The aim of this work was to study antigen expression associated with cancer stem cells at human metastatic melanoma transplantable cell lines received in FSBSI «N.N. Blokhin RCRC». Expression of CD133, CD117, CD34, CD90, CD24, CD44 associated with cancer stem cells has been explored in immunofluorescence reaction with monoclonal antibodies. Isospecific antibodies which values of immunofluorescence didn't increase 0,5%-level of positive cells were used for negative control. The cancer stem cells antigen expression was heterogeneous. If we take 10% antigen-positive cells for discrimination level, we can see CD133-expression at 6 (28,5%) cell lines of 21, CD117 - at 6 (28,5%) cell lines of 21, CD90 - at 12( 57%) of 21 lines, CD34 - at 4(19%) of 21 cell lines, CD44 - at 17(81%) of 21 cell lines and CD24 - at 5 (23,8%) of 21 lines. This antigens expression was less than 10% (from 1 to 8.5%) at other cell lines. After cloning mel Ibr cell line that expressed on surface about 8.5% of CD133 in semi-solid agar, the clone of cells expressed 100% CD133⁺ cells was obtained. Thus, tumor cells expressing a small percentage of CSC-associated antigens are also CSC, and other antigen-negative cells are their descendants.

Previous studies have shown that aranosa-lio initiate cell death by activation of CD95-receptor of apoptosis. The mechanism of liposomal aranosa action is still unknown. Anticancer drugs may cause tumor cell death by the activation of autophagy. The purpose of this study was to determine the influence of liposomal aranosa on autophagy. The study was assessed on human melanoma cell lines mel Mtp, mel Ibr, mel Kor, mel Z and mel Mtp clone X. The cells were treated with aranosa-lio or liposomal aranosa at concentration 450 μg/ml for 24 hours. The expression of autophagy were analyzed the expression of mRNA Beclin 1. Expression of mRNA Beclin 1 was reduced on the majority of cell lines, which might be due to the fact that the liposomal form of the drug caused cell death by intrinsic apoptosis which is suppose to suppress autophagy.

CTA are locate in a group of tumor-associated proteins. In normal conditions their expression is limited to embryonic tissues and reproductive tissues of adults. In other tissues CTA are not exist. Interestingly, that the aberrant CTA overexpression is Observed during malignant transformation. In this work we investigated the mRNA of 20 CTA genes overexpression in biopsy and peripheral blood of 49 patients. Among them a MAGE-A1, MAGE-A2 MAGE-A3 MAGE-A4, MAGE-A5, MAGE-A6 mRNAs were detected in same time by using originally primers, that specifically recognize MAGEA-conservative sequences. Similar system of primers was constructed for GAGE1-8 verification. Furthermore, the expression of SSX1, SSX2, SSX4, XAGE1, NY-ESO1 and MAGEC1 was identified. It is found that the detection of mRNA CTA-coding genes can be used as a monitoring test. The presence of mRNA MAGE-C1 and XAGE1 genes in the blood of patients with colorectal cancer could be a potential marker of a more favorable course of the disease. In other hand, the existence of mRNA MAGE-A1-6, GAGE 1-8 and SSX1,2,4 genes may indicate a poor prognosis.

Glioblastoma multiforme resistance requires a new approach for glioma therapy. Protein p53 is one of the main cellular oncogene, overexpressed in 50% of brain tumor cases. Impact of p53 attenuation was evaluated in the presence of oncolytic adenovirus and temodar, which exhibit anti-glioma effect using in vitro glioma models. Using U87 human glioma cells we observed an additive effect between alkilated chemotherapeutic agent temodar and oncolytic adenovirus which results into p53 inhibition. It occurs that attenuation of p53 using PFTa inhibitor, significantly prolongs cell death type II - autophagy and, therefore improves effect mediated by autophagy induced agents. In conclusion, combination of PFTa and temodar might represent a powerful therapeutic combination which sensibilises glioma cells to the infection with oncolytic vector.

Effective production of pharmaceutical proteins using transgenic poultry may be achieved only from a construction of expression vector for chicken oviduct bioreactor. This study was focused on development of temporary expression system by using primary oviduct epithelial cells in which transfected gene expression can be studied. The present work was aimed to study three manners for preparation of primary oviduct cells, but only one was effective. Selective digestion of chicken oviduct tissue pieces in trypsin at 38° C and mechanical withdrawal of upper cell layer resulted in the isolation of purity primary culture. Experiments indicated that monolayer cultured for 2-3 d were fit to be passaged. Then after 1-2-d, the cells were transfected. Based on our result with primary culture, we did not used complicated enzyme treatment and extra equipment in order to increase the purity of epithelial cells. Liposomes «Lipofectine® 2000» were used for primary oviduct cells transfection of a plasmid designed based on the pIRES EGFP2 vector (Clontech, United States). The ratio of cells carrying GFP activity was 5-8% of the total number of cultured cells. This researches was focused on development in vitro of temporary expression system in which transfected gene expression can be tested for 4-5 d.

Immunoliposomal Mitoxantrone against CD5⁺ T-cell limphoproliferative diseases was developed. Liposomal model with lipid proportion PhCh: Chol: mPEG₂₀₀₀-DSPE: pNP-PEG₃₀₀₀-lipid 11: 9: 0,95: 0,05 was used for immu-noliposome development. We used gel-filtration to remove free Mitoxantrone and unconjugated monoclonal antibody from immunoloposomal construction. One liposome was conjugated with ~16 antibody molecules. Immunoliposomes linked with 80% of antigen+ cells. Immunoloposomal Mitoxantrone killed antigen+ target cells in MTT-assay.

The study evaluates physical, chemical and biological properties of glycosylated derivatives of pyropheophorbide-α and chlorine e₆, cycloimid derivatives of chlorine p₆. Glycosylated derivatives of pyropheophorbide α and chlorine e₆ were characterized by stability in solution without light exposure, but subject to photobleaching during exposure. Derivatives of pyropheophorbide-α showed high photo-induced activity in biological assay, the IC₅₀ value varied from 35±7nM to 82±7 nM (depending on the pendent group and the tumor cell culture). Among the derivatives of chlorine e₆ conjugate with galactose residue in A-pyrrole showed the maximum activity towards HEp2, A549 and HT29 cells (IC_{50 -} 15±8 nM, 70±19 nM and 34±16 nM respectively), its activity 8-10 times exceeding the unconjugated counterpart and the derivatives with different macrocycle modification. Cycloimid derivatives of chlorine p₆ are stable in solution without exposure and relatively stable under light exposure. These compounds showed high photo-induced activity in biological assay (IC₅₀ varied from 35±10 nM to 250±30 nM) significantly exceeding the activity of unmodified derivatives of chlorine p₆.

Efficacy of photodynamic therapy with intraperitoneal introduction photosensitizer Amidoaminchlorine on rat sarcoma M1 model was studied. The aim of this study was to determine minimum effective dose of photosensitizer and parameters of laser radiation for achievement of tumor full regress. Dynamics of photosensitizer accumulation in tumor and healthy tissues was studied to definite the time of photodynamic therapy. Antitumor efficacy different doses of photosensitizer and various parameters of laser radiation were investigated. As a result, optimal term of photodynamic therapy, photosensitizer minimum effective dose and parameters of laser irradiation were established.

The liposomal antitumor drugs cell death induction mechanism has been analyzed in this work. Liposomal dosage forms of doxorubicin, cisplatin and aranosa overcome drug resistance. However, the mechanism of overcoming drug resistance by this drugs is different. Liposomal doxorubicin overcomes multidrug resistance by liposomes binding with the P-glycoprotein in 185 position of glycine. Liposomal cisplatin overcomes monoresistance by external apoptosis gene activation. The liposomal aranosa does not use CD95/Fas signaling pathway of apoptosis. Thus, liposomal drug action mechanism depends on the type of cells, and on the antitumor drug, that is enclosed in a liposome and can be individualized in each case.