RU2729788C2 - Agent, use thereof and a method of increasing resistance of a mammalian organism to ischemic-reperfusion involvement of the small intestine - Google Patents

Abstract

FIELD: medicine.SUBSTANCE: group of inventions relates to an agent for treating ischemic-reperfusion injury of the small intestine, characterized by that it contains a pharmaceutical composition of preparations, consisting of recombinant protein PSH, stabilizer and pharmaceutically acceptable solvent, where components are taken in following ratio, wt %: recombinant protein PSH—0.1–0.4, trehalose stabilizer—0.01–0.04, pharmaceutically acceptable solvent—99.89–99.56, also relates to a method for increasing resistance of mammals to ischemic and reperfusion injures of the small intestine.EFFECT: group of inventions provides preparing an agent which enables increasing intestinal epithelial cell resistance to ischemia reperfusion injury, and the method enables extending the spectrum of use of the agent for reducing organ damage in reperfusion on in vivo models.7 cl, 3 dwg, 5 ex, 1 tbl

Description

Technology area

The invention relates to medicine, in particular to experimental surgery and can be used to correct ischemic and reperfusion injuries of the small intestine. The proposed therapeutic agent and the method of its use can be used in the treatment of pathologies of the small intestine, which are accompanied by ischemia-reperfusion syndrome, including occlusion of the splash vessels, strangulation small bowel obstruction, etc.

State of the art

Ischemic-reperfusion injury of the small intestine plays a fundamental role in various clinical conditions. Most of the cardiac and vascular interventions, occlusion of the mesenteric vessels, postoperative complications, resolution of strangulation obstruction and bowel transplantation are associated with this pathology [Timerbulatov Sh.V. et al., 2010; Yandza T., et al., 2011].

One of the reasons for the defeat of the small intestine in ischemia-reperfusion injury is powerful oxidative stress, and ROS overproduction is the main factor that triggers the mechanism of apoptosis, necrosis and cell damage [Granger D.N. et al., 2015]. Among the internal organs, the small intestine is most sensitive to ischemia-reperfusion injury. In conditions of hyperproduction of reactive oxygen species (ROS), the intestinal antioxidant system is not able to fully cope with the normalization of free radical processes [Menytsikova E.B. et al., 2008]. In this case, one of the ways to reduce the excess of reactive oxygen species is to increase the antioxidant status in the damaged tissue by using exogenous antioxidant enzymes and other antioxidant agents.

Known methods for reducing ischemia-reperfusion lesion of the small intestine in animals in the experiment by preliminary intravenous administration of antioxidant agents before the induced lesion. Allopurinol, remifentanil, lazaroid (21 - amino steroid) U-74389G, selenium, infiximab and others are considered as such agents [Sapalidis K. et al., 2013; Cho S.S. et al., 2013; Flessas I. et al., 2015; Kim Y. et al., 2014; Pergel A. et al., 2012]. Antioxidant agents are inferior in efficiency to antioxidant enzymes, since they have an indirect mechanism of action on the balance of the rare system.

Known methods of using direct inhibitors of ROS overproduction - propofol and tempol to reduce the level of ischemic and reperfusion lesions of the small intestine [Gan X., 2015, Zhang G. et al., 2016] The main drawback of these agents is their low substrate specificity for ROS.

Known methods for the complex use of antioxidant agents. To reduce ischemic-reperfusion damage to organs, an antioxidant composition including sodium ascorbate, N-acetylcysteine and deferoxamine is proposed. The composition is intended for infusion therapy [US Patent No. US 20170333393, 2017]; a composition based on allopurinol, oxypurinol and deferoxamine for intravenous administration in a complex of resuscitation measures [US Patent No. US 4978668,1990].

In clinical practice, attempts are known to use low molecular weight antioxidants Mexidol [Bagov AN, 2009] and Reamberin for antioxidant therapy in ischemia-reperfusion syndrome [Kurbonov KM, 2013]. The main disadvantage of low molecular weight antioxidants, which are actively introduced into the clinic, is their low antioxidant activity; they are significantly inferior in efficiency to antioxidant enzymes.

To effectively reduce ischemia-reperfusion lesions of the small intestine, an agent with a broad substrate specificity for ROS and high antioxidant activity is required.

The antioxidant enzyme peroxiredoxin 6 is suitable for these characteristics. Recombinant human peroxiredoxin 6, in contrast to other antioxidant enzymes, has a wide substrate specificity, since it is capable of neutralizing the entire spectrum of hydroperoxides (inorganic and organic hydroperoxides, lipid peroxides, peroxynitrites). It has a pronounced antioxidant activity (peroxidase activity for H2O2 - 90 nmol / mg / min and for tert-butyl peroxide - 60 nmol / mg / min). The key feature is that the peroxidase activity of Prx6 is manifested at low peroxide concentrations (1-100 μM), at which other antioxidant enzymes are ineffective.

The effectiveness of the use of pharmacological compositions with recombinant peroxiredoxin 6 for the treatment of diseases associated with overproduction of free radicals caused by exogenous and endogenous factors has been shown [RF Patent No. 2280448, 2006]. However, this technical solution is not associated with the use of peroxiredoxin 6 in ischemia-reperfusion lesions of the small intestine.

The well-known work of M.G. Sharapov et al., Which considers the use of recombinant peroxiredoxin 6 in the treatment of ischemia-reperfusion lesions of the small intestine. The work shows that intravenous administration of exogenous peroxiredoxin 6 before ischemia-reperfusion leads to minimization of tissue damage and a decrease in apoptotic death of intestinal cells and mesenteric vessels [Sharapov M.G. et al., 2017].

, наряду с перекисью водорода (Н₂О₂) относиться к потенциально опасным АФК. Для детоксикации потенциально опасных АФК:

и Н₂О₂ в организме существуют супероксиддисмутазы и пероксидазы, которые объединяют в первую линию защиты от АФК [Ланкин В.З. и др., 2001]. С учетом этого факта, для максимальной защиты тонкого кишечника от поражающего действия АФК и торможения сигнальных каскадов, вызванных гиперпродукцией АФК необходим агент, объединяющий свойства супероксиддисмутазы и пероксидазы. В качестве такого агента возможно использование фермента-антиоксиданта PSH (Prx VI-SOD) с совмещенной пероксидазной и супероксиддисмутазной активностями. Раскрыт способ получения предложенного белка путем культивирования клеток штамма E.coli BL21(PSH), трансформированных сконструированным рекомбинантным вектором экспрессии на основе плазмиды pET22b(+). Изобретение позволяет получить высокой выход белка PSH, обладающего высокой антиоксидантной активностью [Патент РФ №2534348, 2014].However, recombinant peroxiredoxin 6, possessing peroxidase activity, is unable to neutralize the superoxide radical. Superoxide radical

, along with hydrogen peroxide (Н ₂ О ₂ ), belong to potentially dangerous ROS. To detoxify potentially dangerous ROS:

and H ₂ O ₂ in the body there are superoxide dismutases and peroxidases, which combine in the first line of defense against ROS [Lankin V.Z. et al., 2001]. Taking this fact into account, an agent combining the properties of superoxide dismutase and peroxidase is needed for maximum protection of the small intestine from the damaging action of ROS and inhibition of signaling cascades caused by overproduction of ROS. An antioxidant enzyme PSH (Prx VI-SOD) with combined peroxidase and superoxide dismutase activities can be used as such an agent. A method for producing the proposed protein by culturing cells of the E. coli BL21 (PSH) strain transformed with a constructed recombinant expression vector based on the pET22b (+) plasmid is disclosed. The invention makes it possible to obtain a high yield of the PSH protein with high antioxidant activity [RF Patent No. 2534348, 2014].

The closest prototype analogue is the use of an antioxidant enzyme PSH (Prx VI-SOD) with combined peroxidase and superoxide dismutase activities in the composition of blood replacement solutions for reperfusion of an isolated heart (ex vivo model). Reperfusion with blood-substituting solutions containing PSH protein is necessary to maintain the functioning of an isolated organ outside the body [RF Patent No. 2534348, 2014]. The disadvantage of this method of use is that the use of PSH only for isolated organs does not reveal its therapeutic efficacy in the whole organism.

The object of the invention is to provide a means and a more effective method for the combined treatment of ischemic and reperfusion injuries of the small intestine of a mammal.

The technical result consists in the fact that the claimed agent makes it possible to increase the resistance of intestinal epithelial cells to ischemia-reperfusion injury. The claimed method of application makes it possible to expand the range of use of the agent for reducing organ damage during reperfusion in in vivo models.

The essence of the invention

The subject of the present invention is an agent for the treatment of ischemia-reperfusion injury, which contains a pharmaceutical composition of preparations consisting of a recombinant PSH protein, a stabilizer and a pharmaceutically acceptable solvent, where the components are taken in the following ratio, wt. %:

recombinant PSH protein 0.1-0.4 stabilizer 0.01-0.04 pharmaceutically acceptable solvent 99.89-99.56

The agent contains trehalose as a stabilizer.

Another object of the present invention is a method for increasing the resistance of the mammalian organism to the action of ischemic and reperfusion injuries of the small intestine. According to the method, a therapeutically effective amount of the agent is administered to the mammal intravenously, once, 15-20 minutes before the induced ischemia of the mammal, at a dose of 5 mg / kg to 20 mg / kg of the mammalian body weight. Preferably at a dose of 10 mg / kg body weight.

Another object of the present invention is the use of the agent as a preparation for the treatment of ischemia-reperfusion injury of the small intestine.

List of figures

FIG. 1. Distribution of exogenous PSH (Prx VI-SOD) in the tissues of the rat small intestine after intravenous administration.

FIG. 2. Histostructure of the small intestine in different experimental groups. Group 1 (period of ischemia 1 hour, period of reperfusion 2 hours, no treatment); group 2 (period of ischemia 1 hour, reperfusion period 24 hours, no treatment); group 3 (ischemic period 1 hour, reperfusion period 2 hours, using PSH (Prx VI-SOD); group 4 (ischemic period 1 hour, reperfusion period 24 hours, using PSH (Prx VI-SOD). In groups 3 and 4, the architecture of the intestinal wall is preserved Microphoto × 100.

FIG. 3. The level of the marker of apoptotic cell death caspase-3 in the intestinal tissue in different experimental groups. The control (the level of caspase-3 in the intact intestine) was taken as 100%. Group 1 (period of ischemia 1 hour, period of reperfusion 2 hours, no treatment); group 2 (period of ischemia 1 hour, reperfusion period 24 hours, no treatment); group 3 (ischemic period 1 hour, reperfusion period 2 hours, using PSH (Prx VI-SOD); group 4 (ischemic period 1 hour, reperfusion period 24 hours, using PSH (Prx VI-SOD).

Description of the invention

When studying the effective composition of the composition for intravenous administration to reduce complications after ischemia-reperfusion injury of the small intestine, it was found that the use of the recombinant PSH protein led to an increase in the resistance of cells to ischemia-reperfusion injury under conditions of normalization of the activity of the endogenous antioxidant system.

The new composition of the composition includes an agent containing a solution containing at least one antioxidant used in a therapeutically effective dose.

The therapeutically effective dose of PSH is determined in accordance with the data on the effective concentration of PSH in the protection of biomacromolecules from reactive oxygen species in vitro, as well as in accordance with the concentration of exogenous peroxiredoxin Prx VI in the intestinal tissue in normal conditions. The range of therapeutically effective doses of PSH in liquid form of the drug can vary depending on the severity of the lesion and range from 5 mg to 20 mg protein per kg of body weight.

The retention of the structure and activity of PSH is a fundamental point of this invention. To prevent a decrease in the activity of the protein during its storage, a method of stabilizing and lyophilizing a protein solution is used.

A lyophilized preparation for preparing a liquid composition using PSH (Prx VI-SOD) and stabilizers has the following composition (wt%):

PSH (Prx VI-SOD) - about 90.0;

Stabilizer - about 10.0

Immediately before use, the dry powder of lyophilized PSH is dissolved in saline at the desired concentration. The solutions are used immediately.

The basis of the pharmaceutical composition included in the product contains the following ratio of components, wt. %

Antioxidant, recombinant PSH protein 0.1-0.4

Trehalose stabilizer - 0.01-0.04

Pharmaceutically acceptable solvent 99.89-99.56

Intravenous administration, single. As solvents PSH use nat. solution and / or other acceptable solvents.

It is possible to use a composition comprising a recombinant PSH protein and a stabilizer trehalose in conjunction with at least one broad-spectrum therapeutic agent. The therapeutic agent used should not inhibit the enzymatic activity of Prx VI-SOD, either peroxidase or superoxide dismutase.

The examples below include, but are not limited to, the use of analogs of the active ingredients included in the product.

The proposed method for the treatment of ischemia-reperfusion lesions of the small intestine may be applicable to different areas of prevention and therapy of the small intestine in ischemia-reperfusion syndrome and include:

A) prophylactic protection of the small intestine and other organs from the overproduction of free radicals during cardiovascular interventions, for example, when simulating myocardial revascularization operations and during surgical interventions on the aorta, with its clamping above the celiac trunk.

B) therapeutic protection of the small intestine from pathological changes in acute forms of abdominal pathologies, for example, when resolving strangulation small bowel obstruction, in acute disturbance of mesenteric circulation.

B) preventive protection of the small intestine to reduce postoperative complications in case of failure of gastrointestinal anastomoses, intra-abdominal hypertension syndrome, hypovolemic shock, etc.

D) preservation of the viability of organs, including the small intestine and its segments, under conditions of extracorporeal perfusion in the body of an asystolic donor before the beginning of the explantation operation.

The method is carried out as follows.

Example 1

Experiments are carried out on white Wistar male rats weighing 200-220 g. Modeling of ischemia-reperfusion injury is carried out under general anesthesia. A mixture of zoletil® 100 (Zoletil® 100, Virbac Same Animale, France) and rometar (Rometar, Bioveta, Czech Republic) is used as anesthesia. Mixture. administered intravenously. In an anesthetized rat, the abdominal cavity is opened 1/3 below the diaphragm by a midline laparotomic incision and access to the superior mesenteric artery and vein is performed. The superior mesenteric artery occlusion model is performed by applying a vascular clamp to the superior mesenteric artery for 60 minutes. Recombinant PSH (Prx VI-SOD) is administered intravenously in liquid form 15-20 minutes before induced ischemia at a dose of 10 mg of protein per kg of body weight. After 2 and 24 hours of reperfusion, intestinal tissue is collected for research. Studies include histological assessment of tissues, enzyme-linked immunosorbent assay and determination of the state of the endogenous antioxidant system by the level of gene expression of antioxidant enzymes by RT-PCR.

Example 2 Obtaining and preparing PSH-based agent (Prx VI-SOD)

Within the framework of the present invention, a bifunctional PSH peptide is obtained from cells of the E. coli BL21 (PSH) strain transformed with a constructed recombinant expression vector based on the plasmid pET22b (+) (RF patent No. 2534348, 2014)

The presence of the His-tag in the structure of the PSH protein simplifies the procedure for the purification of the recombinant protein due to the fact that this domain specifically binds to Ni-NTA-agarose.

E. coli BL21 (PSH) bacteria expressing PSH (Prx VI-SOD) are resuspended and destroyed by ultrasound on an UZDN-2 generator at 4 ° C. Cell debris is removed by centrifugation at 14000 rpm for 10 min, the supernatant is filtered through a 0.45 μm filter (Corning, USA) and applied to a 10 ml column (BioRad, USA) with Ni-NTA agarose (Invitrogen, USA) which has been previously equilibrated with the application buffer. The incubation of the bacterial lysate with Ni-NTA-agarose is carried out for 40 min at + 4 ° C. Then, the column is washed with 100 ml of a solution: 12 mM Tris-HCl pH 7.8, 20 mM imidazole. The protein is eluted with 5 ml of buffer: 12 mM Tris-HCl, pH 7.8, 250 mM imidazole. The protein is concentrated using a VIVASCIENCE 30.000 MWCO membrane concentrator (Sartorius, Germany) and dialyzed against 1 × PBS (1.7 mM KH ₂ PO ₄ , 5.2 mM Na ₂ HPO ₄ , 150 mM NaCl, pH 7.4). The protein purity, assessed by polyacrylamide gel electrophoresis, was about 95%. Typically, about 20-40 mg of purified protein is obtained from one fermentation. The resulting protein is used for further experiments.

The obtained PSH (Prx VI-SOD) can be used in liquid form, however, the protein is unstable during long-term storage in solutions, and after repeated freezing / thawing, it loses its activity. Therefore, the recombinant protein is lyophilized and stabilized in order to maintain the activity of the protein during storage. Lyophilized forms can be used in the future for the preparation of liquid forms for introduction into the blood.

Lyophilized PSH preparation (Prx VI-SOD) is obtained by a technology that includes at least 4 stages: 1) protein purification using affinity chromatography on Ni-NTA agarose; 2) concentration of fractions to a concentration of 5-7 mg / ml containing PSH (Prx VI-SOD) and dialysis against deionized water 3) introduction of at least one stabilizer; 4) the process of lyophilization at -20 ° C until a dry powder is formed.

Stabilizers are necessary to maintain PSH (Prx VI-SOD) activity for a longer period, at least one year, and in more accessible conditions, at temperatures from +4 to -20 ° C, depending on the shelf life. The lyophilized preparation is packaged in vials sterilized in an autoclave at + 120 ° C (depending on the application, in doses from 1 to 5 mg and stored at -20 ° C).

It is known that trehalose performs the function of preserving the secondary structure of peptides and proteins [Singer M.A. et. al, 1998]. The use of trehalose as a stabilizer includes, but does not limit the use of other stabilizers. Stabilizers can be selected from the group consisting of: a) one or more mono- or polysaccharides or sugar alcohols (maltose, glucose, lactose, trehalose, sucrose, mannitol, inositol, galactose, ribose, xylose, mannose, sucrose, cellobiose, raffinose and maltotriose, b) amino acids (alanine, glycine, etc.), c) polypeptides or proteins (albumin).

Due to the fact that PSH (Prx VI-SOD) is a highly soluble protein, for the direct preparation of a liquid form of the drug from a lyophilisate of protein for intravenous administration, nat. solution, Ringer's solution and other balanced salt solutions (Dawson R.M., 1986).

Immediately before use, the dry powder of lyophilized PSH is dissolved in saline at the desired concentration. The solutions are used immediately. To reduce ischemia-reperfusion injury, depending on the severity of the lesion, it is recommended to use (wt%) from 0.1 to 0.4% (from 1 to 4 mg / ml) of the main active ingredient PSH (Prx VI-SOD).

The composition of the pharmaceutical composition included in the product contains the following ratio of components, wt. %

Antioxidant, recombinant PSH protein 0.1-0.4 Trehalose stabilizer 0.01-0.04 Pharmaceutically acceptable solvent 99.89-99.6

Example 3 Distribution of PSH (Prx VI-SOD) in an animal after injection

To determine the distribution of exogenous PSH (Prx VI-SOD) in the tissues of the small intestine when administered intravenously to a rat, PSH (Prx VI-SOD) is injected into a vein in an amount of 20 mg per animal. The 20 mg dose is not therapeutic but is used to identify exogenous PSH (Prx VI-SOD) in the tissues of the small intestine. PSH (Prx VI-SOD) was injected intravenously using a 24-G catheter and 20 minutes later, small intestine samples were taken for histological preparation. For immunohistochemical study of the distribution of endogenous PSH (Prx VI-SOD), samples of intestinal tissue of a healthy rat were used; His-tag domain was used as a detection tag, which was linked to the Prx VI-SOD construct.

For immunolabeling of exogenous Prx VI-SOD-His-tag, the sections are incubated with primary mouse monoclonal antibodies to the His-tag domain Prx6 (USBiological, USA) (1: 100) .The obtained data show that 20 minutes after PSH injection (Prx VI -SOD), is observed in the blood vessels of the submucosa and in the lamina propria of the mucous membrane. In addition, its diffuse distribution along the intestinal mucosa is noted (Fig. 1).

The study was carried out on 40 male Wistar rats. The animals were divided into four experimental groups. Ten animals received the introduction of recombinant PSH (Prx VI-SOD) according to our proposed scheme, followed by the induction of ischemic and reperfusion lesions of the small intestine. Induction of I-P lesions was performed by occluding the superior mesenteric artery.

Experimental groups:

1. Clipping of the superior mesenteric artery for 1 hour (period of ischemia), followed by its decamping to reproduce the reperfusion of the small intestine for 2 hours - 10 individuals.

2. Clipping of the superior mesenteric artery for 1 hour (period of ischemia), followed by its decamping to reproduce the reperfusion of the small intestine within 24 hours - 10 specimens

3. Intravenous administration of PSH (Prx VI-SOD) at a dose of 10 mg / kg, followed by clipping of the superior mesenteric artery for 1 hour (period of ischemia), followed by its decamping to reproduce reperfusion of the small intestine for 2 hours - 10 specimens

4. Intravenous administration of PSH (Prx VI-SOD) at a dose of 10 mg / kg, followed by clipping of the superior mesenteric artery for 1 hour (period of ischemia), followed by its decamping to reproduce reperfusion of the small intestine for 24 hours - 10 specimens

After the expiration of the reperfusion period for a period of 2 and 24 hours, the animals were decapitated and the tissues of the small intestine were harvested for morphological examination and assessment of the state of the endogenous antioxidant system. The morphological examination of the tissue was carried out using the standard histological technique with eosin / hematoxylin staining. Microscopic analysis of sections of intestinal and vascular tissues was performed using a Leica DM 6000 microscope. The final histological picture was assessed in accordance with the criteria described by Chiu C.J. et al [Chiu C.J. et al., 1970]. The study of the general state of cells was carried out using enzyme immunoassay to assess the level of the marker of apoptotic cell death caspase-3 in the intestinal tissue. ELISA was performed according to the standard method using polyclonal rabbit antibodies against the apoptosis marker caspase-3 (1: 1000) (Imgenex, USA).

Example 4

Histological studies have shown that the use of the proposed method for restoring the consequences of ischemic and reperfusion injuries of the small intestine has a positive effect on the preservation of the architecture of the intestinal wall (Fig. 2, tab. 1).

The degree of intestinal damage in ischemic and reperfusion injuries is expressed in accordance with the Chiu C.J. criteria. et al [Chiu C.J. et al., 1970] ischemic and reperfusion injuries. Where: Grade 0 - normal mucous; degree 1 - development of subepithelial spaces at the apex of the villi and vascular occlusion; grade 2 - expansion of subepithelial spaces with moderate detachment of the epithelium from the lamina propria; degree 3 - significant detachment of the epithelium along the entire length of the villi and bare areas of the villi: degree 4 - exposure of the villi with the appearance of their own lamina and expansion of capillaries; and grade 5, submucosal disintegration, hemorrhage, and ulceration.

Table 1 clearly shows that the use of exogenous PSH effectively reduces the degree of ischemia-reperfusion injury after 2 and 24 hours of reperfusion.

Example 5 Data of enzyme-linked immunosorbent assay of the content of a marker of apoptotic cell death.

Enzyme immunoassay of the content of the marker of apoptotic cell death caspase-3 showed that after 2 hours of reperfusion, the level of this marker increases by 3.5 times relative to the intact control (p <0.05), after 24 hours of reperfusion this indicator decreases by 1.75 times , relative to the values of the first group, but does not correspond to the control values. In the second group, the content of the marker of apoptotic cell death is 2 times higher than the control values (Fig. 3)

The use of exogenous PSH in the third experimental group reduces the level of caspase - 3 (1.4 times) relative to the level of this marker in the first experimental group (p <0.05), where, at equal time periods of ischemia and reperfusion, exogenous PSH was not used. After 24 hours of reperfusion, after the use of PSH (fourth experimental group), the level of caspase-3 decreases by 1.5 times compared to the values of the second experimental group, where exogenous PSH was not used at equal time periods of ischemia and reperfusion (p <0.05). In addition, in the fourth group, the rate of apoptotic death is reduced by almost 2 times, relative to the values of the third group (p <0.05), and corresponds to the values of the content of this marker in the control. Thus, a decrease in the degree of intestinal damage when using PSH according to the proposed treatment regimen occurs against the background of a decrease in the level of apoptotic cell death in the third and fourth experimental groups relative to the experimental groups, where, with equal periods of ischemia and reperfusion, PSH treatment.

The results obtained indicate that a therapeutic agent containing an effective concentration of the recombinant antioxidant PSH (Prx VI-SOD) and its method of using it to correct the consequences and complications of ischemic and reperfusion injuries of the small intestine by intravenous administration according to the scheme, contributed to a decrease in the degree of damage to intestinal tissues in acute and chronic experiment. In addition, it hindered the growth of the level of apoptotic death of cells of the small intestine, thereby preventing the progression of pathological changes in the organ during its ischemia-reperfusion injury.

Thus, the use of the proposed therapeutic agent and the method of its application for the correction of the consequences and complications of ischemic and reperfusion injuries of the small intestine makes it possible to prevent the progression of pathological changes in the organ during surgery, which justifies its use with therapeutic and prophylactic purposes for anti-ischemic and antiperfusional protection of the small intestine. ...

Industrial applicability

The method of treatment by intravenous administration of the antioxidant protein of recombinant PSH at a dose of 5 mg / kg to 20 mg / kg of body weight can find prophylactic and / or therapeutic use for correcting the consequences and complications of ischemia-reperfusion syndrome of the small intestine in various clinical conditions. The use of the method of treatment by using an agent based on PSH makes it possible to increase the effectiveness and shorten the duration of treatment, which is explained by the high effectiveness of PSH in comparison with known antioxidants. PSH is readily soluble in aqueous solutions, non-toxic and biocompatible with mammals, including humans, since it uses recombinant human peroxiredoxin VI as a leader compound.

The use of the recombinant PSH protein increases the protective functions of the body in ischemic and reperfusion injury, which is currently an urgent task for the treatment of such clinical conditions. The combination of two antioxidant enzymes with different targets for action can provide maximum protection against ROS overproduction through additive effects.

Sources of information

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Claims (8)

1. An agent for the treatment of ischemia-reperfusion lesions of the small intestine, characterized in that it contains a pharmaceutical composition of preparations consisting of a recombinant PSH protein, a stabilizer and a pharmaceutically acceptable solvent, where the components are taken in the following ratio, wt%: recombinant PSH protein 0.1-0.4 trehalose stabilizer 0.01-0.04 pharmaceutically acceptable solvent 99.89-99.56 2. The agent according to claim 1, characterized in that it contains trehalose as a stabilizer. 3. A method of increasing the resistance of the mammalian organism to the action of ischemic and reperfusion injuries of the small intestine, characterized in that a therapeutically effective amount of the agent according to PP. 1-2 is administered to the mammal once. 4. The method according to claim 3, characterized in that the therapeutically effective amount of the agent is administered intravenously 15-20 minutes before the induced ischemia of the mammal. 5. The method according to claim 3, characterized in that the agent is administered in a dose of 5 mg / kg to 20 mg / kg of the mammalian body weight. 6. The method according to claim 3, characterized in that the agent is administered in the most effective therapeutic dose of 10 mg / kg of body weight. 7. Application of the agent described in paragraphs. 1-2, as a drug for the treatment of ischemia-reperfusion injury of the small intestine.

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