RU2046340C1 - Method for selecting medicinal preparations possessing anticancerogenic activity - Google Patents

Abstract

FIELD: medicine. SUBSTANCE: method involves studying relationship between biological object survival and interinjection interval for a preparation under study. Selection is performed for preparations showing resonance type relationships with respect to the mentioned parameters and shown therapeutic effectiveness of the preparation when used in modes including periodical application of the preparation with period provided maximum of the resonance function. EFFECT: enhanced selectivity of the method. 3 dwg

Description

 The invention relates to medicine, in particular to experimental medicine, and can be used in the study of the pathogenesis of cancer, in the development of effective therapeutic courses of new chemicals exhibiting cytotoxic activity, as well as in the study of the mechanisms underlying the restoration of the functional state of the body and associated with violation of the cell cycle.

 It is known that the cells of the tumor tissue and normal tissue (not affected by the disease) are characterized by different average generation times. The latter allows quite effective use in the treatment of tumors of a whole class of cyclo- and phase-specific drugs. However, the chemicals used are typically highly toxic. The most vulnerable to cytostatics are cells of regenerating tissues (for example, hematopoietic and small intestine tissue).

 The difficulty in correcting the functional state of the body, which would extend the time of observation of the experimental animal during its “recovery” period, lies not only in choosing an adequate drug, but also in optimizing the procedure for its administration: in choosing the course of exposure with the drug used.

 Particular hopes are associated with the optimization of the administration regimen of cyclically specific and phase-specific cytotoxic agents. Unfortunately, the regulation of the proliferative activity of normal tissues and tumors is poorly studied, and for this reason the problem has not yet been found a satisfactory solution.

 The proposed method essentially has no analogues in technical solution. It is based on the use of periodic exposure courses with a phase-specific cytotoxic drug with an adequately selected exposure period. Theoretical calculations show that with phase-specific effects with a period close to the average or doubled average cycle length of actively proliferating cells of critically normal tissue, it is possible to significantly increase the efficiency of theration (correction) and achieve selective destruction of tumor cells. This effect is based on reliably established normal fast-renewing tissues of tumor cells and actively proliferating cells of various durations of the average cell cycle and is associated with a significant "resonant" decrease in damage to normal tissues using the above effects (hereinafter, under the term "normal tissue critical for a given preparation) "refers to normal tissue, limiting the use of this drug, ie tissue, the defeat of which under the influence of the drug leads to ultimately to the death of the body).

 The described model was adapted to the conditions of the task set in this application and verified experimentally. It turned out that under the conditions of the experiment, the high efficiency of the functioning of the biological model (experimental oncology) can be achieved, which suggests the high efficiency of the proposed approach in the practice of treating cancer patients.

 The known method by which an attempt is made to optimize the mode of administration of phase-specific cytotoxic drugs by their repeated administration. However, the result of the studies described cannot be considered satisfactory enough, since the treatment courses are accompanied by large side effects due to the high toxicity of the drugs to critical normal rapidly renewing tissue.

 This is due to the lack of a causal relationship between the treatment regimen and the appropriate mechanism of action of the drug.

 The purpose of the invention is to increase the efficiency of the method by reducing the toxicity of the drug in relation to normal cells (cells not affected by pathology).

режим Б; + режим С; Δ контрольная группа. На фиг. 5 торможение роста опухоли меланомы в-16 (а) и кинетика гибели мышей (б) при различных режимах введения АРА-Ц. По осям абсцисс и ординат.In FIG. Figure 1 shows the dependence of the survival rate of CFUs of mice (CBAx57BL) F1 on the period of 6-fold administration of OM (single dose 1 g / kg). On the abscissa axis, the period of OM administration (h), on the ordinate axis, the number of surviving CFUs (% of the initial level). Different symbols correspond to independent experiments; confidence intervals with significance level p <0.3 are indicated. The solid line corresponds to the calculated dependence. In FIG. 2 the dependence of the defeat of the epithelium of the small intestine of mice (CBAx57BL) F1 on the period of 8-fold introductions of OM (0.25 g / kg). On the abscissa, the period of the introduction of OM (h); along the ordinate axis: and the number of crypts per section of the intestine; b the total number of crypt enterocytes per section of the intestine (x10); in the number of cell positions on the villi. M + m (m average error) are indicated 30 hours after the end of OM administration. The ranges M ± m are shaded for control animals. For control animals, the number of cell positions on the villi is 108 ± 6. The calculated curves are indicated by a solid line. In FIG. Figure 3 shows the survival of mice (CBAx57BL) F1 versus the period of OM administration (single dose 0.25 g / kg). On the abscissa, the period of administration of OM; along the ordinate axis, the number of surviving mice (% of baseline); Δ 8 injections; · 6 injections. In FIG. 4 inhibition of tumor growth of Lewis lung cancer (a) and the kinetics of mouse death (b) under various modes of administration of ARA-C. On the abscissa axis, days after tumor inoculation, on the ordinate axis, the tumor volume in cm ³ (a) and the number of live mice (b). Tumor volume was calculated as the product of three independent linear sizes. · Mode A;

mode B; + mode C; Δ control group. In FIG. 5 inhibition of tumor growth of melanoma B-16 (a) and the kinetics of death of mice (b) under various modes of administration of ARA-C. On the axes of abscissa and ordinate.

 PRI me R 1. Resonance effect for early hematopoietic predecessors. In the experiments, the dependences of the survival of mouse hematopoietic precursors (CBA57BL line F1), forming colonies in the spleen (CFUc), on the period of 6-fold injections of high doses (single dose 1 g / kg) of oxyurea (OM) were investigated.

 The experimental results are shown in FIG. 1.

 According to the results obtained, there is a pronounced resonance effect with a resonance maximum of CFU survival during the administration of OM with a period of 12 hours. It should be noted that earlier attempts to evaluate the cycle parameters of these unidentifiable directly cells did not lead to the desired results.

 PRI me R 2. Resonance effect for the epithelium of the small intestine. In another series of experiments, the dependence of cell damage on the period of OM injection was studied for the second “critical” tissue epithelial tissue of small intestine for phase-specific cytotoxic agents. In the experiments used mice line (CBAx57BL) F1.

 The state of the epithelium was evaluated by morphometric indices on formalin-fixed sections of the small intestine. The defeat of various cellular epithelial compartments corresponding to cells of different stages of differentiation (from stem to functionally mature) was controlled. For all cell compartments, clear resonance dependences of cell survival on the period of OM administration were revealed with survival maxima corresponding to OM administration with periods close to 8 or 16 hours.

 Thus, it was found that the resonance effect is manifested not only for individual cell populations, but also at the level of renewed tissue.

 The results of this series are presented in FIG. 2.

 Data on the number of surviving crypts reflect the survival of stem (cryptogenic) cells, the total number of crypt enterocytes corresponds to the number of proliferating epithelial cells, the number of cell positions on the villi is a generally accepted indicator of the number of non-proliferating functionally mature enterocytes.

 For a more complete understanding of the dependence of enterocyte damage on the period of OM management, morphometric data were supplemented by electron microscopic studies of the ultrastructure of villus enterocytes. According to the data obtained, this dependence is actually even more contrasting than it follows from the indicators reflected in FIG. 2.

 PRI me R 3. The dependence of the survival of mice (CBAx57BL) F1 from the period of administration of oxyurea. In this series of experiments, the correctness of the transition from organism survival to damage to one tissue and vice versa was confirmed.

 Mice were injected with a single dose of 0.25 g / kg of oxyurea.

 According to the results obtained, the survival of mice in a resonant manner depends on the period of OM administration, and when the period varies, it changes in phase with the survival of the epithelium of the small intestine and cryptogenic cells of this tissue (see Fig. 3).

 Similar resonance dependences were obtained for the survival of mice in the case of the S-phase-specific agent cytosine-arabinoside (ARA-C), which is widely used in antitumor therapy, as shown in examples 4 and 5.

 PRI me R 4. Inhibition of tumor growth of Lewis lung cancer and the kinetics of mouse death. We used BDF1 mice (males). The animals were intraperitoneally injected with ARA-C in saline. Three modes of administration were used. Mode A 3 courses of 4 injections of ARA-C at a dose of 75 mg / kg every 11 hours, the interval between courses is 3.5 days. Mode B every 3 hours, 8 times at 16 mg / kg on the 2nd, 6th, 10th and 14th day. Mode C 5 injections of ARA-C at 75 mg / kg every 24 hours and 3 injections of ARA-C at 75 mg / kg every 24 hours (the interval between courses is 4.5 days). As control was used animals that were not exposed to the drug.

 All courses began in 2 days. after inoculation, the tumors had approximately the same duration (11–13 days) and showed toxicity close to LD 10, with minimal toxicity observed for regime A.

 The results are presented in FIG. 4.

 The average lifespan of Lewis lung cancer tumor-bearing mice was: for control animals, 30.0 + 1.1 (hereinafter, mean values and mean error per day); with mode A 45.0 + 1.4; with mode B 34.7 + 2.0; in mode C 29.9 + 1.6. From here and from FIG. 4 it follows that injections of ARA-C with a period corresponding to a resonant decrease in side toxicity contribute to the manifestation of increased antitumor efficacy of the drug, which is manifested both in inhibition of tumor growth and in lengthening the life time of mice.

 PRI me R 5. Inhibition of tumor growth of melanoma B-16 and the kinetics of the death of mice with various injections of ARA-C.

 In an additional series of experiments, the effect of (ARA-C on the growth of a tumor of B-16 melanoma vaccinated in mice of the BDF1 line (males) and the kinetics of the death of mice under various modes of administration of this drug were investigated.

 The administration modes A and B described in the previous example were used.

 The results are presented in FIG. 5.

 The average lifespan of B-16 melanoma tumor-bearing mice was: for control animals, 24.2 + 1.5; with mode B 33.8 + 3.1; with mode B 26.3 + 1.8.

 Thus, the result obtained fully confirms the conclusion made in the previous example.

 The proposed method for the correction of the functional state allows to achieve a significant reduction in the toxicity level of the used anti-cancer drugs.

Claims (1)

 METHOD FOR CORRECTION OF THE FUNCTIONAL CONDITION OF THE ORGANISM WITH ONCOLOGICAL DISEASE, including a course of drug exposure, characterized in that the drug effect includes courses of periodic drug administration with a period that provides a resonant maximum survival, critical for this drug to average tissue, or corresponding to the average normal tissue or appropriate the duration of the cycle of actively proliferating cells of critical normal tissue and is determined either independently data on the duration of the cycle of these cells, or based on the results of a study of the dependence of the damage of critical normal tissue on the period of multiple periodic injections of the drug.

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