Медицина/6.Экспериментальная и клиническая фармакология

Babiy O.P., Grehirchak N.M.¹, Shpak E.G.²

¹National University of Food Technologies, Ukraine
²R.Y. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology NAS of Ukraine, Kyiv

SCREENING ADJUVANTS ORGANIC AND INORGANIC NATURE FOR CONSTRUCTING CANCER VACCINES

Problem of tumor growth is the most important in modern medicine and biology. Above it are dealt hundreds of medical laboratories around the world. In this area has been done much research, but its solution is still far ahead. Therefore, the search for new drugs to treat cancer is an open topic. At present the views of scientists around the world aimed at using alternative means, namely the methods of biotherapy, especially immunotherapy of cancer patients [1]. The last has several areas, including the use of cytokines, monoclonal antibodies and specific anti-tumor vaccines. The use of anticancer vaccines is very promising approach, it’s made on the tumor associated antigens, which are based on the formation of specific reactions of antitumor immunity. It should be noted that most of the tumor associated antigens have low immunogenicity, which leads to the need to find different ways to improve the efficiency of anticancer vaccines [2]. The one of ways to enhance the immune response to this antigens is using of adjuvants, which spectrum is quite broad. However, the impact of adjuvant on immunogenicity of tumor associated antigens and the dynamics of nonspecific and specific antitumor responses remains is still unspecified.

The work was aimed on the selection of potential adjuvant for designing antitumor vaccines and study their effects on the immune system in animal experiments with Lewis lung carcinoma (LLC).

In experiment were used male Balb / c line 2-2.5 months old and average weight 18 - 20 g mice obtained from vivarium of R.E. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology NAS of Ukraine. As a model of tumor growth was used the Lewis lungs metastatic epidermoid carcinoma.

A series of experiments, namely triple immunization of animals by chicken embryonic proteins (0.1 mg of protein per injection) were carried out in mono or in combination with adjuvant: lipids from cell B. subtilis B-7025 molecular weight 18.5 kDa and 70 kDa (0.006 mg/injections), microbial cell BCG (0,3×10⁸ CFC/injections), colloidal silver (Ag) and suspension of iron oxide (Fe₃O₄) in 2% solution of polidekstran (0.06 mg/injections). For intact control (IC) were used animals injected with NaCl.

Immunological examination included: determination of cytotoxic activity and antibody-dependent cytotoxic activity of lymphocytes and macrophages, cooperative cytotoxic activity of effecter cells, cooperative antibody-depended cellular cytotoxicity of lymphocytes and macrophages, ELISA detection of generated antibodies specific to chicken embryonic proteins or tumor antigens LLC.

As the results of investigations evidenced, the introduction of chicken embryonic proteins by themsalve independently, so do in combining with adjuvant caused inhibition of growth of LLC in experiments on animals. Stability of this effect remains at all stages of growth of experimental tumors. The comparative analysis of the size of primary tumors in animals from different groups at the end of the experiment (34th day) showed that in animals, who received the vaccine based on CEB with glycoproteins B. subtilis B-7025 tumor volume was 13% lower than in the IC . It is necessary to note that the degree of inhibition of tumor growth in terms of different tumor process was uneven.

The dynamic of growth of the LLC after interruption to the animals studied vaccines was different. In the primary stages of tumor process the interruption of all the studied substances resulted inhibition of tumor growth. Through the development of the tumor suppressive effect of the studied preparations has been gradually decreased. The most expressive effect was in animals that received the vaccine based on CEP and glycoproteins as adjuvant.

Within the immunological experiments there were established that the maximal synthesis of antibodies was observed in groups of animals, which as an adjuvant to CEP got metabolite B. subtilis B-7025 with mol. weight 18.5 kDa and 70 kDa and peptidoglycan of S.aureus cells. In the group of animals where as adjuvant were used BCG synthesis of antibodies was lower, than in group with chicken embryonic proteins (Fig. 1). According to this we can conclude that BCG activates the cellular immunity and suppress of humoral.

Figure 1. ELISA detection of serum in experimental groups specific to chicken embryonic proteins provided immunization CEP with adjuvants.

Similar results were also obtained in assessing the accumulation of antibodies to proteins of Sarcoma 37 (Fig. 2).

It was found that in all experimental groups, the level of medium molecular circulating immune complexes (CIC) in serum was higher compared with intact animals. In mice immunized with CEP and Fe₃O₄, CIC level exceeded the same index in animals that received no adjuvant chicken embryonic proteins. Add to CEP almost all investigated adjuvant (except a mixture of lipids B. subtilis B-7025) led to a decrease in titer of antibodies against protein S-37. As a result of the test to determine cytotoxic activity of lymphocytes against cells S-37, demonstrates that the introduction of CEP is not likely led to its change.

Figure 2. ELISA detection of serum proteins in experimental groups against Sarcoma 37.

It is shown that a mixture of lipids B. subtilis B-7025 has immunotoxic effects on the mice Balb/c and does not cause inflammatory reactions. Introduction of CEP with adjuvants, mainly with lipids of B. subtilis B-7025, induces the formation of specific IgG in the serum of animals. These data suggest the feasibility study of lipids as potential immunomodulating agents for their further use in oncology practice.

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