Krebs-2 solid carcinoma was cured using a new “3+1” strategy for eradication of Krebs-2 tumor-initiating stem cells. This strategy was based on synchronization of these cells in a treatment-sensitive phase of the cell cycle. The synchronization mechanism, subsequent destruction of Krebs-2 tumor-initiating stem cells, and cure of mice from a solid graft were found to depend on the temporal profile of the interstrand cross-link repair cycle. Also, the temporal profile of the Krebs-2 interstrand repair cycle was found to have a pronounced seasonal cyclicity at the place of experiments (Novosibirsk, Russia). As a result, the therapeutic effect that is based on application of the described strategy, originally developed for the “winter repair cycle” (November−April), is completely eliminated in the summer period (June−September). We conclude that оne of the possible and the likeliest reasons for our failure to observe the therapeutic effects was the seasonal cyclicity in the duration of the interstrand repair cycle, the parameter that is central to our strategy.

Stem-like tumor cells of ascites carcinoma Krebs-2 and Epstein-Barr virus–induced B-lymphoma were shown to possess the innate capability of binding and internalizing the TAMRA-labeled double-stranded DNA (dsDNA) probe. The process of binding and internalizing is rather complicated and composed of the following successive stages: 1) initiating electrostatic interaction and contact of a negatively charged dsDNA molecule with a positively charged molecule(s) on the surface of a stem-like tumor cell; 2) binding of the dsDNA probe to a tumor stem cell surface protein(s) via the formation of a strong chemical/molecular bond; and 3) the very internalization of dsDNA into the cell. Binding of DNA to cell surface proteins is determined by the presence of heparin/polyanion-binding sites within the protein structure, which can be competitively blocked by heparin and/or dextran sulfate, wherein heparin blocks only the binding, while dextran sulfate abrogates both binding and internalization. The abrogation of internalization by dextran sulfate implies the role of scavenger receptors in this process. Cells were shown to uptake DNA in amounts constituting ∼0.008% of the haploid genome. Inhibitors of caveolae-dependent internalization abrogate the DNA uptake in Krebs-2 cells, and inhibitors of the clathrin/caveolar mechanism block the internalization in B-lymphoma cells. In the present report, it is shown for the first time that in contrast to the majority of committed tumor cells, stem-like tumor cells of Krebs-2 and B-lymphoma carry a general positive charge on their surface.

Background and Aims: A new technology based on the chronometric administration of cyclophosphamide and complex composite double-stranded DNA-based compound, which is scheduled in strict dependence on interstrand crosslinks repair timing, and named “Karanahan”, has been developed. Being applied, this technology results in the eradication of tumor-initiating stem cells and full-scale apoptosis of committed tumor cells. In the present study, the efficacy of this novel approach has been estimated in the model of Lewis carcinoma.Methods: To determine the basic indicative parameters for the approach, the duration of DNA repair in tumor cells, as well as their distribution along the cell cycle, have been assessed. Injections were done into one or both tumors in femoral region of the engrafted mice in accordance with the developed regimen. Four series of experiments were carried out at different periods of time. The content of poorly differentiated CD34+/TAMRA+ cells in the bone marrow and peripheral blood has been determined. Immunostaining followed by the flow cytometry was used to analyze the subpopulations of immune cells.Results: The high antitumor efficacy of the new technology against the developed experimental Lewis carcinoma was shown. It was found that the therapy efficacy depended on the number of tumor growth sites, seasonal and annual peculiarities. In some experiments, a long-term remission has been reached in 70% of animals with a single tumor and in 60% with two tumors. In mice with two developed grafts, mobilization capabilities of both poorly differentiated hematopoietic cells of the host and tumor stem-like cells decrease significantly. Being applied, this new technology was shown to activate a specific immune response. There is an increase in the number of NK cell populations in the blood, tumor, and spleen, killer T cells and T helper cells in the tumor and spleen, CD11b+Ly-6C+ and CD11b+Ly-6G+ cells in the tumor. A population of mature dendritic cells is found in the tumor.Conclusion: The performed experiments indicate the efficacy of the Karanahan approach against incurable Lewis carcinoma. Thus, the discussed therapy is a new approach for treating experimental neoplasms, which has a potential as a personalized anti-tumor therapeutic approach in humans.

Background/Aim: We previously have described the "3+1" tumors cure approach consisting of individual time schedule of cyclophosphamide and dsDNA preparation administrations. The aim of the study was to adapt the "3+1" approach based on eradication of cancer stem cells to the model of murine ascitic cyclophosphamide-resistant lymphosarcoma (RLS). Materials and Methods: Adaptation of the "3+1" approach includes the identification of the timing to disrupt the tumorigenic potential of a certain tumor. Results: The proposed therapeutic scheme allowed complete reduction of primary RLS ascites in experimental animals. However, reduction of primary ascites due to the complementary action of cyclophosphamide and dsDNA was inevitably followed by the development of a secondary one, most likely arising from a solid carcinomatous formation in the peritoneal wall. Conclusion: The "3+1" approach resulted in the elimination of cancer stem cells, and, as a consequence, in the complete reduction of RLS ascites.Malignant neoplasms rank second in the world after cardiovascular disorders in overall mortality rates. Recent studies indicate a much higher complexity of the disease than previously thought. The main characteristic of this complexity determining the unpredictable response of neoplasms to a variety of therapeutic procedures and the difficulties in their cure is the heterogeneity of malignant cells that include a subpopulation of almost "indestructible" cells, which possess an incredible survival and proliferative potency, and is designated as tumor-initiating stem cells or cancer stem cells (CSCs) (1, 2).The main hallmarks of cancer stem cells are their capabilities of (1) self-renewal in through unlimited replicative cycles, (2) producing a progeny of committed cells with high proliferative activity, but incapable of inducing a new tumor, and (3) retaining a potency to induce new tumors with similar histological properties in a series of transplantations (3). CSCs possess a number of features allowing their dominance in co-existence with organism: self-sufficiency in proliferative stimuli (4), reduced sensitivity to anti-proliferative ones (5), immortalization (6, 7), dedifferentiation (8), genomic instability (9), increased efflux and metabolism of xenobiotics (10, 11), reversed "Warburg effect" (12, 13), suppression of pro-apoptotic signals ( 14), as well as stimulation of pathways causing apoptosis evasion (15). Another essential for disease development property of CSCs is their migratory capacity, allowing exit into the bloodstream, and formation of new growth foci in distant organs (16)(17)(18)(19)(20). Thus, the aforementioned properties of CSCs are such that they create 795