Efficient prophylactic vaccines have been developed to prevent cervical cancer, a frequent female oncological disease caused by carcinogenic human papillomaviruses. World Health Organization worked out the program for prophylaxis of cervical cancer and control of the disease. It includes preventive adolescent vaccinations, screening of precancerous cervical lesions in women as well as cervical cancer treatment if originated. Cervical cancer diagnostics is being improved, development of therapeutic human papillomaviruses vaccines is in progress. The review deals with major achievements and certain challenges in the field of cervical cancer prevention.

Part I see: Volgareva G.M. Papillomaviral carcinogenesis. Major achievements and certain challenges. Part I. General notions of papillomaviruses. Human papillomavirusesassociated cancers. Russian Journal of Biotherapy 2020;19(1):6–12.

Part 2 see: Volgareva G.M. Papillomaviral carcinogenesis. Major achievements and certain challenges. Part 2. HPV-associated cancers in Russia. Preventive HPV vaccines. Russian Journal of Biotherapy 2020;19(2):31–8.

Phytoadaptogens (ginseng, eleutherococcus, rhodiola rosea, aralia, etc.) have a wide spectrum of biological activity, supporting the body in a state of nonspecific increased resistance to stress. The second part of the review provides information of the phytoadaptogens normalizing effects on the cardiovascular system. Analyzes current data on the antitumor properties of phytoadaptogens in relation to the effect on the onset and development of tumours, metastasis, recurrence, development of cytostatic disease, confirming the prospect of their using in drugs for prevention and biotherapy of tumour diseases.

Past 1 see: Phytoadaptogens in the tumours biotherapy and geriatrics (Past 1). Russian Journal of Biotherapy 2020;19(2):13–21.

Chewing gum medicines are solid dosage forms intended for chewing for a certain period of time without subsequent ingestion to provide local and systemic effects. This article describes the basic technologies for producing chewing gum medicinal products, analyzes and compares the technological aspects of production. In addition, the advantages and disadvantages of the various technologies used, the features of the use in pharmaceutical technology, as well as additional technological methods for creating modifications of the release of drugs from chewing gum medicinal gums are given. It is concluded that the application of various technologies for the production of chewing gums and the methods for improving individual stages of the process are promising.

Introduction. There is evidence that disseminated tumor cells (DOCs) in the bone marrow (BM) are precursors of subsequent distant metastases. The detection of DOCs in non-small cell lung cancer (NSCLC) will provide important information about the features of metastasis, as well as the possibilities of identifying new targets for the treatment of NSCLC.

Purpose of the study is to evaluate the possibility of detection DOCs in BM and to identify the frequency of BM involvement in patients with NSCLC.

Materials and methods. Morphological and immunological methods studied 62 BM samples from patients with NSCLC. DOCs analysis was performed using flow cytometry (FACSCanto II, USA, Kaluza Analysis v 2.1 software), monoclonal antibodies to CD45, cytokeratins directly labeled with various fluorochromes were used.

Results. DOCs (EPCAM+CD45–) in the BM were found in 43.5 % of patients (threshold level: 1 cell per 10 million myelocaricytes). The presence of DOCs did not correlate with tumor size, lymph node status, stage of the tumor process. The highest detection rates of DOCs were observed at stages IA and IIA: 60.7 % (3/5) and 58.3 % (7/12) respectively. BM involvement in adenocarcinoma was observed in 45 % (15/33) cases, in squamous cell carcinoma – in 37 % (10/27) samples (p = 0.501). It was found that DOCs are more often detected in more differentiated tumors (p = 0.023). Significant correlations between the presence of DOCs in the BM and myelogram parameters have not been established. A decrease in the number of granulocyte germ cells was observed in 4 % of BM involvement (p = 0.036).

Conclusion. The possibility of detecting DOCs in the BM of NSCLC patients has been established. BM involvement was 43.5 %. DOCs are detected even in the early stages of NSCLC. Relationship between BM involvement and the degree of tumor differentiation was found. More frequent BM involvement was observed in adenocarcinoma compared with squamous cell carcinoma of the lung.

Compliance with patient rights and principles of bioethics

The study protocol was approved by the biomedical ethics committee of N.N. Blokhin National Medical Research Center of Oncology of the Ministry of Health of Russia. All patients gave written informed consent to participate in the study.

Introduction. The molecular basis of radio- and photodynamic therapy (PDT), the mechanism of action of a number of antitumor chemotherapy drugs is oxidative stress (OS). The enzyme hemoxygenase-I (НO-1), a molecular marker of OS, is a key participant in the system of protection and adaptation of tumor cells under stress.

Objective. To find out whether the sensitivity of human melanoma tumor cells to OS depends on the basal and modulator-induced levels of НO-1 expression

Material and methods. Human melanoma cell lines were used in the study. The expression of mRNA НO-1 in cells was studied by real-time RT-PCR, the reactive oxygen species content in cells – by flow cytometry and the cytotoxicity of drugs – by MTT assay.

Results. According to our data, human melanoma cells have different basal levels of HO-1 transcription: high (3.0–3.5 o. u.) in lines MelIL, MelP, medium (1.5 o. u.) in lines MeWo, MelZ, MelIbr and low (0.5 o. e.) – MelMe, A375).There is no direct correlation between the level of basal cell expression of HO-1 and their sensitivity to the OS inducer – Н₂О₂. The hemin-induced increase in HO-1 expression in cells is accompanied by doubled resistance to Н₂О₂. It was found that HO-1 repression in the presence of apigenin was registered in melanoma cells with different basal levels, but sensitization to Н₂О₂ (2–4 times) was observed only for cells with medium (MeWo) and low (A375) levels of basal HO-1 expression. It was found that the decrease in basal expression of HO-1 induced by apigenin is accompanied by an increase in the reactive oxygen species content in cells.

Conclusions. The results of our research allow us to recommend natural flavon apigenin, a modulator of HO-1 expression, for inclusion in the chemotherapy and PDT regimens to increase the effectiveness of human melanoma treatment.

Introduction. Alginate gel encapsulation is widely used in biomedical research to protect cells from mechanical stress and contact with the recipient’s immune system. The use of alginate for transplanting biomedical cellular products to humans imposes high requirements on the sterility of the final product. To select the optimal protocol for cell encapsulation for further in vitro use or transplantation of cells enclosed in an alginate gel, it is necessary to study how sterilization processes affect such characteristics of the obtained gel as the ability to maintain cell viability and proliferation.

The purpose of the study was to compare the effect of ultraviolet radiation and autoclaving on the biological properties of alginate gel.

Materials and methods. MOLT-4 culture cells were encapsulated in 1, 2 and 4 % alginate beads at a concentration of 0.75 × 10⁶ cells/ml. On the eighth day of cultivation, we measured the size of the colonies and determined the viability of the cells in an automatic cell counter.

Results. The colony area for all variants of the concentration of alginate gel was significantly higher for a gel sterilized by UV treatment compared with autoclaving. The difference between the colony sizes for the two methods of alginate sterilization was significant at the accepted level of significance for all variants of gel concentrations (α = 0.05, df = 198, t = 1.972). Moreover, the number of viable cells after dissolution of alginate did not significantly differ between the experimental variants.

Conclusion. Thus, autoclaving sterilization leads to proliferation retardation of encapsulated cells and cannot be recommended for use in protocols for the preparation of alginate capsules with encapsulated cells for in vitro and in vivo cultivation.

Introduction. The introducing of tumor molecular profiling into clinical practice has revealed the need for development of new analytical methods for estimating marker expression in solid tumors, as routinely used method of immunohistochemistry has a number of significant drawbacks.

Objective. Analytical validation of immunofluorescence staining and flow cytometry method developed by the authors for the examination of tumor protein markers in the solid tumors tissue.

Materials and methods. Method validation was carried out by quantitative estimation of βIII-tubulin (TUBB3) expression in single-cell suspensions of non-small-cell lung cancer obtained from surgical tumor samples. Primary antibodies to TUBB3 (ab7751) and secondary DyLight 650-conjugated antibodies (ab98729) were used for immunofluorescent staining. The «Navios» flow cytometer (Beckman Coulter) was used to measure the fluorescence. The validation parameters were assessed by the coefficient of variation calculated as the ratio of standard deviation of TUBB3 level to its mean value.

Results. Two parameters were analyzed: intra-assay precision and time stability of the results of the TUBB3 expression assessment. It was demonstrated that the mean coefficients of variation of the marker expression level in the tumor tissue did not exceed 20 % for both parameters. According to recommendations on the analytical validation of methods based on flow cytometry, it proves the validity of the method for these parameters.

Conclusions. The intra-assay precision and time stability were demonstrated for the results of a quantitative estimation of TUBB3 expression in solid tumor tissue using immunofluorescence staining and flow cytometry method developed by the authors. The practical value of the time stability of immunofluorescence stain during 24 h storage of a stained cells suspension in the dark at 4 °C was highlighted. It shows the possibility of adjusting the time interval between completion of the analytical study part and flow cytometer measurement.

Introduction. Amphotericin B remains the drug of first choice in the treatment of most severe systemic fungal infections. However, it is characterized by very low solubility and high toxicity. Amphamide – semisynthetic derivative of Amphotericin B – have been prepared at the Gause Institute of New Antibiotics. It showed several advantages over amphotericin B in vivo.

Objective. The aim of the study was to investigate the toxicological safety of amphamide drug formulation in chronic experiment on rats.

Materials and methods. The study was performed in male and female Wistar rats. Amphamide drug formulation was injected intraperitonealy at the total doses of MTD and LD₅₀ (30 × 0,07 mg/kg or 30 × 0,17 mg/kg with 24-h interval). During the experiment body weight, hematological parameters, blood biochemical parameters, electrocardiography and urinalysis were performed. Animals were sacrificed on 1st and 30th day after the end of treatment. At necropsy, the mass coefficients of heart, liver, kidneys, spleen and thymus were calculated. The internal organs were subjected to histological evaluation.

Results. It has been shown that the treatment with total dose of amphamide produces an increase of urea and creatinine level in serum, changes in urine composition and its specific gravity. Microscopic pathology observation showed dose-dependent structure abnormalities in kidneys, liver, lungs, stomach, and testes. Multiple administration of low dose of the drug produces transient toxic effects completely reversible within 30 days. When amphamide was used in a high dose, morphological signs of toxic cardiomyopathy were found.

Conclusion. The results of the clinical and laboratory studies and microscopic pathology observation of kidneys demonstrate that nephrotoxicity is the main limiting type of drug toxicity. Dose dependence and reversibility within a month of toxic effects of amphamide allows us to recommend it to further advance.

Introduction. Immunoregulatory functions of myeloid-derived suppressor cells have been extensively studied over the recent decades. Additionally, myeloid-derived suppressor cells have been investigated as a prognostic factor.

Objective. To optimize the measurement of monocytic myeloid-derived suppressor cells in the peripheral blood of patients with B-cell chronic lymphocytic leukemia.

Materials and methods. The number of myeloid-derived suppressor cells with CD14⁺HLA-DR^low/–-phenotype was determined in the peripheral blood of patients with B-cell chronic lymphocytic leukemia and healthy donors by flow cytometry. The measurement was carried out at two points, which differed in the concentration of anti-HLA-DR antibodies − 15 and 4 μl.

Results. The median amount of myeloid-derived suppressor cells in the peripheral blood of B-cell chronic lymphocytic leukemia patients with 15 μl of anti-HLA-DR antibody was 1.9 %, and with 4 μl of antibody concentration – 7 %. Healthy donors had that median of 0.15 % with 15 μl of antibody and 0.3 % with 4 μl concentration.

Conclusion. The number of CD14⁺HLA-DR^low/–-cells in the blood of patients with B-cell chronic lymphocytic leukemia is sensitive to the concentration of the HLA-DR antibody.

Compliance with patient rights and principles of bioethics. The study protocol No 3 of 13.02.2020 was approved by the biomedical ethics committee of Research Institute of Experimental Diagnostics and Therapy of Tumors N.N. Blokhin National Medical Research Center of Oncology of the Ministry of Health of the Russian Federation. All patients gave written informed consent to participate in the study.