Animal tumor models are used for preclinical studies of drugs and cancer therapy. The aim of this work was to analyze the growth of murine pancreatic tumor cells Pan02, carrying GFP marker, injected subcutaneously (s. c.), intraperitoneally (i. p.) or orthotopically into the pancreas (ortho) of C57BL/6 mice. Mice were injected with 2 × 105 cells: s. c. in the right flank; i. p. with a syringe into the abdominal cavity, or ortho surgically under the pancreas capsule. The weight of mice was determined in the dynamics of tumor growth, and blood serum was taken to analyze the antibody response to the GFP reference protein. At the 2nd and 4th weeks of tumor growth, some mice were slaughtered and the expression of GFP by the tumor cells, as well as the composition of the immune cells in the tumor, were analyzed by flow cytometry and confocal microscopy. It was shown that with the different localization, the pancreatic tumors grew at different rates and lethality. When the tumor was injected i. p., mice lost weight with rapid tumor growth. In the ortho model, the mice increased their weight. Mortality in the s. c. and i. p. groups was comparable. In the s. c. model, the tumor grew slowly to a volume of 200-400 mm3 and stopped growing. There was no mortality in this group during the follow-up period (2 months). The same antibody response to GFP was formed with all injection schemes. The subpopulation composition of immune cells varied greatly in the different models of tumor cell administration. Regardless of the type of immune response, Pan02-GFP cells rapidly suppressed GFP gene expression in vivo. The data obtained showed that murine pancreatic tumor Pan02 is immunogenic and causes the formation of an adaptive immune response. Regardless of the presence or absence of an immune response and elimination of GFP+ cells, the tumor continued to grow in the i. p. and ortho models, but not in the s. c. one, and caused the death of mice. When conducting preclinical studies, it is necessary to use several ways of tumor cell injection to obtain a more objective result.

The family of genes containing C2H2 zinc finger domains, which has more than 700 members, is one of the largest in the genome. Of particular interest are C2H2 genes with potential tissue-specific transcription, which determine the functional properties of individual cell types, including those associated with pathological processes. The aim of this work was to identify C2H2 family genes with tissue-specific transcription and analyze changes in their activity during tumor progression. To search for these genes, we used four databases containing data on gene transcription in human tissues obtained by RNA-Seq analysis. The analysis showed that, although the major part of the C2H2 family genes is transcribed in virtually all tissues, a group of genes has tissue-specific transcription, with most of the transcripts being found in the testis. After having compared all four databases, we identified nine such genes. The testis-specific transcription was confirmed for two of them, namely ZBTB32 and ZNF473, using quantitative PCR of cDNA samples from different organs. A decrease in ZBTB32 and ZNF473 transcription levels was demonstrated in germ cell tumors. The studied genes can serve as candidate markers in germ cell tumors.

The emergence of genetic diseases and evolutionary processes are associated with the flow of genetic information from one generation to another, in which genetic information carried by gametes can be changed by the appearance of de novo germline mutations. The rate of germline mutations determines the rate of evolution and the incidence of heritable disorders. Despite the great theoretical and practical importance, the problem of establishing mutation rates and their dependence on different factors remains scarcely studied, and the mutation rate values obtained by different methods vary considerably. The review discusses different ways of estimating the rate of these mutations and makes an attempt to explain the reasons for discrepancies in the data obtained. Three levels of the mutation formation are considered: (1) mutations that are formed during the development of a given individual during gametogenesis (basic mutations); (2) mutations transmitted to offspring and determining differences in the genomes of consecutive generations (parents and offspring), which include basic mutations and possible changes resulting from complex processes of sperm transfer to oocyte, fertilization, and subsequent events that lead to only one viable offspring of hundreds of millions spermatozoa and oocytes; (3) mutations which are formed at level 2, fixed in evolution, and determine evolutionary processes and differences between genomes, in particular, of hominoids, hominids, and hominins.

The embryonic development of all body systems is controlled by coordinated interactions of a group of genes encoding master regulators of development, which determine the fate of cells in the process of differentiation. These genes are often involved in the processes of malignant degeneration of cells. Hence, they are considered as targets of targeted gene therapy in the development of modern approaches to cancer treatment. One of the most aggressive forms of cancer is pancreatic ductal adenocarcinoma (PDAC). Thus, the study of pancreas master regulators genes is a challenging task. This review is devoted to SOX9, which is one of the key master regulators of pancreatic development and is a crucial factor in the induction and progression of PDAC.

One of the most popular nonviral delivery systems for gene therapy constructs are carriers based on polyethylenimine (PEI) DNA complexes. A number of disadvantages associated with the lack of targeted delivery and increased cytotoxicity are overcome by adding auxiliary molecules to the complexes. An example of this is chondroitin sulfate (CS). The purpose of this work was to assess the effect of CS on the transfection properties of DNA–PEI complexes under different conditions of their preparation and transfection protocols. All complexes were prepared in solutions with high and low ionic strength. Transfection of C26 cells was performed according to two protocols differing in the presence of serum in the medium. The portion of transfected cells, transgene expression level, and cell viability were the main parameters of assessing the transfection efficiency. In binary DNA–PEI complexes prepared in different salt conditions, using different transfection protocols, the difference in the portion of transfected cells reached ten times. Addition of CS improved this transfection efficiency indicator up to 6.5 times, while the maximum difference in this indicator for the corresponding ternary complexes was reduced to 2.5 times. Changes in the proportion of CS in the composition of the complexes had an insignificant effect on their transfection properties. In the case of complexes prepared in high ionic strength solutions, the order of CS addition was also important. The best results of transfection efficiency were achieved with ternary complexes prepared in low ionic strength solutions, using a serum-free protocol, while these indicators were comparable with the data for Lipofectamine 2000. The addition of chondroitin sulfate improves the transfection properties of DNA–PEI complexes and makes them less dependent on the methods of preparation and transfection.

An artificial double tandem tumor-specific promoter based on survivin and human telomerase reverse transcriptase gene promoters was constructed. Studies in in vitro and ex vivo therapeutic systems showed that the designed promoter exhibits a high activity in tumor cells, which is comparable to the activity of the CMV constitutive promoter.

The review discusses the causes of multiple failures in cancer treatment, which might primarily result from the excessive variability of cancer genomes. They are capable of changing their spatial and temporal architecture during tumor development. The key reasons of irreproducibility of biomedical data and the presumable means for improvement of therapeutic results aiming at targeting the most stable tumor traits are suggested.

Contents of J-peptide of secreted human polymeric immunoglobulins may vary considerably with different pathologies, reflecting the state of the adaptive immune system. In this work assessed the content of J-peptide in various tissues of healthy people to use as a baseline for studies related to the change in the content of J-peptide in pathologies.

The content of the J peptide of secretory polymeric immunoglobulins can significantly change during various pathologies, reflecting the state of the adaptive immune system. In this study, the content of J peptide was determined in various tissues of healthy people. These results can be used as basic data for investigations of the changes in the content of J peptide during different pathologies.

С помощью системы позитивно-негативной селекции в области хромосомы 19 человека длиной 106 пар оснований, расположенной между генами FXYD5 и COX7A1, идентифицированы 10 потенциальных инсулято∙ ров, обладающих способностью блокировать взаимодействие энхансера с промотором. Взаимное расположение инсуляторов и генов согласуется с гипотезой, согласно которой инсуляторы подразделяют геномную ДНК на независимо регулируемые петлевые домены.

A positive-negative selection system revealed 10 potential insulators able to block enhancer interaction with promoter in the 10(6) bp human chromosome 19 region between genes FXYD5 and COX7A1. Relative positions of insulators and genes are in accord with the hypothesis that insulators subdivide genomic DNA into independently regulated loop domains.

Secreted polymeric immunoglobulins, IgA dimers and IgM pentamers, are unique in having not only the L and H chains but also the J chain, which is responsible for oligomerization. These antibodies belong to the local adaptive immune system, located in the mucosas of the respiratory and digestive systems and acting first to protect the body from infectious agents. Polymeric immunoglobulins are produced by highly specific B cells and are actively transported to the mucosa surface through epithelial cells. Hence, their production and, accordingly, the J-chain content are associated with the transporting function of the epithelium and depend on its state, which dramatically changes upon malignant transformation. The content of the J chain and the transcription level of its gene were studied in normal and tumor cells at various stages of squamous cell cancer and adenocarcinoma of the lungs by RT-PCR and immunoblotting.

S/MARs (scaffold/matrix attachment regions) are the DNA regions that are involved in the interaction with the nuclear matrix and are identified by in vitro methods. According to the available information, S/MARs possess an insulating activity, i.e., the ability to block the interaction between the enhancer and promoter in vivo, and are, probably, intact insulators or their fragments. Nevertheless, there is still no direct proof for this correspondence. To obtain additional information on the insulator activity of S/MARs, we selected five DNA fragments of different lengths and affinities for the nuclear matrix from a previously constructed library of S/MARs and tested their ability to serve as insulators. Two of five elements exhibited an insulator (enhancer blocking) activity upon the transient transfection of CHO cells. None of the S/MARs displayed either promoter or enhancer/silencer activities in these cells.

Destabilase-lysozyme (DL) from salivary gland secretion of the medicinal leech (Hirudo medicinalis) is as a member of the invertebrate lysozyme family, which sharply differs from other lysozyme families. In this study, DL lysozyme function was confirmed during expression of a gene encoding DL in Escherichia coli. Several constructs of the expression vectors pKK OmpA and pET-3A with or without bacterial, leech, or yeast signal peptides (SP) were used. The use of a construct without signal peptide genes resulted in normal growth of the transformed cells. Transformation of E. coli cells with the constructs containing SP was accompanied by the disruption of the forming cells. The use of the expression vector pET-32 LTC-System for production of DL as a fusion protein with thioredoxin also resulted in normal cell growth. However, specific activity of DL isolated from such cells was significantly lower than that of enzyme purified from extracts of Spodoptera frugiperda cells, which were infected with the baculovirus vector carrying DL cDNA. It is shown that the action mechanism of invertebrate lysozyme does not differ from that of other families: recombinant DL from S. frugiperda extracts catalyzed cleavage of synthetic substrate, hexamer of N-acetylglucosamine, to di- and tetramers, which is typical for enzymatic function of other lysozyme families.

Reporter gene analysis of HERV-K solitary long terminal repeats (LTRs) showed that they retain detectable activity in human teratocarcinoma cells, and can direct the transcription in both orientations relative to the reporter gene. Deletion analysis demonstrated the possible existence of alternative promoters within the LTR as well as a silencer-like element in the U5 region. Our results indicate also that all-trans-retinoic acid is capable of modulating expression of the reporter gene directed by a HERV-K LTR in NT2/D1 cells.

Intrinsic lysozyme-like activity was demonstrated for destabilase from the medicinal leech supported by (1) high specific lysozyme activity of the highly purified destabilase, (2) specific inhibition of the lysozyme-like activity by anti-destabilase antibodies, and (3) appreciable lysozyme-like activity in insect cells infected with recombinant baculoviruses carrying cDNAs encoding different isoforms of destabilase. Several isoforms of destabilase constitute a protein family at least two members of which are characterized by lysozyme activity. The corresponding gene family implies an ancient evolutionary history of the genes although the function(s) of various lysozymes in the leech remains unclear. Differences in primary structures of the destabilase family members and members of known lysozyme families allow one to assign the former to a new family of lysozymes. New proteins homologous to destabilase were recently described for Caenorhabditis elegans and bivalve mollusks suggesting that the new lysozyme family can be widely distributed among invertebrates. It remains to be investigated whether the two enzymatic activities (isopeptidase and lysozyme-like) are attributes of one and the same protein.