| Method to enhance delivery of glutathione and atp levels in cells -> Monitor Keywords |
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Method to enhance delivery of glutathione and atp levels in cellsRelated Patent Categories: Drug, Bio-affecting And Body Treating Compositions, Designated Organic Active Ingredient Containing (doai), O-glycoside, , Nitrogen Containing Hetero RingMethod to enhance delivery of glutathione and atp levels in cells description/claimsThe Patent Description & Claims data below is from USPTO Patent Application 20060105972, Method to enhance delivery of glutathione and atp levels in cells. Brief Patent Description - Full Patent Description - Patent Application Claims
BACKGROUND OF THE INVENTION [0001] The protective mechanisms of mammalian cells against exogeneous and endogenous stressors that generate harmful free radicals employ the antioxidant co-enzyme, glutathione (GSH). GSH is important in maintaining the structural integrity of cell and organelle membranes and in the synthesis of microtubules and macromolecules. See C. D. Klassen et al. Fundamental and Applied Toxicology, 5, 806 (1985). Stimulation of GSH synthesis in rat renal epithelial cells and stomach cells has been found to protect the cells from the toxic effects of cyclophosphamide and serotonin, respectively. Conversely, inhibition of glutathione synthesis and glutathione depletion has been found to have the following effects: (a) decreased cell viability, (b) increased sensitivity of cells to the effects or irradiation, (c) increased sensitivity of tumor cells to peroxide cytolysis, (d) decreased synthesis of prostaglandin E and leukotriene C and (e) selective destruction of trypanosomes in mice. [0002] Biosynthesis of glutathione (GSH) involves two sequential reactions that utilize ATP and that are catalyzed by the enzymes .gamma.-glutamylcysteine synthetase and glutathione synthetase (GSH-synthetase) using the three precursor amino acids L-glutamic acid, L-cysteine, and glycine, as shown in FIG. 1. [0003] All substrate-level reactants occur at near enzyme-saturating concentrations in vivo with the exception of L-cysteine, whose cellular concentration is exceedingly low. Therefore, the first reaction in which L-cysteine is required, i.e., the synthesis of .gamma.-L-glutamyl-L-cysteine, is the rate-limiting step of glutathione biosynthesis. Thus, the availability of intracellular L-cysteine is a critical factor in the overall biosynthesis of GSH, are sufficient stores of ATP. [0004] In the synthesis of ATP via the nucleotide salvage pathway, the nucleotide precursors that may be present in the tissue are converted to AMP and further phosphorylated to ATP. Adenosine is directly phosphorylated to AMP, while xanthine and inosine are first ribosylated by 5-phosphoribosyl-1-pyrophosphate (PRPP) and then converted to AMP. [0005] Ribose is found in the normal diet only in very low amounts, and is synthesized within the body by the pentose phosphate pathway. In the de novo synthetic pathway, ribose is phosphorylated to PRPP, and condensed with adenine to form the intermediate adenosine monophosphate (AMP). AMP is further phosphorylated via high energy bonds to form adenosine diphosphate (ADP) and ATP. [0006] During energy consumption, ATP loses one high energy bond to form ADP, which can be hydrolyzed to AMP. AMP and its metabolites adenine, inosine and hypoxanthine are freely diffusible from the muscle cell and may not be available for resynthesis to ATP via the salvage pathway. [0007] The availability of PRPP appears to control the activity of both the salvage and de novo pathways, as well as the direct conversion of adenine to ATP. Production of PRPP from glucose via the pentose phosphate pathway appears to be limited by the enzyme glucose-6-phosphate dehydrogenase (G6PDH). Glucose is converted by enzymes such as G6PDH to ribose-5-phosphate and further phosphorylated to PRPP, which augments the de novo and salvage pathways, as well as the utilization of adenine. [0008] Many conditions produce hypoxia. Such conditions include acute or chronic ischemia when blood flow to the tissue is reduced due to coronary artery disease or peripheral vascular disease where the artery is partially blocked by atherosclerotic plaques. In U.S. Pat. No. 4,719,201, it is disclosed that when ATP is hydrolyzed to AMP in cardiac muscle during ischemia, the AMP is further metabolized to adenosine, inosine and hypoxanthine, which are lost from the cell upon reperfusion. In the absence of AMP, rephosphorylation to ADP and ATP cannot take place. Since the precursors were washed from the cell, the nucleotide salvage pathway is not available to replenish ATP levels. It is disclosed that when ribose is administered via intravenous perfusion into a heart recovering from ischemia, recovery of ATP levels is enhanced. [0009] Transient hypoxia frequency occurs in individuals undergoing anesthesia and/or surgical procedures in which blood flow to a tissue is temporarily interrupted. Peripheral vascular disease can be mimicked in intermittent claudication where temporary arterial spasm causes similar symptoms. Finally, persons undergoing intense physical exercise or encountering high altitudes may become hypoxic. U.S. Pat. No. 6,218,366 discloses that tolerance to hypoxia can be increased by the administration of ribose prior to the hypoxic event. [0010] Hypoxia or ischemia can also deplete GSH. For example, strenuous aerobic exercise can also deplete antioxidants from the skeletal muscles, and sometimes also from the other organs. Exercise increases the body's oxidative burden by calling on the tissues to generate more energy. Making more ATP requires using more oxygen, and this in turn results in greater production of oxygen free radicals. Studies in humans and animals indicate GSH is depleted by exercise, and that for the habitual exerciser supplementation with GSH precursors may be effective in maintaining performance levels. See L. L. Ji, Free Rad. Biol. Med., 18, 1079 (1995). [0011] Tissue injury, as from burns, ischemia and reperfusion, surgery, septic shock, or trauma can also deplete tissue GSH. See, e.g., K. Yagi, Lipid Peroxides in Biology and Medicine, Academic Press, N.Y. (1982) at pages 223-242; A. Blaustein et al., Circulation, 80, 1449 (1989); H. B. Demopoulos, Pathology of Oxygen, A. P. Autor, ed., Academic Press, N.Y. (1982) at pages 127-128; J. Vina et al., Brit. J. Nutr., 68, 421 (1992); C. D. Spies et al., Crit. Care Med., 22, 1738 (1994); B. M. Lomaestro et al., Annals. Pharmacother., 29, 1263 (1995) and P. M. Kidd, Alt. Med. Res., 2, 155 (1992). [0012] It has been hypothesized that delivery of L-cysteine to mammalian cells can elevate GSH levels by supplying this biochemical GSH precursor to the cell. However, cysteine itself is neurotoxic when administered to mammals, and is rapidly degraded. In previous studies, it was shown that N-acetyl-L-cysteine, L-2-oxothiazolidine-4-carboxylate, as well as 2(R,S)-n-propyl-, 2(R,S)-n-pentyl and 2(R,S)-methyl-thiazolidine-4R-carboxylate can protect mice from heptatotoxic dosages of acetaminophen. See H. T. Nagasawa et al., J. Med. Chem., 27, 591 (1984) and A. Meister et al., U.S. Pat. No. 4,335,210. L-2-Oxothiazolidine-4-carboxylate is converted to L-cysteine via the enzyme 5-oxo-L-prolinase. As depicted in FIG. 2, compounds of formula 1, e.g., wherein R=CH.sub.3, function as prodrug forms of L-cysteine (2), liberating this sulfhydryl amino aciuc by nonenzymatic ring opening and hydrolysis. However, the dissociation to yield L-cysteine necessarily releases an equimolar amount of the aldehyde (3), RCHO. In prodrugs in which R is an aromatic or an alkyl residue, the potential for toxic effects is present. [0013] U.S. Pat. No. 4,868,114 discloses a method comprising stimulating the biosynthesis of glutathione in mammalian cells by contacting the cells with an effective amount of a compound of the formula (1): wherein R is a (CHOH).sub.nCH.sub.2OH and wherein n is 1-5. The compound wherein n is 3 is 2(R,S)-D-ribo-(1', 2', 3', 4'-tetrahydroxybutyl)thiazolidone-4(R)-carboxylic acid (Ribose-Cysteine, RibCys). Following in vivo administration, RibCys releases cysteine by non-enzymatic hydrolysis. RibCys has been demonstrated to be effective to protect against acetaminophen-induced hepatic and renal toxicity. A. M. Lucus, Toxicol. Pathol., 28, 697 (2000). RibCys can also protect the large and small bowel against radiation injury. See M. P. Caroll et al., Dis. Colon Rectum, 38, 716 (1995). These protective effects are believed to be due to the stimulation of GSH biosynthesis, which elevates intracellular GSH. However, a need exists for methods to restore or maintain intracellular GSH stores in mammalian tissues subjected to hypoxic conditions in which the ATP stores necessary to drive the biosynthesis of GSH and its precursors are depleted. SUMMARY OF THE INVENTION [0014] The present invention provides a method to treat a mammal threatened by, or afflicted with a hypoxic condition (hypoxia) comprising administering an effective amount of a compound of formula (Ia): (RibCys) or a pharmaceutically acceptable salt thereof, effective to counteract the effects of said hypoxia in the tissue(s) of said mammal. Although depressed glutathione levels have been implicated in a number of hypoxic conditions, as discussed above, the use of RibCys or its salts to prevent, counteract or otherwise treat such conditions has not been reported. It is believed that simply administering a GSH precursor such as cysteine will not be as effective in many instances of hypoxia, when the depletion of ATP stores contributes to inhibition to the biosynthesis of GSH. As well as functioning as a prodrug for cysteine, administration of effective amounts of RibCys can deliver amounts of ribose to ATP-depleted tissues that stimulate the in vivo synthesis of ATP and that also can stimulate the synthesis of NADPH (nicotinamide adenine dinucleotide phosphate, reduced). This coenzyme supplies the electrons to glutathione reductase, which in turn recycles oxidized GSH via GSSG, to free GSH, which resumes its protective role as a cofactor for antioxidant enzymes in the cell. Optionally, compound (Ia) can be administered with an additional amount of free ribose. Preferably, administration will be by oral administration, particularly in prophylactic or pre-loading situations, but parenteral administration, as by injection or infusion, may be necessary in some situations. BRIEF DESCRIPTION OF THE DRAWINGS [0015] FIG. 1 depicts the metabolic synthesis of glutathione (GSH) from L-glutanic acid. [0016] FIG. 2 depicts the in vivo dissociation of a compound of formula I to yield cysteine and an aldehyde. DETAILED DESCRIPTION OF THE INVENTION [0017] As used herein, the term RibCys refers to 2(R,S)-D-ribo-(1', 2', 3', 4'-tetrahydroxybutyl)thiazolidine-4(R)-carboxylic acid, as well as the 2R or 2S enantiomers of (Ia), and its pharmaceutically acceptable salts. Such salts include alkali metal salts of the carboxylic acid moiety as well as stable acid addition salts of the NH moiety, including salts of both inorganic and organic acids, such as citrate, malate, gluconate, glutamate, hydrochloride, hydrosulfate and the like. [0018] As used herein, the term "hypoxia" or "hypoxic condition" is defined to mean a condition in which oxygen in one or more tissues of a mammal is lowered below physiologic levels, e.g., to a less than optimal level. Hypoxia also includes conditions in which oxygen levels are lowered in tissues due to stress such as aerobic exercise, physical weight pressure, anesthesia, surgery, anemia, acute respiratory distress syndrome, chronic illness, chronic fatigue syndrome, trauma, burns, skin ulcers, cachexia due to cancer and other catabolic states and the like. Hypoxia also includes "ischemia" or "ischemic conditions" in which tissues are oxygen-deprived due to reduction in blood flow, as due to constriction in, or blockage of, a blood vessel. Ischemia and/or ischemic conditions include those caused by coronary artery disease, cardiomyopathy, including alcoholic cardiomyopathy, angioplasty, stenting, heart surgery such as bypass surgery or heart repair surgery ("open-heart surgery" ), organ transplantation, prolonged weight pressure on tissues (pressure ulcers or bedsores), ischemia-reperfusion injury which can cause damage to transplanted organs or tissue, and the like. The present invention is effective to treat the GSH and ATP depletion due to hypoxia and thus to increase a subject's energy level strength and well-being, even though the underlying cause of the hypoxic condition, such as viral or bacterial infection, exposure to bacterial or other toxins, low red-cell counts, aging, cancer or continued exercise, is not affected. [0019] The term "treating" or "treatment" as used herein includes the effects of RibCys administration to both healthy and patients afflicted with chronic or acute illness and includes inducing protective affects as well as decreasing at least one symptom of a past or ongoing hypoxic condition. [0020] Effective doses of RibCys will vary dependent upon the condition, age and weight of the patient to be treated, the condition to be treated and the mode of administration. Both cysteine, as released in vivo from RibCys in animal models, and ribose, as administered directly to human subjects, have been found to be essentially non-toxic over wide dosage ranges. For example, ribose has been reported to increase exercise capacity in healthy human subjects when taken orally at dosages of 8-10 g per day by an adult. See U.S. Pat. No. 6,534,480. RibCys administered to mice at 8 mmol/kg i.p., increased glutathione levels in numerous organs, including heart (1.5.times.) and muscle tissue (2.5.times.). See, J. C. Roberts, Toxicol. Lett., 59, 245 (1991). Likewise, RibCys at 8 mmol/kg has been found to deliver effective protective amounts of cysteine to mice exposed to cyclophosphamide. This dose can deliver about 70-80 g of ribose and about 60-70 g of cysteine to an adult human. See J. C. Roberts, Anticancer Res., 14, 383 (1994). Doses of 2 g/kg RibCys were reported to protect mice against acetaminophen hepatic and renal toxicity by A. M. Lucas et al., Toxicol. Pathol., 20, 697 (2000). Doses of 1 g/kg RibCys were reported to protect mice against irradiation-induced bowel injury (see J. K. Rowe et al., Dis. Colon Rectum, 36, 681 (1993). J. E. Fuher (U.S. Pat. No. 4,719,201) reported that doses of ribose of about 3 g/day for at least 5 days effectively restored and maintained ATP levels in dogs subjected to ischemia (heart attack model), doses that delivered about 550-700 mg/kg of ribose to an 30 kg dog. Continue reading about Method to enhance delivery of glutathione and atp levels in cells... Full patent description for Method to enhance delivery of glutathione and atp levels in cells Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Method to enhance delivery of glutathione and atp levels in cells patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. 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