The use of cytokines as anticancer drugs is limited due to their toxicity. It is possible to reduce toxicity and to increase the therapeutic index by using cytokines in the form of conjugates with antibodies – immunocytokines. The results of preclinical studies have shown increased efficacy and reduced toxicity of various immunocytokines compared to the original, unconjugated cytokines. The potential of immunocytokines as anticancer agents is currently being studied in clinical trials. The therapeutic efficacy of immunocytokines depends on their physicochemical parameters, which determine the in vivo biodistribution, and biological activity as a result of the mechanisms of the antibody action and cytokine sites incorporated in the design. There is a need for methods that allow to assess the biological activity of both individual sites and the entire immunocytokine molecule when characterizing immunocytokines at an early stage of research.
This review considers the existing approaches for assessing the biological activity of immunocytokines in vitro in the course of preclinical studies, such as monolayer cultures, reporter cell lines, co-cultures, three-dimensional (3D) tumor models. Monolayer cultures are sufficient to confirm the mechanism of action of separate sites of immunocytokines used in the design, and the “gold standard” test systems for determining the specific biological activity of the cytokine and the effector functions of the antibody site remain in demand. Commercial reporter cell lines remain an alternative option for assessing the biological activity of cytokine and antibody sites at the level of activation of signaling pathways. Co-cultures of tumor and effector cells make it possible to evaluate the cytotoxic and immunomodulatory effects of antibody and cytokine sites without using 3D cultivation methods. The use of 3D tumor models makes it possible to replace several tests for the biological activity of separate sites of immunocytokines conducted on monolayer cultures and co-cultures with one comprehensive study, however, such models require significant time and material costs.

The review analyzes some parameters of CBA mice-males as model of spontaneous carcinogenesis characterizing adhesive and adaptive disorders. A weakening of the hepatocytes mutual adhesiveness force was noted already in early ontogenesis (5–10 days of postnatal development). This violation persisted and enhanced during hepatocarcinogenesis. A decrease of the β2 leukocyte integrins LFA-1 and Mac-1 expression on peripheral blood cells as well as an increase of the interleukins 6 and 10 in blood serum were determined during ontogenesis. It is significant for weakening the liver cells contact interactions (mutual adhesiveness) as well as immunity effectors and tumor cells interactions. A disbalance of the adaptive reactions and life quality important components was revealed in the CBA mice-males ontogenesis. Number of dopaminergic neurons and the neurogenesislevel in CBA micemales were decreasing. This does not contradict the dynamics of chronic stress and the aging process: an increase in the catabolic stress hormone corticosterone, a decrease in the anabolic hormone testosterone in the blood serum, a decrease in motor activity, signs of cachexia and alopecia, as well as a violation of immunological parameters.
CBA mice-males with an assessment of parameters characterizing adhesive and adaptive disorders during spontaneous carcinogenesis (the hepatocytes mutual adhesiveness forсe, the expression of β2 leukocyte integrins LFA-1 and Mac-1 on peripheral blood cells, the content of interleukins 6, 10, corticosterone and testosterone in blood serum, the number of dopaminergic neurons in the midbrain during ontogenesis) as well as the frequency and size of tumours, lifespan and somatic status of animals can be used as a scientifically- and evidence-based test system to study cytostatic drugs as well as non-toxic geroprotective medications for prevention and treatment of cancer in individuals with an increased risk of malignant neoplasms developing especially hepatocellular carcinoma.

Background. CD437, an analog of vitamin A, is an agonist of the retinoic acid γ-receptor (RARγ). CD437 is also known to cause p53-independent DNA damage by a mechanism independent of the RAR-mediated pathway. In cancer patients, iron deficiency is constantly detect, the delivery of iron to tissues is also destroyed.
Aim. To study the effect of CD437 on iron metabolism in metastatic melanoma cells, Mel Z.
Materials and methods. In this study 2D cultivation of metastatic Mel Z melanoma cells, phase-contrast and fluorescence microscopy, flow cytofluorimetry were used.
Results. In control cells without the addition of CD437 CD71, transferrin receptor, expressed 40 ± 4 % (p <0.05) of Mel Z cells, in the presence of CD437 CD71 expression increased to 80 ± 6 %. Next, we have studied the expression of ferritin. Iron, which is not involved in cell metabolism, is bound by ferritin. In control experiments, ferritin was expressed by 84 ± 6 % (p <0.05) of cells. When the cells grew in the presence of CD437, ferritin was expressed by all the cells (100 %, p <0.05). Such a scenario indicates that CD437 may contribute to the accumulation of free, unbound iron in the cell, which can induce ferroptosis. In control experiments without the addition of CD437, the level of membranes lipid peroxidation, an indicator of ferroptosis, was insignificant. Lipid peroxidation induced by CD437 was 55 ± 5 % (p <0.05) of the fluorescence intensity induced by erastin, positive control.
Conclusion. CD437 increases the iron uptake by metastatic melanoma cells. The low level of membranes lipid peroxidation induced by CD437 does not allow it to be considered as an inducer of ferroptosis. Additional investigations are needed to find iron-binding targets alternative to ferritin.

Background. Zinc alloys have advantages for use as biodegradable implantable orthopedic metal structures due to the absence of gas formation in comparison with magnesium alloys. But their mechanical properties are often has lower values.
Aim. Investigation of effect of high-pressure torsion (HPT) on strength, ductility, corrosion resistance, antimicrobial properties, surface cell colonization and biocompatibility of Zn-based alloys.
Materials and methods. The alloys of the Zn-x%Mg system (where x = 0; 1 and 1.7 %) in the initial undeformed state and after HPT were investigated in this work. Mechanical properties were studied on an Instron 3382 testing machine at room temperature. The biocompatibility of the alloys was evaluated by hemolytic activity and cytotoxicity assesment. We also studied the stimulation of colonization of the surface of the samples by mesenchymal multipotent stromal cells, as well as the presence of antimicrobial properties relative to the Escherichia coli culture. To study the degradation rate, the alloy samples were incubated in a standard nutrient medium for 8 days, assessing the change in their mass relative to the initial value.
Results. It has been established that HPT leads to an increase in the strength of pure Zn 2 times, and of Zn-1%Mg and Zn-1.7%Mg alloys by 3 and 5.5 times, respectively, with an increase in their ductility. At the same time, deformation treatment has practically no effect on the corrosion resistance of the initial materials. No significant increase in the hemolytic activity and bactericidal activity of the alloys was revealed during studies. However, a significant decrease in the ability of cells to colonize the surface of pure zinc was observed after HPT.
Conclusion. HPT leads to a significant increase in the strength and ductility of studied materials. At the same time, a decrease in the biocompatibility of zinc-based alloys after HPT did not observed. It was found that the discovered cytotoxic effect was obviously caused not so much by the alloy processing method as by its chemical composition. This makes it possible to evaluate the studied alloys of the Zn-x%Mg system treated by HPT (and, in particular, the Zn-1.7%Mg alloy) as a promising structure for the development of biodegradable orthopedic products.

Background. Expression evaluation of somatostatin receptors (SSTRs) in tumor cells is necessary for the reasonable use of therapy directed at such receptors.
Aim. The affinity determination of the original analogue of somatostatin cyphetrylin for SSTRs of transplanted mice mammary adenocarcinoma Ca-755.
Materials and methods. Cyphetrylin was synthesized in the Chemical Synthesis Laboratory of the Research Institute of Experimental Diagnostics and Therapy of Tumors, N.N. Blokhin National Medical Research Center of Oncology of the Ministry of Health of Russia. Cyphetrylin in tablet form was administered orally at a therapeutic dose of 10 mg/kg for 7 days to female F1 (C57Bl/6 × DBA/2) tumor-grafted Ca-755 mice. Animals of the control group were not administreted with сyphetrylin. Tumor tissue samples were obtained from animals on the 9th and 14th days after Ca-755 transplantation and sent for immunohistochemical study, which was performed on serial paraffin sections by the immunoperoxidase method using primary antibodies to various types of SSTRs.
Results. The high frequency of positive expression of SSTR1, SSTR2, and SSTR5 (in 80, 100 and 100 % of tumor samples, respectively) was shown in tumor samples of the control group animals. As a result of cyphetrylin introduction in tumor samples obtained on the 9th day after Ca-755 inoculation, a change in the tumor receptor status was found towards a decrease in the level of expression of SSTR2 (80 % of samples) and SSTR5 (80 % of samples); SSTR1 expression did not change (80 % of samples). Compared to the controlled, in tumor samples after cyphetrylin administration, obtained on day 14 from Ca-755 transplantation, a decrease in the expression level of SSTR2 (80 % of samples), SSTR1 and SSTR5 (60 % of samples for SSTR each type) was noted, due to cyphetrylin binding to SSTRs of tumor cells. The receptors SSTR3 and SSTR4 did not show a high level of expression in the studied Ca-755 tumor samples.
Immunohistochemical staining of Ca-755 cells with antibodies to SSTRs showed a tendency to reduction of antigen-positive cells number from 15–50 % in control to 10–40 % on day 9 after Ca-755 transplantation and 10–30 % on day 14 after Ca-755 transplantation.
Conclusion. The data obtained indicates the presence in mice transplanted mammary adenocarcinoma Ca-755 of SSTR1, SSTR2 and SSTR5 high level expression due to the binding to which the direct cyphetrylin antitumor effect is realized.

Background. A glycoside derivative of indolocarbazole LHS-1269, one of the new drugs selectively affecting tumors, which was first synthesized at the N.N. Blokhin National Medical-Research Center of Oncology of the Ministry of Health of Russia, is of particular scientific interest. Experimental studies demonstrated a multi-target mechanism of action of this compound. LHS-1269 interacts with several intracellular targets and induces various pathways of cell death. Several innovative models of the dosage forms were designed to achieve the highest antitumor activity of the compound and to perform further preclinical studies.
Aim. To develop the methods for the quantitative determination of LHS-1269 in pharmaceutical compositions proposed as a result of the search for the optimal dosage form.
Materials and methods. The study analyzed the spectrophotometric characteristics of LHS-1269 solutions in dimethylformamide, dimethylsulfoxide (DMSO) and in the mixture of solvents DMSO–ethyl alcohol, as well as electronic absorption spectra of the excipients in the mixture of solvents DMSO–ethyl alcohol. Spectrophotometric measurements were performed on a Cary 100 spectrophotometer (Varian, Inc., Australia) in the wavelength range from 200 to 500 nm. The standard sample is the substance LHS-1269 (N.N.  Blokhin Oncology Research Center of the Ministry of Health of Russia).
Results. The carried out studies showed that LHS-1269 solutions in dimethylformamide, DMSO and mixture of DMSO– ethyl alcohol are suitable for spectrophotometric measurements. Several variants of the methodology for the assay of LHS-1269 in various dosage form models that differ in the content of the active substance and the excipients composition have been developed: LHS-1269 concentrate for solution for injection and infusion; lyophilisate for solution for injection; liposomal lyophilisate for dispersion for injection.
Conclusion. Techniques for the assay of LHS-1269 in dosage form models have been developed. It has been shown that the developed techniques are applicable for LHS-1269 quantitative determination in innovative dosage forms containing polymeric low molecular weightsolubilizers, lipids, cholesterol, mono- or oligosaccharides as excipients.

Background. Hydrogels are promising for use in tissue engineering for the restoration and regeneration of various tissues, since they are able to perform the functions of bulk scaffolds, providing the formation of 3D cell structures. Population of such scaffolds with autologous or heterogeneous mesenchymal multipotent stromal cells in vitro makes it possible to localize these cells in the area of target tissues after implantation in a patient. One of the difficult tasks is the choice of the method and mode of sterilization of the hydrogel, which does not change its properties.
Aim. Study of the effectiveness of hydrogel sterilization by an accelerated electron beam in various modes, changes in the structure and biocompatibility of the scaffold, to assess the prospects for its use for medical purposes, including as a platform for mesenchymal stromal cells.
Materials and methods. We used a hydrogel based on 4 % solutions of sodium alginate and sodium salt of carboxymethyl cellulose, cross-linked with calcium chloride, which was developed, obtained and provided for our research by the team of the Research and Educational Center for Biomedical Engineering of the National University of Science and Technology “MISIS”. Hydrogel samples loaded with Escherichia coli, Lactobacillus acidophilus, Saccharomyces cerevisiae were subjected to electron beam treatment in the range of 5–100 kGy. After electron beam treatment of hydrogel, the presence of living microorganisms and its structure were evaluated by IR-Fourier spectroscopy, as well as the phenotype and formation of 3D structures by mesenchymal multipotent cells.
Results. It was found that the treatment of hydrogels with an electron beam at a mode of 25 kGy ensures the death of microorganisms, but does not destroy the structure of the hydrogel and does not inhibit the ability to form capillary-like structures by mesenchymal multipotent cells.
Conclusion. Treatment with an accelerated electron beam at a 25 kGy can be used to sterilize hydrogels to obtain bulk scaffolds for cell engineering implants.

Background. The effectiveness of cancer neoantigen peptide vaccines depends on the presence of an adjuvant in their composition. Poly(I:C), a TRL-3 agonist, is used as an adjuvant in mouse models of cancer vaccines, but has limitations for use in humans. Therefore, the search for new effective adjuvants for inclusion in the composition of cancer neoantigen peptide vaccine is relevant. Ridostin Pro is a domestic drug that contains a natural complex of sodium salts of double-chiral and single-chiral ribonucleic acids, is an agonist of TLR-3, an inducer of interferon, its antiviral activity is shown. In this regard, the study of Ridostin Pro as an adjuvant in the composition of neoantigen peptide vaccines is of interest.
Aim. To evaluate the ability of Ridostin Pro and Poly(I:C) adjuvants enhance the specific T-cell response to neoantigen synthetic peptides; to study the antitumor efficacy of a neoantigen peptide vaccine with Ridostin Pro or Poly(I:C) adjuvants.
Materials and methods. Immunogenicity of peptides after vaccination with Ridostin Pro or Poly(I:C) adjuvants evaluated with ELISpot. Antitumor effect of Ridostin Pro or Poly(I:C) adjuvants were evaluated on a mouse model of the B16-F10 tumor by the effect on the tumor growth rate and survival of mice.
Results. Vaccination of mice with Ridostin Pro or Poly(I:C) with neoantigen peptides contributed to the appearance of a specific immune response to peptides that were part of the vaccine. Ridostin Pro, both as part of a vaccine model and when administered without a peptide, inhibits tumor growth and increases the life expectancy of mice with melanoma B16-F10.
Conclusion. Ridostin Pro promotes the formation of a specific immune response to the peptide vaccine, enhances the antitumor effect of the vaccine. These results confirm that Ridostin Pro may prove to be an effective adjuvant for personalized neoantigen peptide vaccines.