INTRODUCTION:

The increase in cancer is a global trend. Cancer is a common disease in all countries of the world. Stress and poor environment are the main causes of cancer. Initial treatment with chemotherapy and radiation therapy may shrink the tumor. However, their longer use never leads to the destruction of the tumor and complete recovery.

Therefore, the search and development of effective natural sources, means of prevention and therapy of malignant neoplasms is one of the global and urgent problems of the world community.

Currently, medicinal substances of natural origin, which include snake venoms, are attracting more and more attention. In the literature there are multilateral studies on the study of the physicochemical properties of viper venom, on the influence of environmental pollutants on the pharmacological properties and toxicity of the biologically active components of the venom, but many questions remain in the study of the pharmacological properties of these invaluable poisonous products of snake biosynthesis^1-10. Every year, scientific research opens up new possibilities for the impact of these valuable products of snakes on the human body.

Tumors of hematopoietic and lymphoid tissues account for approximately 8% of all malignant neoplasms, and together are among the 6 most common types of malignant diseases¹¹.

The basis for diagnosing blood cancer is laboratory testing. Suspicions of the disease may arise when assessing the results of general and biochemical blood tests, which reveal: a decrease in the level of red blood cells and hemoglobin (anemia); low platelet count (thrombocytopenia); increased or decreased level of white blood cells; the phenomenon of “leukemic failure” is possible (the absence of transitional forms of cells, only the youngest forms of immature and fully mature blood elements are detected); the presence in the peripheral and venous blood of young, immature formed elements at different stages of development; disorders of the blood coagulation system¹².

To determine the presence of cancer in experimental animals, the following types of diagnostics are used: palpation and visual examination, x-ray examination; ultrasonography; blood analysis; analysis for histo- or cytology¹³.

The study of red blood cells in the pathogenesis of tumor growth attracts the attention of scientists because red blood cells are carriers of information about the processes occurring in the body. Tumor growth can lead to disruption of the metabolism of red blood cells, which may result in a decrease in their viability and a change in their elastic properties. The accumulation of highly toxic metabolites in the membranes of red blood cells due to the activation of lipid peroxidation and the autocatalytic increase in the amount of its products contributes to the deterioration of membrane parameters and a decrease in the ability of red blood cells to deformations in the microvasculature¹⁴.

Blood testing can provide minimally invasive diagnostics for many diseases, including cancer¹⁵.

Any disease, pathological process, as well as a number of physiological changes can, to one degree or another, affect the quantitative and qualitative characteristics of the composition of circulating blood¹⁶. This determines the great importance of the need to study blood. White blood cells, having high reactivity, quickly become involved in adaptive reactions. They are capable of a nonspecific response in response to alternating influences¹⁷.

Microelements are the most important substances on which the vital activity of organisms depends. They are not a source of energy, but are responsible for vital chemical reactions. Among all microelements, a special group includes the so-called essential microelements, the regular intake of which with food or water into the body is absolutely necessary for its normal functioning.

One of these essential microelements is manganese (Mn).

Manganese actively affects the metabolism of proteins, carbohydrates and fats. Its ability to enhance the action of insulin and maintain a certain level of cholesterol in the blood is also considered important.

Manganese controls and supports many vital functions of the body.

In the presence of manganese, the body uses fats more fully, and the absorption of copper increases. The microelement also regulates hematopoietic processes, enhances the synthesis of thyroid hormones - thyroxine and triiodothyronine, participates in the synthesis of interferon and strengthens the immune system and maintains normal blood clotting.

Manganese is necessary for the formation and functioning of the musculoskeletal system, cell division, and effective brain function (ensuring neurochemical processes in the central nervous system). It is also an antioxidant. Manganese supports the balance of sex hormones, the health of the nervous system and immune resistance.

Manganese deficiency leads to various forms of anemia, reproductive dysfunction in both sexes, growth retardation in children, manifestations of underweight, etc. Currently, deficiency of this mineral is quite common, which is associated with improper and unbalanced nutrition, as well as environmental pollution¹⁸.

It is known that some nanoparticles (NPs) tend to accumulate in tumors. In particular, NPs of iron and manganese oxides can penetrate the brain and, with significant accumulation, cause toxic effects. These facts served as a prerequisite for studying the effects of NPs of these elements on the viability of glioma cells. Gliomas are the most common malignant type of brain tumor. Exposure to manganese oxide NPs led to a significant decrease in the level of glycolysis of U-87 MG glioma cells against the background of a decrease in their viability, while a cumulative radiation dose of 40 Gy did not have a significant effect on glycolysis and viability of U-87 MG glioma cells. There was also no cumulative effect from exposure to radiation and NPs. The results obtained may be of interest in the light of the development of new treatment regimens for gliomas, where manganese oxide NPs can be used as an additive inhibitory component in complex chemotherapy for tumor cells¹⁹.

On the other hand, over the past decade, scientists have created several fundamentally new methods for treating cancer and other diseases, which are based on the action of various organic or inorganic nanoparticles. In some cases, these structures are directly involved in destroying the tumor or eliminating the source of the disease. They serve as unique “targets” that are targeted either by immune cells or by laser radiation. Russian scientists have created a new type of nanoparticles based on iron, zinc and manganese that can “burn out” cancerous tumors when interacting with a magnetic field without killing healthy cells. The first experiments on animals and cell cultures showed that although this “burning out” is very effective, it has quite serious side effects. They are due to the fact that the amount of heat generated by nanoparticles is quite difficult to control. If they release too much energy, then they begin to damage not only the tumor, but also neighboring healthy tissue. Russian physicists have found an ingenious solution to this problem by noting that the magnetic properties of many materials strongly depend on the ambient temperature. Russian researchers synthesized a test batch of nanoparticles from a mixture of iron oxide, manganese and zinc and confirmed that magnetic fields do not heat them to temperatures higher than 45°C. Now magnetite nanoparticles are obtained using vapor deposition or chemical methods. At the same time, impurities inevitably form in them, therefore, in order to obtain a heating effect, it is necessary to influence them with a magnetic field of higher frequency and higher power. Because of this, a person experiences very strong discomfort. An organism weakened by illness is especially vulnerable. You can obtain reference-grade pure magnetite by oxidizing pure iron, and it is necessary to grind it to nanosize so that the substance gets inside the diseased cell and is heated under the influence of a magnetic field. The method becomes especially effective in combination with radiation and chemical therapy - even if not all tumor cells die during a “heat attack,” the survivors are significantly weakened, and the subsequent use of traditional methods can occur in a more gentle manner for the patient. Compared to chemotherapy and radiotherapy, magnetic hyperthermia is more microinvasive, targeted and local, since only cells that have captured magnetic nanoparticles are exposed. However, magnetic hyperthermia itself has very limited application in clinical oncology. It is used to treat certain types of tumors or local metastases in the liver, kidneys, prostate and pancreas. At the same time, it is very difficult to control the distribution of particles in the tumor itself. The method is not designed for radical treatment of most malignant neoplasms^20,21,22.

Snake venom, poisonous secretions of the salivary glands of some species of snakes - a thin, colorless, straw-yellow or greenish liquid; consists of albumins, globulins, enzymes, water, salts, etc. When dry, it can be stored for up to 23 years. The properties of the venom of different snakes are not the same: viper venom contains hemorrhagin, which causes hemorrhages, cobra venom is rich in a neurotoxin that affects the nervous system. Therefore, the consequences of a bite from different snakes manifest themselves differently. When bitten by a viper - pain, swelling in the area of the bite, skin hemorrhages with the formation of blisters (sometimes with tissue necrosis); a cobra bite causes shortness of breath, palpitations, dizziness, nausea, and sometimes vomiting; death occurs from paralysis of the respiratory center. The venom of cobra, copperhead and rattlesnake has an anticoagulating effect on the blood, while the venom of viper has a coagulating effect²³. In the literature there are some assumptions about the properties of the components of snake venom to destroy sarcoma cells.

Conflicting data were obtained from experimental studies of the effect of snake venoms on malignant neoplasms in animals²⁴.

Karimov Z.N.²⁵ found that cobra venom, when administered intravenously to rats, causes an average antitumor effect against sarcoma 45 and M-1; Viper venom, when administered intravenously or intramuscularly, has weak antitumor activity.

Kasenova K.U. and Kunaeva K.S.^26,27 noted that the venoms of snakes of the viper, pit viper and asp families cannot be classified as antitumor forms. The authors recommend the use of snake venoms in oncology as analgesics. It is also known that protein components isolated from cobra venom selectively act on tumor cells without affecting normal cells^28,29,30.

There are conflicting data on the use of snake venom in medicine, in particular in antitumor therapy, and these issues require additional research

From the contradictory data it follows that the use of snake venoms in antitumor therapy requires additional research.

Based on the above, the purpose of this work was to identify the antitumor activity of whole viper venom in an experiment on rats using manganese nanoparticles.

MATERIALS AND METHODS:

The experiments used the poison of the viper Macrovipera lebetina obtusa and manganese nanoparticles.

Experiments were carried out on 20 white outbred laboratory rats, weighing in the range of 200-220g, which were intraperitoneally injected with a mixture of viper venom (Macrovipera lebetina obtusa Linnaeus, 1758) at a dose of 0.01mg/kg body weight with manganese nanoparticles.

The use of a suspension of magnetic particles in a mixture of biologically active components of viper venom, introduced into the tumor area, and exposure to a magnetic field until a temperature of 42°C was reached in the tumor area led to increased destruction of the tumor, while the surrounding tissues were preserved, and the magnetic effect decreased.

The work used a method of inducing hyperthermia using high-frequency magnetic field, which is applied to the tumor with the capture of exogenous nano-sized particles in it³². This technique generates local heat near a magnetic nanoparticle around the tumor in a time-varying magnetic field, which occurs due to the transfer of energy from the external magnetic field into heat. As a result, tumor cells are heated to the Curie temperature, while benign tissue is preserved. This method ensures uniform heating and destruction of tumor tissue and reduces the side effects of magnetic nanoparticles.

Laboratory blood tests were carried out using an Auto Hematology Analyzen Ratyo RT -7600 device, made in China, 2019. Quantitative indicators of red blood cells, leukocytes, lymphocytes and platelets were determined in the blood of white rats as the most informative indicators of the blood being tested.

The activity of enzymes was also determined in the blood of rats - ALT and AST, ALP, γ-GTP, lactate dehydrogenase (LHD), using reagent kits produced by "HUMAN" on a BIOSKREM MS 2000 microanalyzer, manufactured in the USA. Blood content of total protein (TP), total bilirubin (TB), medium molecular peptides (MMP), C-reactive protein (CRP), products of lipid peroxidation (LPO) - hydroperoxide (HP), concentration of diene conjugates (DK), malonic dialdehyde (MDA), total antioxidant status (TAS), catalase (Kat) and superoxide dismutase (SOD) activity were determined on a BIOSKREM MS 2000 microanalyzer, manufactured in the USA, using reagent kits manufactured by “HUMAN”.

The content of triglycerides (TQ), total cholesterol (TC), low-density lipoproteins (LDL), intermediate-density lipoproteins (VLDL), high-density lipoproteins (HDL) was determined by the enzymatic colorimetric method using a set of chemical reagents produced by Human, Germany. Determinations were carried out on an FP-9019 analyzer (made in Finland).

An electrocardiogram was performed using an Elektrocardigraph Carawell ECG-II03GVet device, made in China.

Statistical analysis of quantitative data was carried out using nonparametric methods - the Wilcoxon-Mann-Whitney test. When studying the dependencies between indicators, the Spearman formula for the rank correlation coefficient was applied:

where: d – difference of ranks, n – number of ranks.

When applying the above methods, MS EXCEL and S-PLUS programs were used for statistical processing.

RESULTS AND DISCUSSION:

Viper venom in combination with manganese nanoparticles was injected into a cancerous tumor and exposed to a magnetic field, heating to a temperature of 42℃. As a result, after the introduction of viper venom in combination with iron oxide nanoparticles, significant changes were noted in the blood of experimental animals, as well as the death of diseased cells.

The content of leukocytes in the blood decreased by 12.1%, 19.5%, 26%, granulocytes, monocytes and neutrophils increased, respectively, by 8.6%, 3.8%, 3.9%.

Blood tests are often used to monitor the effectiveness of cancer treatment. An increase in the number of white blood cells indicates the immune system's response to the tumor. A decrease in this value may indicate blood cancer. A decrease in red blood cells and hemoglobin indicates anemia, which often accompanies the development of malignant neoplasms. Liver function tests measure the levels of enzymes and proteins produced by the liver. They include: alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase, gamma-glutamyltransferase (GGT). They may be increased when there is blockage of the liver or bile ducts ¹⁷.

After intraperitoneal administration of viper venom at a dose of 0.02mg/kg body weight of rats mixed with manganese nanoparticles, supraventricular extrasystoles were noted on the ECG. Atriventricular conduction slowed down, and a first-degree block was noted.

Experimental data on the dynamics of changes in blood parameters indicated leukocytosis, lymphopenia, monocytosis, and granulocytosis. The content of leukocytes in the blood decreased by 12.1%, 19.5%, 26%, granulocytes, monocytes and neutrophils increased, respectively, by 8.6%, 3.8%, 3.9%.

Analysis of daily blood showed that the content of leukocytes in the blood began to decrease, tending to intact values. The content of erythrocytes in the blood, on the contrary, during the entire study period increased by 2.5%, 4.2% and 8.5%, respectively. The content of red blood cells in the blood increased by 17.5% after 72 hours. The changes are statistically significant at P<0.05. With the combined use of viper venom and manganese nanoparticles, inhibition was observed in the development of tumor cells, which contributed to the partial recovery of experimental animals.

Subsequently, the condition of the animals improves. Data from laboratory studies confirmed that after the introduction of a complex mixture of viper venom with manganese nanoparticles, all determined blood parameters of experimental rats practically reached intact values after 72hours.

We found that 72 hours after the start of incubation with manganese nanoparticles, the viability of tumor cells significantly decreased, which was confirmed by changes in the data, the content of leukocytes in the blood decreased by 23%, lymphocytes increased by 9.5%, granulocytes, monocytes and neutrophils decreased, respectively, by 5.7%, 0.9%, 1.2%.

The results of experimental studies have shown that significant changes occur in the content of formed elements in the blood, which do not pose a threat to the body and prevent the development of tumor cells This study may be useful for the development of new therapy and diagnosis of malignant neoplasms. However, at present, there is no data on clinical studies of biologically active proteins of viper venom together with manganese nanoparticles that affect the elementary composition of the blood and thereby prevent the proliferation of tumor cells. Thus, in conclusion, it should be noted that, despite the existing achievements and shortcomings, a number of peptides and proteins of viper venom that affect the blood have good prospects as a basis for the creation of new drugs with anticancer activity.

The results of laboratory studies showed that significant changes occur in the content of leukocytes and erythrocytes in the blood, which do not pose a threat to the animal’s body. Intraperitoneal administration of microdoses of viper venom mixed with manganese nanoparticles leads to quantitative changes in the composition of blood cells. Thus, intraperitoneal administration of a mixture of viper venom and manganese nanoparticles to white laboratory rats helps to normalize the composition of blood cells and stops the process of growth and reproduction of tumor cells and thereby eliminates the threat of tumor growth.

CONCLUSIONS:

1. It was experimentally revealed that after intravenous administration of viper venom at a dose of 0.01mg/kg body weight with manganese nanoparticles and exposure to a magnetic field, heating to a temperature of 42℃, after 1hour and 24hours and 72hours a change in percentage was noted the content of leukocytes in the blood of experimental animals. The content of leukocytes in the blood decreased by 12.1%, 19.5%, 26%, granulocytes, monocytes and neutrophils increased, respectively, by 8.6%, 3.8%, 3.9%.

2. It was found that after intravenous administration of viper venom at a dose of 0.01mg/kg body weight with manganese nanoparticles and exposure to a magnetic field, heating to a temperature of 42℃, after 1 hour and 24 and 72hours there was a change in the percentage of red blood cells. The content of erythrocytes in the blood during the entire study period increased by 2.5%, 4.2% and 8.5%, respectively.

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Received on 31.07.2024 Revised on 12.10.2024

Accepted on 19.12.2024 Published on 08.03.2025

Available online from March 12, 2025

Res.J. Pharmacology and Pharmacodynamics.2025;17(1):13-18.

DOI: 10.52711/2321-5836.2025.00003

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