AP928A - Cytokine and hemopoietic factor endogenous production enhancer and methods of use thereof. - Google Patents

Abstract

The invention concerns to medicine, in particular to pharmacology and therapy. According to the invention method of stimulating endogenous production of cytokine and . hemopoietic factors comprising introducing to a mammalian body in need of stimulation of cytokine or hemopoietic factor or both, an effective amount of oxidized glutathione, its pharmaceutically acceptable salt form, or/and its pharmaceutically acceptable derivative, for a period of time to stimulate said endogenous production to obtain a therapeutic effect, wherein said oxidized glutathione, or/and its pharmaceutically acceptable salt form, or/and its pharmaceutically acceptable derivative is introduced parenterally or topically. According to the invention, oxidized glutathione or/and its pharmaceutically acceptable salt form, or/and its pharmaceutically acceptable derivative is introduced along with an extender of the half life of said oxidized glutathione or/and its pharmaceutically acceptable salt form,-or/and its pharmaceutically acceptable derivative.

Description

CYTOKINE AND HEMOPOIETIC FACTOR ENDOGENOUS PRODUCTION ENHANCER AND METHODS OF USE THEREOF

Field of the Invention

The present invention relates to medicine and more particularly to pharmacology and therapy, and is intended to be used for preventing and treating various diseases by way of increasing and/or modulating endogenous production of cytokines and hemopoietic factors.

Background of the Invention

It has been known that a number of endogenously produced mammalian humoral factors, i.e. cytokines and hemopoietic factors possess important biological activities that are considerably helpful in treating various human diseases ',2. Many of these factors are being tested in man, those with proven efficiency being commercially available as medicinal agents.

The following cytokines and hemopoietic factors are being most extensively researched in oncology: interleukin 2 (IL-2)3’4, tumor necrosis factor alpha (ΤΝΈ-a)5, erythropoietin, macrophage-granulocyte and granulocyte colony-stimulating factors (GM-CSF and G-CS.F, respectively6-7). No less actively is being studied the use of cytokines and hemopoietic factors for the treatment of infectious disease: interferons (EFN-g and IFN-b)8’9-10, colony-stimulating factors11'12, and the like13. Colony-stimulating factors and erythropoietin are broadly used in hematology14,ls.

However, the medicinal use of these exogenously administered agents has its limitations associated -with the lack of acceptable drug formulations or their exorbitant cost, a short half-life of these substances in biological media, difficulties in dose finding as well as numerous toxic and allergic effects16'17, since even the recombinant products are more or less immunogenic to the human organism because of the processing fluctuations in the course of the artificial synthesis.

In this regard, in view of achieving a more invariable and significant therapeutic effect free of adverse reactions, it is preferable to induce the endogenous production of the autologous cytokines and hemopoietic factors immediately within the organism of a subject. The remedial effect due to such intrinsic stimulation is free of all the disadvantages associated with exogenously introduced cytokines and hemopoietic factors. A number of compounds are currently being evaluated that stimulate endogenous production of cytokines and hemopoietic factors in both experimental and clinical settings. There are universally known cases, including successful ones, of using microbial products for cancer therapy which in recent decades has been shown to be mediated via stimulation of the tumor necrosis factor endogenous production18. The products capable of evoking concomitant production of various cytokines and hemopoietic factors have presently come to be known as multi-cytokine inducers. Among these are a killed streptococcal preparation, Nocardia Opaca, and other bacterial products19'20'21. However, virtually all the substances possessing such capability are either killed microorganisms or microbial products or compounds having irregular composition, which results in their limited medicinal utility or even renders their therapeutic use impracticable. Thus, the problem of finding a medically and pharmaceutically acceptable inducer of the cytokine and hemopoietic factor endogenous production has not heretofore been resolved.

Oxidized glutathione (also known as glutathione disulfide and GSSG) will often be referred to as GSSG in this application. GSSG is known as a dimmer of tripeptide glutathione (g-glutamyl-cysteinyl-giycine) where two molecules of the tripeptide with the above structure are linked via a covalent disulfide bond between the cystein residues. Therefore, both the tripeptide glutathione (glutathione, reduced glutathione, GSH; hereinafter referred to as GSH) and its dimmer GSSG are natural metabolites present in animal tissues and biological fluids. At the same time, the natural blood level of GSSG is not sufficient for inducing the cytokine endogenous production in both normal and pathological conditions. GSH is known to.be one of the most important intermediates in the amino acid metabolism and a factor maintaining the intracellular homeostasis22,23. The reducing properties of GSH and its function as a donor of reduction equivalents, which is due to the sulfhydryl moiety of the cystein residue, are of key importance. This characteristic of GSH is responsible for the substance playing a crucial part in one of the most important intracellular antioxidant systems, consisting of GSH as such and two enzymes of its reversible conversion into GSSG: glutathione peroxidase and glutathione reductase24,25. The permanent functioning of said system is essential for inactivating or reducing endogenously generated oxidants as well as active metabolites of foreign substances26,27. * GSH is also known to participate in detoxification reactions involving a group of enzymes collectively known as glutathione S-transferase2s. These enzymes are capable of conjugating the GSH molecule with various xenobiotics by forming a bond between the latter and glutathione via the thiol moiety of the cystein residue of the tripeptide. The subsequent degradation of the conjugate is catalyzed by the g-glutamyl cycle enzymes, and may vary considerably depending upon the nature of the xenobiotic.

Under natural conditions, GSSG does not accumulate in amounts sufficient for inducing cytokine and hemopoietic factor production, due to a constant reduction of GSSG to GSH. The GSSG reduction to GSH also actively progresses in the intestines and liver upon GSSG oral administration, and as any product made of amino acids, the substance is proteolyticaily degradable in the gastrointestinal tract. GSSG is known to be used as a components of a nutritional supplement utilized as an adjunct diet in treating patients . However, being a peptide substance, most of the orally administered GSSG is digested in the gastrointestinal tract with the remainder being reduced in the intestinal and hepatic cells to GSH which can partially enter the circulation only as reduced glutathione. Thus, the main goal of the orally administering GSSG along with a number other compounds29 is to favour increasing in the blood and tissue level of the reduced glutathione (GSH).

According to the present invention, the delivery of GSSG into the organism through the gastrointestinal tract eliminates the possibility of the realization of its activity as a stimulator/modulator of endogenous production of cytokines and hemopoietic factors. Therefore, only parenteral route of GSSG administration can preserve the oxidized state of glutathione in the blood and other body liquids.

Moreover, using some agents capable of extending GSSG half life in biological media is important for realization of the multi-cytokine inducing activity of GSSG.

An elevation of the GSH endogenous levels for medicinal utility is known to be suggested for boosting immunity30 and treating toxemias, poisonings, diabetes mellitus, cardiovascular, infectious and other disorders31’32,33. Possible functions of GSH and GSSG appear in the literature.

Exogenous GSH or its direct (g-glutamyl-cystein, n-acetyl-cystein, and n-acetyl-cystein-glycine) or indirect (2-oxothiazolidine-4-carboxylate) biochemical precursors, or their salts and esters, are reportedly used as medicinal agents and dietary supplements in treating various diseases34'35'36’37,38. GSH is also claimed to be useful as a chemoprotective agent that prevents neurotoxicity in cancer chemotherapy39 as well as in combination with antineoplastics in order to augment their effect40.

No reference, however, is currently available to GSSG as a medicine in its own right (sole - * substance) used to induce the endogenous production of cytokines and hemopoietic factors. The , substance is known neither to have medicinal effects in human and animal diseases nor to be applied as a pharmaceutical agent for treating illnesses.

Summary of the Invention

It is an object of the present invention to provide an active substance, and advantageous combinations of said substance and/or its derivatives with extenders and/or enhancers or modulators of its ♦ activity which are capable of inducing endogenous cytokine and hemopoietic factor production to an individual or a subject in need thereof. «Subject in need thereof» as used in this application is intended to mean a mammal, e.g., man, domestic animals and livestock including cats, dogs, cattle and horses, having one or more manifestations of a disease in which stimulation of endogenous cytokine or hemopoietic factor (or both) production would be considered beneficial by those skilled in the art. «Therapeutic agent» as used in this application is meant to include any drug form of GSSG-containing material or GSSG alone, which has a therapeutic effect on neoplastic, infectious, hematologic, immunologic or other diseases. Therapeutic effect, as will be further defined, indicates any effect in man and other mammals which is beneficial, including curative, preventative, allowing maintenance at a beneficial level, or is in any way advantageous in connection with the body of man and othermammals.

In accordance with the present invention, it is GSSG that upon parenteral administration induces the endogenous cytokine and/or hematopoietic factor production in an individual or subject in need thereof, in both health and disease.

Having performed studies in search for a medically and pharmaceutically acceptable inducer of the cytokine and hemopoietic factor endogenous production, the applicants discovered a new property of a previously known substance, oxidized glutathione (oxidized glutathione, glutathione disulfide, GSSG; hereinafter often referred to as GSSG).

Being administered parenterally or acting on isolated ceils, the substance is capable of inducing production of several cytokines and hemopoietic factors in mammals (animals and humans) in both health and disease.

The inducer or stimulator/modulator of the endogenous cytokine and hemopoietic factor production is oxidized glutathione (GSSG) which is a dimmer of reduced glutathione having the structure g-glutamyl-cysteinyl-glycine, where the two molecules of the tripeptide are linked via a covalent disulfide bond between the cystein residues.

According to the invention, a method is provided for stimulating endogenous production of cytokine and hemopoietic factors by introducing to a mammalian body in need of stimulation of cytokine or hemopoietic factor or both, an effective amount of oxidized glutathione for a sufficient period of time to stimulate said endogenous production to obtain a therapeutic effect. -

Preferably, the glutathione is introduced parenterally or topically. In a preferred form, the method is carried out by introducing the oxidized glutathione (GSSG) or its derivatives with an extender of half life and/or enhancers or modulators to enhance the desired effect of stimulating endogenous production of cytokines and hemopoietic factors and producing a therapeutic effect in a body.

Preferably, the GSSG derivative is selected from the group of compounds representing a molecule of GSSG chemically modified by binding covalently as for example: with cysteamine - (2-mercaptoethylamine), lipoic acid (6,8-thioctic acid), camosine (b-alanyl-hystidine), adenosine (9-β-ϋ-ribofuranosyladenine), methionine (2-amino-4-[methylthio]butanoic acid); and both the D and L forms of the amino acids set forth in this paragraph can be used.

Particularly desirable derivatives are GSSG covalently bound either to cysteamine (S-thioethylamine-glutathione disulfide ), or to lipoic acid (bis-[6,8-thiooktanil]lglutathione disulfide), or to camosine ([b-alanyl-hystidiljlglutathione disulfide), or to adenosine ([9-β-ϋ- ribofuranosyladenil]lglutathione disulfide), or to methionine (bis-[2-amino-4- [methylthio]butanoil]lglutathione disulfide), or mixtures thereof and including tire D and/or L forms of amino acids herein.

Preferably, the extender is selected from the group consisting of pharmaceutically acceptable prooxidant compounds, (hydrogen peroxide, ascorbic acid) compounds capable of forming both weak ionic and coordinating links which stabilize molecule of GSSG (dimethyl sulfoxide), or materials which are competitors of NADP-H-dependent reduction of GSSG into GSH catalyzed by glutathione reductase, compounds capable of producing reversible inhibition of reduction of NADP+ into NADP-H catalyzed by glucose-6-phosphate-dehydrogenase or by other NADP-H-dependent enzymes, or mixtures thereof.

Particularly desirable extenders are hydrogen peroxide, inosine, ascorbic acid, dimethyl sulfoxide, or cystamine or mixtures thereof.

Preferably, the enhancer/modulator is selected from the group consisting of methyl moiety donators (such as choline-chloride{[2-hydroxyethyl]trimethylammonium chloride} or S-adenosyl-methionine), representatives of intracellular redox-oxidative pairs (such as Iipoic/dehydrolipoic, folic/dehydrofolic, ascorbic/dehydroascorbic acids). An enhancer or modulator or enhancer/modulator as used herein is meant to be a material which increases or changes beneficially in terms of curative outcomes the therapeutic effect of GSSG or its derivatives, but is not an extender of half life of the GSSG.

Particularly desirable enhancers or modulators are choline-chloride, S-adenosyl-methionine, lipoic (6,8-thioctic) and folic (pteroylglutamic) acids.

In the preferred form, GSSG is introduced to the body at a dose of from 0.01 to 0.5 mg of GSSG base per kg of body weight for GSSG base and its salts, and from 0.01 to 1.0 mg for GSSG derivatives, at least one time during each 24 hour period, although it can be continuously injected or otherwise introduced to the body to have a 24 hour total dosage of from 0.01 to 0.5 mg per kg of body weight-for * GSSG base and its salts, and from 0.01 to 1.0 mg for GSSG derivatives each 24 hour period. Preferably, < administration and introduction to the body is carried out until a desired stimulating effect increasing ' production of cytokines and hemopoietic factors and providing a therapeutic effect is obtained.

According to the invention, a therapeutic agent for treating neoplastic, infectious, hematologic, , immunologic and other diseases is provided, comprising an effective amount of oxidized glutathione, along with a pharmaceutically acceptable excipient. Preferably, the oxidized glutathione for parenteral use ; is in a pharmaceutically acceptable solvent as, for example, an aqueous solution including water, glucose solution, isotonic solutions of sodium chloride, buffered salt solutions. Preferably, a pharmaceutically acceptable extender capable of enhancing and prolonging therapeutic effect;as by increasing the half life of oxidized glutathione; or a pharmaceutically acceptable enhancer or modulator of GSSG activity by mechanisms other than increasing the GSSG half life, is used along with the GSSG.

The applicants have for the first time shown that an immediate action of exogenous GSSG or its salts on mammalian (human and laboratory animal) cells capable of producing cytokines and hemopoietic factors, exerts stimulation on the synthesis of these molecules and their increased level in the blood serum (in vivo conditions) or culture media {ex vivo or in vitro conditions). Additionally, the use of GSSG or its salts ensures the modulation of intracellular transmission pathways of cytokine signals and thus reproduce effects of those cytokines. The method suggested can bring about the effect of stimulating production of cytokines and hemopoietic factors, and this effect is elicited by the administration of GSSG into the organism or entering into the cultural media, as well as by the administration of GSSG in combination with pharmacologically active formulations mediating either the prolongation of glutathione’s retaining the oxidized form or enhancing or beneficially modulating its activity. The studies performed by the applicants have revealed GSSG and its formulations to possess a therapeutic effect in various experimental and clinical pathological conditions.

The revealed GSSG-induced stimulation of the endogenous cytokine and hemopoietic factor production in the body results in antitumor, anti-infective, hemopoietic, immunomodulatory and other pharmacological effects resulting, in turn, to a greater or lesser extent therapeutic or preventive effect in various diseases.

Brief Description of the Drawings

The above and other objects, features and advantages of the present invention will be better understood from the following specification when read in connection with the accompanying drawing in which:

Figures la, lb, lc and Id are charts showing fluorescent flow cytometric analysis of cells HL-60 (control or in the presence of the preparation of this invention), and fluorescent flow cytometric analysis of human lymphocytes (control or in the presence of the preparation of this invention), respectively, as will be described in the discussion of Example 4, relating to research of apoptosis-induced preparation activity in cultivated mammalian cells; and

Figure 2 is a drawing of GSSG structure with notification of sites for chemical modifications when GSSG salts and derivatives are reproduced; and

Figures 3, 4, 5, 6 and 7 are drawings of compounds where GSSG is covalently bound to: to cysteamine (S-thioethylamine-glutathione disulfide, Fig. 3); lipoic acid (bis-[6,8-thioktanil]lgiutathione disulfide, Fig. 4); camosine ([b-alanyl-hystidiljlglutathione disulfide, Fig. 5), adenosine ([9-p-D- * ribofuranosyladeniljlglutathione disulfide, Fig. 6), or to methionine (bis-[2-amino-4- [methylthiojbutanoiljlglutathione disulfide, Fig. 7).

Description of Preferred Embodiments

In accordance with the present invention, the medicinal agent suggested for treating neoplastic, infectious, hematologic, and other diseases, in which stimulation of the endogenous cytokine and hemopoietic factor production is appropriate, has an effective amount of GSSG and/or its pharmaceutically acceptable salts, and/or its pharmaceutically acceptable derivatives as its active principle It is also advantageous to'prepare a drug form of the medicinal agent as an injectable solution containing 0.01 to 2.0% of GSSG base for GSSG itself ant its salts, or 0.01 to 4.0% for GSSG derivatives.

The GSSG used as a therapeutic or medicinal agent in accordance with the present invention is shown in Figure 2. GSSG-and/or its pharmaceutically acceptable salts, and/or its pharmaceutically acceptable derivatives is preferably used in a carrier or solution as, for example, isotonic solution of sodium chloride, glucose solution, other buffer and salt solutions. Any aqueous based or solvent based earner or solvent can be used as long as the overall solution or dispersion is compatible with the body and pharmaceutically acceptable i.e., it does not cause any unwanted side effects in the body or unwanted interaction with GSSG and/or its pharmaceutically acceptable salts, and/or its pharmaceutically acceptable derivatives.

In the structural formula of Figure 2, points X,, X2, X3, X4, Xj, and X4 are noted as sites for chemical modification of the GSSG. Generally, the GSSG and/or its pharmaceutically acceptable salts, and/or its pharmaceutically acceptable derivatives is used in the form shown in solution or can be any of its physiologically and pharmaceutically acceptable soluble salts. The disodiura and di-lithium salts where Xb X4, are either sodium ions or lithium ions or a mixture, are preferred for best solubility of the drug.

Xi, X2, X3, and X4 can each be hydrogen if other substitutes are not used. Other salts of GSSG can be used, so long as they are pharmaceutically acceptable, i.e., do not adversely affect the body, for example, X1; X2, X3, and X4 can all be (or one or more of them can be) potassium, calcium, zinc, molybdenum, vanadium, fluoride, mixtures thereof or any other pharmaceutically acceptable substitutes. Water soluble salts are preferred for use in this invention. '

I

In the structural formula of Figure 3, points Xb (or probably Xt, and X2) is noted as site for-covalent binding with cysteamine molecule(s) (2-mercaptoethylamine). In the structural formula of Figure 4, points X5, and X$ are noted as sites for covalent binding with molecules of lipoic acid (6,8-thioctic acid). In the structural formula of Figure 5, point X3 is noted as site for covalent binding with molecule of camosine (b-alanyl-hystidine). In the structural formula of Figure 6, point X2 is noted as site for covalent binding with molecule of adenosine (9-3-D-ribofuranosyladenine). In the structural formula of Figure 7, points X5, and X^ are noted as sites for covalent binding with molecules of methionine (2-amino-4-[methylthiojbutanoic acid).

In accordance with the present invention, it is expedient to use such GSSG or its derivative drug forms and/or pharmaceutical compositions that either prolong oxidized glutathione half-life in tissues and biological fluids, or augment, or beneficially modulate the revealed biological and therapeutic properties of GSSG.

In accordance with the present invention, with the purpose of augmenting, beneficial modulating, and/or prolonging the therapeutic effect of GSSG, its drug form (injectable solution) is suggested to contain GSSG and/or its derivatives as described above (see Fig. 3-7) together with pharmaceutically acceptable component (extender, enhancer/modulator), capable of extending the half-life of GSSG and/or its derivatives or enhancing/modulating their biological and therapeutical effects. GSSG and/or its salts, and/or its derivatives can either be present in a single drug form together with the above mentioned extenders, enhancers/modulators (single injectable solution prepared beforehand or ex tempore), or be delivered into the body using separate drug forms: injectable solutions for GSSG and/or its salts, and/or its derivatives and/or derivatives salts; and any pharmaceutically acceptable drug forms, dosage regimens, and administration routes tor the above mentioned extenders, enhancers/benencial modulators.

As a pharmaceutically acceptable GSSG derivative, one of the compounds, or of their pharmaceutically acceptable salts, where GSSG is covalently bound to: either cysteamine (S-thioethylamine-glutathione disulfide, see Fig. 3 for structural formula), or lipoic acid (bis-[6,8-thiooktaniljlglutathione disulfide, see Fig. 4), or camosine ([b-alanyl-hystidil]lglutathione disulfide, see Fig. 5), or adenosine ([9-P-D-ribofuranosyladenil]lglutathione disulfide, see Fig. 6), or methionine (bis-[2-amino-4-[methylthio]butanoil]lglutathione disulfide, see Fig. 7), can be offered for application.

This is because the presence of one of the aforementioned molecules (cysteamine, lipoic acid, camosine, adenosine, or methionine) as a constituent part of a modified GSSG molecule, stabilizes structure of the corresponding derivative making it more resistant against proteolysis and/or reduction to GSH. As another way of stabilizing molecule of GSSG, its salts, or its derivatives/derivative salts, and protecting them against proteolysis and/or reduction, a replacement of one or more of L-amino-acids constituting the molecule of both GSSG and the aforementioned derivatives with their D-forms, can be implemented. c

All of pharmaceutically acceptable GSSG or derivatives most preferably can be used as the~ 4 medicinal agents in the injectable form of 1.0% solution with dosage range of from 0.01 to 0.5 mg of * GSSG base per kg of body weight for GSSG base and its salts, and from 0.01 to 1.0 mg for GSSG , derivatives, with preferable concentration range of from 0.5% to 5.0% one or more times a day, by one or more day pulses or continuously until a desired therapeutic effect has been achieved.As a pharmaceutical acceptable component or extender to prolong glutathione permanence in oxidized form, 0.003% hydrogen peroxide and/or 5.0% ascorbic acid can be offered for application. This is.-because in the presence of hydrogen peroxide or ascorbic acid, a donor of reactive oxygen intermediates (that is an oxidant), GSSG is reduced by glutathione reductase to GSH at a lesser speed, thereby conditioning a slower reduction of GSSG introduced exogenously into biological media.

Hydrogen peroxide preferably can be used in amounts of from 0.03 to 0.0003% by weight of solutions used (from 1.0 to 5.0 ml of solutions, regardless whether they contain or do not contain GSSG and/or its salts, and/or its derivatives/derivative salts). Ascorbic acid preferably can be used in amounts of from 0.1 to 10% by weight of solutions used (from 1.0 to 10.0 ml of solutions, regardless whether they contain or do not contain GSSG and/or its salts, and/or its derivatives/derivative salts).

Usage of an acceptable concentration of hydrogen peroxide (H2O2) and/or ascorbic acid in formulation of the drug form for parenteral administration, as well as usage of any other prooxidant compounds (donors of active oxygen form), makes it possible to realize only one of possible methods of the prolongation of oxidized glutathione and/or its derivative half-life in the biological fluids and tissues and, thereby, to enhance and prolong the pharmaceutical effect of GSSG and/or its derivatives.

We have also found some other pharmaceutically acceptable components or extenders capable of mediating the slowdown of the reduction of exogenous GSSG and/or its derivates into GSH in biological media. Such, in particular, are: the compounds capable of forming weak ionic and/or coordinating links which stabilize molecules of GSSG, for example, dimethyl sulfoxide; the factors capable of setting up competitive relations with a reduced form of the nicotinamide adenine dinucleotide phosphate or NADP-H, for example, inosine (and other derivatives of hypoxanthine); as well as the agents reversibly inhibiting the processes of reduction of the oxidized form of NADP+ into NADPH, for example, cystamine (2,2'-Dithio-bis[ethylamine]) and other inhibitors of glucose-6-phosphate-dehydrogenase.

Besides hydrogen peroxide and ascorbic acid, one of other pharmacologically accepted components capable to prolong the oxidized glutathione half-life can be dimethyl sulfoxide, which stabilize GSSG or its derivate molecules by forming both weak ionic and coordinating links with atoms of GSSG. Dimethyl sulfoxide is used most preferably as 7.0% (v/v) solution and preferably as a solution of from 0.1% to 30% by volume (from 1.0 to 30.0 ml of solutions or more when applied epicutaneously/through instillations, regardless whether they contain or do not contain GSSG/GSSG salts and/or its derivates/derivate salts. (

Since reduced NADP-H is the key cofactor of glutathione reductase system catalyzing the -reduction of GSSG into GSH, any pharmaceutically acceptable compounds or biophysical influence retarding the reduction of GSSG or blocking biological oxidation of NADP-H by glutathione reductase will facilitate preservation of GSSG/GSSG salts and/or its derivates/derivate salts from reduction in biological media and, therefore, will enhance and prolong its curative effect.

Due to conducted research we were the first to discover that GSSG pharmaceutical and medicinal effect will reinforce, when GSSG used in combination with agents capable of competition with NADP-H, as well as with compounds reversibly inhibiting the enzymatic reaction, catalyzed by glucose-6-phosphate-dehydrogenase which mediates the reduction of the oxidized form of NADP+. Reversible inhibitors of pentose phosphate pathway of glucose oxidation can be used.

Thus, besides hydrogen peroxide, ascorbic acid and dimethyl sulfide one of other pharmacologically accepted components capable to prolong the oxidized glutathione half-life can be inosine (hypoxanthine-9-D-ribofuranoside) used most preferably as 0.1 % solution and preferably as a solution of from 0.1% to 5% by weight (from 1.0 to 5.0 ml of solutions, regardless whether they contain GSSG/GSSG salts and/or its derivates/derivate salts.

The investigations carried out showed inosine to facilitate biological and therapeutical effects of GSSG. It was demonstrated that this property of inosine is based on its ability to compete with NADP-H, and thereby, to retard GSSG reduction into GSH. Moreover, we have also found that other hypoxanthine derivatives (including inosine, nucleoside ones, hypoxanthine riboside and other nucleoside derivatives of inosine) possess this property as well.

Also, besides hydrogen peroxide, ascorbic acid, dimethy sulfoxide and inosine, cystamine (2,2’-

Claims (1)

1. Original document published without claims.