A Universal Tumor-Specific Promoter for Cancer Gene Therapy.

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.

Expression of the FAP gene in non-fibroblast human cell lines. Development of cancer-associated fibroblast models.

The fibroblast activation protein (FAP) is selectively expressed in cancer-associated fibroblasts (CAFs) and facilitates tumor progression, which makes this protein an attractive therapeutic target. There are difficulties in obtaining CAFs for studying the function and suppression of FAP. In this work, the expression level of FAP was determined by PCR assay in 25 human cell lines and 8 surgical samples of tumor stroma. The expression of FAP was observed in all tumor stroma samples and in four cell lines: NGP-127, SJCRH30, SJSA-1, and A375. The level of FAP expression in NGP-127, SJCRH30, and SJSA-1 lines as well as in CAFs of patients was comparable, which makes these cell lines a possible model for studying FAP.

[The cause of cancer mutations: Improvable bad life or inevitable stochastic replication errors?]

Despite substantial progress in understanding the mechanisms of carcinogenesis and fighting oncology diseases, cancer mortality remains rather high. Therefore, there is a striving to reduce this mortality to the level determined by endogenous biological factors. The review analyzes the mutations that lead to cell malignant transformation and describes the contribution that self-renewal of adult tissues makes to tumorigenesis. Cancer progression is considered as a development of a complicated system where cells mutate, evolve, and are subject to selection. Cancer paradoxes are described in conclusion.

[FIBROBLAST ACTIVATION PROTEIN (FAP) AS A POSSIBLE TARGET OF THE ANTITUMOR STRATEGY.]

This review was devoted to the use of the versatile component oftumoral stroma (fibroblast activation protein, FAP) as a target of the versatile tumor therapy. The tumor is a coevolution system, which includes the microenvironment or reactive stroma differing from the normal tissue by the phenotypic and genotypic features. Important elements of the tumor microenvironment are cancer-associated fibroblasts (CAFs), which contain typical marker FAP (serine proteinase with the enzymatic activity of dipeptidyl peptidase and endopeptidase). According to the literature, more than 90% of tumors contain FAP-positive activated fibroblasts. FAP is virtually absent in normal tissues, but it is present in the embryonic and tumor tissues, which makes it a selective and versatile model. In this work, basic approaches to affecting the CAF using FAP as a target were discussed. The use of FAP as a target provides an important advantage: its proteolytic activity can be used along with the protein-targeted agents. The main directions in the therapeutic use of FAP were discussed in this work.

[Fundamentally low reproducibility in molecular genetic cancer research].

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.

[ADULT STEM CELLS AND CELLS OF MALIGNANT ORIGIN. PART I].

Recent data on adult stem cells are reviewed. According to the present dominant paradigm, it is most probable that cancer predisposition arises or cancer is initiated in these cells.

[ Super-enhancers. Are they regulators of regulatory genes of development and cancer?].

Enhancers make up a huge class of genome regulatory elements that play an important role in the formation and maintenance of specific patterns of gene transcriptional activity in all types of cells. In recent years, high-throughput methods for the genome-wide epigenetic analysis of chromatin have made it possible to identify structural and functional features of enhancers and their role in the spatial and functional organization of the genome and in the formation and maintenance of cell identity, as well as in the pathogenesis of certain diseases. Special attention has been focused on genome regions called super-enhancers, or stretch enhancers, which consist of clusters of elements with properties of classic enhancers. This review considers current data on specific properties of super-enhancers and their role in the formation of interconnected autoregulatory circuits with positive feedback that regulates the most important genes, the activity of which underlies the formation and maintenance of specialized cellular functions.

[ADULT STEM CELLS AND CELLS OF MALIGNANT ORIGIN. PART II].

Recent data on adult stem cells are reviewed. According to the present dominant paradigm, it is most probable that cancer predisposition arises or cancer is initiated in these cells.

[Cytotoxicity of cytosine deaminase and herpes simplex virus thymidine kinase genes in melanoma cells is independent on promoter strength].

In preparation of the therapeutic genetic constructs aimed to the gene-programmed enzymatic transformation of the non-toxic prodrug into toxin within cancer cells the right choice of regulatory elements (promoters and enhancers) is essential. This is widely accepted that the efficiency of the gene therapy constructions is dependent, in particular, on the strength of promoters driving the expression of the therapeutic genes. In this work we demonstrated, using the melanoma-specific promoters and enhancers of human melanoma inhibitory activity and mouse tyrosinase gene, that for the development of cytotoxic effect the promoter strength is not of primary importance. In the case of HSVtk, coding for the herpes simplex virus thymidine kinase, and FCU1, coding for cytosine deaminase/uracil phosphoribosyltransferase hybrid protein genes, their cytotoxic activity was determined by the quantity of the added prodrug.

[Melanoma: surface markers as the first point of targeted delivery of therapeutic genes in multilevel gene therapy].

Melanoma is one of the most malignant tumors, aggressively metastasizing by lymphatic and hematogenous routes. Due to the resistance of melanoma cells to many types of chemotherapy, this disease causes high mortality rate. High hopes are pinned on gene therapeutic approaches to melanoma treatment. At present, one of the main problems of the efficient use of the post-genomic generation therapeutic means is the lack of optimal techniques of delivery of foreign genetic material to the patient's target cells. Surface specific markers of melanoma cells can be considered as promising therapeutic targets. This review describes currently known melanoma specific receptors and its stem cells, as well as contains data on melanoma antigens presented on the cell surface by major histocompatibility complex proteins. The ability of surface proteins to internalize might be successfully used for the development of methods of targeted delivery of gene therapeutic constructs. In conclusion, a concept of multilevel gene therapy and the possible role therein of surface determinants as targets of gene systems delivery to the tumor are discussed.

Promoters with cancer cell-specific activity for melanoma gene therapy.

Melanoma is one of the most aggressive tumors. It develops from pigment-forming cells (melanocytes) and results in a high number of lethal outcomes. The use of genetic constructs with the ability to specifically kill melanoma cells, but not normal cells, might increase the lifespan of patients, as well as improve their quality of life. One of the methods to achieve a selective impact for therapeutic genes on cancer cells is to utilize a transcriptional control mechanism using promoters that are specifically activated only in cancerous cells. In this review, promoters of the genes that are preferentially expressed in melanoma cells are described. These promoters, and other highly melanoma-specific regulatory elements, reduce the unspecific expression of therapeutic genes in normal tissues. Moreover, cancer-specific promoters and their elements are advantageous for the development of universal anticancer drugs. Examples of the use of double promoters that have a high potential as instruments in cancer gene therapy are also given in this review.

[Functional analysis of the HERV-K LTR residing in the KIAA1245/NBPF subfamily].

Long terminal repeats (LTRs) of human endogenous retrovihuses (HERs) might affect transcription regulation of neighboring genes. In our previous study, we showed that the solitary LTR residing in the KIAA1245/NBPF gene subfamily displayed high enhancer activity in a transformed embryonal carcinoma cell line Tera1. In this study, we performed a functional dissection of the LTR and studied its deletion series. Using transient transfection assay, we confirmed the ability of the LTR to drive the expression of the luciferase reporter gene in Teral cells. At the same time, in two other transformed cell lines tested, NGP and NT2/D1, the full-size LTR and its fragments showed no or low enhancer activity, thus demonstrating cell type specificity of the LTR enhancer activity. The functional dissection of the LTRrevealed a specific region within the U3 part appeared to be responsible for the enhancer properties. We showed that the identified enhancer was able to work in a highly cell type specific manner. The data obtained are in line with the hypothesis suggesting that KIAA1245/NBPF LTR may affect the regulation of the KIAA1245/NBPF subfamily genes transcription.

[Transcription of the KLRB1 gene is suppressed in human cancer tissues].

The KLRB1 gene encodes the CD161 receptor of natural killer cells (NK-cells). The gene is also expressed in the NKT-cells. The two cell types are cytotoxic and capable of recognizing and eliminating various cell species, e.g. tumorous, virus-infected, and allotransplants. The biological function of human CD161 is still insufficiently understood; probably it is involved in regulation of the cytotoxic functions of the cells and in regulation of cytokine production. Because immune system, in particular, the activity of cytotoxic cells, is suppressed in most cancer patients, it was suggested that the KLRB1 expression might be suppressed in cancerous cells. The results of this work demonstrated that the transcription of the KLRB1 was suppressed in tumor tissues in 68% patients with nonsmall-lung-cancer (p < 0.0001) and 57% patients with esophageal squamous-cell carcinoma (p = 0.0003). High frequency of the KLRB1 transcription suppression upon cancers makes it possible to use this parameter as a highly informative marker of lung and esophageal cancers.