcitabine (GEM) is its toxic effects on the human body and the occurrence of severe adverse reactions in patients [1,2,3,4]. Among the variety of modern methods of targeted drug delivery and local therapy of diseases at the level of organs, cells and genes, the most developed at this time are methods using magnetically sensitive nanostructures conjugated with antitumor drugs of different mechanism of action. A concept has been substantiated of chemical construction of magnetosensitive nanocomposites (NC) with multilevel hierarchical nanoarchitecture, characterized by the functions of “nanoclinics” and medicobiological nanorobots [12,13,14,15,16,17]: recognition of microbiological objects in biological media; targeted delivery of drugs to specific cells and organs, and deposition; complex local chemo-, immuno-, neutron-capture, hyperthermic, photodynamic therapy and magnetic resonance imaging in real time, detoxification of the body by adsorption of cell decomposition residues, viral particles, heavy metal ions, etc. For the manufacture of magnetically sensitive multifunctional NC, a significant interest of researchers is drawn to nanostructures of core-shell type based on single-domain magnetite (Fe3O4), which are characterized by a unique set of physical, chemical and biological properties, the capability to create magnetic fluids (MF) on their basis, containing oncological remedies with various functional purpose and mechanisms of action [25, 26]
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