RU2348416C2 - Composition with immunopotentiating properties - Google Patents

Abstract

FIELD: medicine; pharmacology.

SUBSTANCE: substance of technical decision seeks to produce immunopotentiating composition based on nanotechnology products, specifically fullerenes and superdispersed diamond powders (nanodiamonds) produced by detonation synthesis. Composition contains modified fullerene of general formula C60-X, where C60 is fullerene nucleus, X is formylporphyrin, as well as superdispersed diamond powder in ratio as follows, wt %: porphyrin modified fullerene C60 - 50; superdispersed diamond powder - 50.

EFFECT: synergetic effect of immunopotentiating components activity with simultaneous considerable reduction in price concerning prototype composition.

2 tbl, 1 dwg

Description

The invention relates to the field of immunology and may find application in medicine and biotechnology as a means to stimulate the functional activity of immunocompetent cells and to correct immunodeficiency states of the body.

Nanomaterials are known, including fullerene-60 modified with porphyrin and exhibiting biological activity [V.M. Elinson et al. Synthesis of new medical materials based on polymers with a nanostructured surface modified with C60 organic derivatives. Materials of the 12th scientific and technical conference "Vacuum Science and Technology". Sudak, Crimea, Ukraine, 2005, p.183-187].

The known composition has a pronounced biological activity, but it has not been identified and not used independent immunostimulating properties of fullerene.

The biological activity of ultrafine diamonds is known (Pat. RF No. 2203068. Biologically active ultrafine diamonds of detonation synthesis, A61K 33/44. Publ. 2003.04.27), however, their immunomodulating properties are not found in combination with other biologically active substances.

The use of fullerenes and their modifications in the manufacture of vaccines is known, for example, the biological activity of a composition comprising fullerene C60 (Pat. RF No. 2184566, Method for producing a vaccine. IPC 7 A61K 39/00, A61K 38/05. Published 2002.07.10, accepted prototype).

In the known composition, fullerene enhances the effect of antigen (extract from microbial biomass), but its immunomodulating properties are insufficient.

An object of the invention is the creation of an immunostimulating composition based on nanotechnology products, namely, the preparation of a preparation having immunobiological activity based on fullerenes with a nanostructured surface (NSP) and ultrafine diamond powders (nanodiamonds) obtained by detonation synthesis. At the same time, it becomes possible to modify the fullerene core with various porphyrins, which, as you know, easily form extremely strong complexes with a number of elements (iron, copper, cobalt), for example formylporphyrin, hemoglobin, cobalamin (vitamin B ₁₂ ), etc.

The problem is solved by using a new composition not previously used: a mixture of fullerene and ultrafine diamond powders in an effective quantitative ratio, in particular in a percentage ratio of 50:50.

Both preparations are finely divided powders capable of long-term storage.

The identified composition includes a modified fullerene of the general formula C60-X, where C60 is the fullerene core, X is porphyrin, as well as ultrafine diamond powder in the following components, wt.%:

- fullerene modified with porphyrin - fifty; - ultrafine diamond powder - fifty.

The identified composition has a synergistic effect of immunostimulating activity of the components while significantly reducing the cost relative to the composition of the prototype.

Thanks to modern methods of cleaning detonation UDDs, they are relatively cheap (Pat. RF No. 2083490, Method for the separation of ultrafine diamonds from an aqueous suspension, 1997; Pat. RF No. 2109683, Method for the separation of synthetic ultrafine diamonds, 1998).

An analysis of the specialized literature did not reveal the work of other authors on the fact of stimulating the functional activity of immunocompetent cells using modified fullerene, in particular in combination with nanodispersed diamonds.

The question of the optimal ratio of components requires special studies to identify the synergistic effect of the special physicochemical properties of this group of carbon structures on the functioning mechanism of cellular biosystems.

So far, it can be argued that the optimal range of the ratio of the claimed components in each case will depend on the type of modifier, the degree of modification of the structure, the method of producing nanodiamond powder, the degree of its purification and other factors.

The results of studies of the above mixtures in various ratios, in particular 50:50, showed high immunomodulating properties of the claimed composition and the presence of immunobiological activity, as illustrated by examples.

Example 1. To the powder of fullerene modified with porphyrin, was added the powder of ultrafine diamonds (UDD) obtained by detonation synthesis, with a particle size of 2 to 7 nm, in a percentage weight ratio of 1: 1. The composition was tested for immunobiological activity.

The effect of the composition on immunocompetent cells was tested in the reaction of blast transformation of human lymphocytes stimulated by phytohemagglutinin (PHA) after the action of drugs in the concentration range (1.5-15) · 10 ^-3 mg / ml. Evaluation of the effect of drugs on the proliferative activity of lymphocytes in a spontaneous and induced PHA test was carried out in accordance with the methodological recommendations of the Pharmaceutical Committee of the Ministry of Health of the Russian Federation [Vedomosti Pharmacological Committee, 1999, No. 1], as well as in the standard test of cytotoxicity by trypan blue staining.

The immunomodulatory activity of the composition according to example 1 is presented in the attached graph.

The graph clearly shows that the claimed composition in the studied concentrations has a stimulating effect on the spontaneous activity of lymphocytes in the range of 30-55% relative to the prototype and up to 120% relative to the control. In addition, it is seen that the claimed composition does not inhibit the PHA-induced proliferative activity of lymphocytes.

We conducted additional studies of the cytotoxic effect of the claimed composition in a concentration of (1.5-15) · 10 ^-3 mg / ml on human lymphocytes (average data for 6 observations), as a result of which it was found that the composition does not have a cytosuppressive effect on immunocompetent cells (see table 1).

Studies were also conducted on the effect of the composition on the functional activity of neutrophilic peripheral blood leukocytes from donors in a wide range of concentrations (0.15-15) · 10 ^-3 mg / ml. A well-known literature method for modeling phagocytosis using the Staphylococcus aur strain was used. H-209 (Novikov D.K., Novikova V.N. Assessment of the immune status. Moscow-Vitebsk, 1996, 281 pp.). The percentage of active neutrophils (PAN) and the percentage of completed phagocytosis (PZP) were calculated in the control test without drug exposure. The results are presented in table 2, which shows that after exposure to the composition increases the percentage of active neutrophils and their phagocytic activity in comparison with the prototype, which is important for immunotherapy and correction of immunodeficiency states of the body.

The literature does not contain information on the use of the claimed compositions in clinical practice, in particular, their immunomodulating properties are not currently known from the prior art, which indicates the novelty of the technical solution, and the non-obviousness of the immunomodulating activity of the composition suggests that its criterion is “inventive step”.

Literature

1. V.M. Elinson et al. Synthesis of new medical materials based on polymers with a nanostructured surface, modified with organic derivatives of C60. Materials of the 12th scientific and technical conference "Vacuum Science and Technology". Sudak, Crimea, Ukraine, 2005, p. 183-187.

2. Pat. RF №2203068. Biologically active ultrafine diamonds of detonation synthesis, А61К 33/44. Publ. 2003.04.27.

3. Pat. RF №2184566, A method of obtaining a vaccine. IPC 7 A61K 39/00, A61K 38/05. Publ. 2002.07.10, taken as a prototype.

4. Pat. RF №2083490, The method of separation of ultrafine diamonds from an aqueous suspension, 1997; Pat. RF №2109683, The method of separation of synthetic ultrafine diamonds, 1998.

5. Sheets of the pharmacological committee, 1999, No. 1.

Table 1
The results of determining the cytotoxic effect of the claimed composition on lymphocytes The percentage of viable cells after 24 hours of incubation (%) No drugs Prototype The claimed composition 97 90 95

table 2 Dose, 10 ^-3 mg / ml % PAN % PZF Prototype Composition Prototype Composition the control 34 34 fifty fifty 0.15 45 47 31 70 1,5 49 51 45 71 fifteen 37 35 51 54

Claims (1)

A composition having immunostimulating properties, including modified fullerene of the formula C60-X, where C60 is the fullerene core, X is formylporphyrin, and ultrafine diamond powder with the following components, wt.%:
porphyrin modified fullerene fifty ultrafine diamond powder fifty