Patent Publication Number: US-2006008487-A1

Title: Method of skin care and/or treatment using thioredoxin

Description:
RELATED APPLICATION DATA  
      This application claims the benefit of U.S. Provisional Application No. 60/587,217, filed on Jul. 12, 2004. 
    
    
     BACKGROUND OF THE INVENTION  
      Free radicals are formed in tissues in multiple ways, for example, as a result of exposure to sunlight. Free radicals initiate chain reactions that produce hydroxyl, superoxide and other radicals, which can damage DNA and other macromolecules. Other processes that generate free radicals are respiration and phagocytosis.  
      To control the damage done by these free radicals, mammals have a number of antioxidant systems. Some of them are obtained from the diet, like vitamin C, E, carotenoids, and polyphenols; and others are produced endogenously (glutathione, thioredoxin). Antioxidant enzymes, like superoxide dismutases and glutathione peroxidases, are also required.  
      Insufficient or unsuitable antioxidants may lead to oxidative stress and the consequent oxidation of membrane lipids, damage to DNA, etc. Cellular dysfunctions resulting from oxidative stress include excessive cell proliferation, cell death by apoptosis, lipid deposition, and mutagenesis. By creating such disorders, oxidative stress is partly responsible from a great number of age-related diseases as cancer, cardiovascular disorders, and neurodegenerative diseases as Alzheimer&#39;s disease.  
      Damage by free radicals formed in response to ultraviolet (UV) light is now believed to be a major cause of skin aging, with the term “photo-aging” being used to describe the multitude of effects of sunlight on the skin, including wrinkles, changes in the matrix proteins elastin and collagen, “age” spots (also called sun spots) and an increase in the incidence of skin cancer.  
      The treatment for sunburn, i.e., erythema in skin caused by solar exposure, used to be the topical use of lubricants and emollients. Thanks to a better understanding of the processes involved, new and more efficient treatments to treat skin damage caused by free radicals have been devised, and they include the topical application and/or ingestion of a variety of natural and synthetic antioxidants. Sometimes, the efficacy of these treatments may be limited because endogenous thioredoxin may be insufficient. Topical application of thioredoxin will help alleviate skin problems caused by solar or other types of free radiation producing free radicals, like radiation therapy for cancer patients.  
      Thioredoxins are small proteins with a redox active disulfide bridge present in the characteristic active site sequence: -Trp-Cys-Gly-Pro-Cys- (tryptophane-cysteine-glycine-proline-cysteine). Thioredoxins have a molecular mass of approximately 12,000. Animals, plants, and most microorganisms possess a system in which thioredoxin is reduced by NADPH and the enzyme NADP-thioredoxin reductase, as follows: 
 
NADPH+H + +Thioredoxin ox →NADP + +Thioredoxin red  
 
      Reduced thioredoxins have been found to serve as electron donors in a variety of cellular redox reactions. Thioredoxins are involved in many physiological processes like cell division, hormone action, transcription, meiosis, DNA replication, protein assembly and repair, and early embryo development. There is also abundant scientific research suggesting an important role of the thioredoxin system in the control of free radicals in animal tissues. Some examples:  
      1) Thioredoxin may be required for recycling of oxidized ascorbate (May et al. 1998)  
      2) Thioredoxin reductases associated with cell membranes has been shown to reduce free radicals at the surface of the skin (Schallreuter et al. 1986b)  
      3) Cell cultures of human keratinocytes contain a membrane-associated thioredoxin reductase that is extremely active in reducing radicals on the outer plasma membrane (Schallreuter et al. 1986a)  
      4) It has been speculated that the susceptibility of neurons to premature death in some diseases is partly due to an insufficient supply of mitochondrial thioredoxin (Yamagata et al. 2002)  
      5) The defense against oxidative damage by UV-generated free radicals in both guinea pig and human skin has been found to be mediated by thioredoxin reductase (Schallreuter et al. 1986b, Sundaram et al. 1990)  
      6) The thioredoxin reductase/thioredoxin system has been shown to reduce superoxide anion radicals through hydrogen peroxide to water (Sundaram et al. 1990). 
    
    
     DESCRIPTION OF THE INVENTION  
      The objective of this invention to provide a method and composition for a preventive regimen and/or therapy based upon the topical application to exposed or affected skin areas of at least one active agent, in association with a dermatologically acceptable carrier or vehicle. This invention is based upon the finding that reduced thioredoxin is useful for the disarming of active oxidative species. Reduced thioredoxin may also increase the efficacy of other ingredients in topical compositions for the prevention and treatment of burns.  
      These and other objectives are accomplished by the present invention, which provides methods and compositions for the prevention and/or treatment of skin damage caused by ultraviolet radiation or other radiations or by any kind of stress leading to the formation of free radicals, by applying topically to the exposed or affected skin areas an effective amount of thioredoxin, preferably in a dermatologically acceptable carrier.  
      As used herein, the term “thioredoxin” encompasses thioredoxins from any source, plant animal or from microorganisms, purified from natural sources such as spinach leaves or obtained by genetic engineering means, i.e., by over-expression of a protein, natural or foreign to the organism used for its production.  
      Thioredoxins obtained by over-expression in yeast or bacteria are preferred because they are less expensive than those obtained by purification from natural materials such as spinach leaves.  
      Many embodiments incorporate at least one other active ingredient with the thioredoxin. These include reduced glutathione, to ensure that the exogenous thioredoxin is in the reduced state. Other ingredients include natural or synthetic antioxidant molecules such as tocotrienols, vitamin E, ascorbic acid, astaxanthin, and/or lycopene. Other desirable ingredients are proteins capable of alleviating oxidative stress such as catalase and/or superoxide dismutase.  
      In the preferred practice of the invention, thioredoxin is applied in admixture with a dermatologically acceptable carrier or vehicle (e.g., as a lotion, cream, ointment, serum) so as to facilitate topical application and, in some cases, provide additional therapeutic effects as might be brought about by moisturizing the affected skin areas. As noted, other ingredients, particularly reduced glutathione, are advantageously included in the compositions.  
      The amount of thioredoxin necessary to bring about prevention and/or therapeutic treatment of damage by oxidative stress is not fixed, and is dependent upon the source and purity of thioredoxin employed, the amount and type of any additional ingredients used, particularly those that appear to exhibit synergistic effects, the skin type of the user, and, where present, the severity and extent of skin damage. Generally, the thioredoxin or composition containing it is topically applied in effective amounts to skin areas which have been damaged, or which are susceptible to damage, because of exposure to a cause of oxidative stress.  
      In one embodiment, the composition contains from about 0.0001% to about 1% (weight per volume), preferably from more than 0.001% or 0.002% to about 0.01% thioredoxin.  
      While the carrier for thioredoxin can be very simple (such as saline solution), it is generally preferred that the carrier be a composition that will facilitate topical application, and particularly one which will form a film or layer on the skin to which it is applied so as to localize the active ingredient. Many such compositions are known in the art, and can take the form of lotions, creams, gels, etc. Typical compositions include lotions containing water and/or alcohols and emollients such as natural oils and waxes, silicone oils, hyaluronic acid, glyceride derivatives, fatty acids or fatty acid esters or alcohols or alcohol ethers, lanolin and derivatives, polyhydric alcohols or esters, wax esters, sterols, phospholipids and the like, and generally also emulsifiers (nonionic, cationic or anionic), although some of the emollients inherently possess emulsifying properties. These same general ingredients can be formulated into a cream rather than a lotion, or into gels, or into solid sticks by utilization of different proportions of the ingredients and/or by inclusion of thickening agents such as gums or other forms of hydrophilic colloids. Such compositions are referred to herein as dermatologically-acceptable carriers.  
      Many preferred embodiments of this invention contain at least one or two, and sometimes several, other active ingredients in addition to thioredoxin, provided that the ingredients are not acids present in concentrations high enough to denature and inactivate thioredoxin, a protein.  
      Some embodiments may include reduced glutathione to maintain thioredoxin in its reduced state.  
      Tocotrienol, lycopene, astaxanthin, ascorbic acid, and/or vitamin E may also be added to the thioredoxin composition, alone or in combination with reduced glutathione in some embodiments.  
      In terms of a possible explanation for the effectiveness of the active ingredients in the prevention or treatment of damage to the skin, it is noted that thioredoxin, as an antioxidant, will help scavenge free radicals such as the oxygen radicals created by exposure of skin cells to ultraviolet radiation. Just like in the living cell, where a number of antioxidants work in a concerted fashion, some embodiments of this invention also use the synergistic effect of antioxidants.  
      The method of the present invention is particularly useful for the prevention and treatment of sunburn and other skin damage resulting from exposure to ultraviolet radiation. Thioredoxin, alone or with other active ingredients can thus be added to dermatological creams and emollients as well as to commercial sunscreens to enhance their sun protection activity, or to creams used to treat sunburn or burns produced by therapeutical radiation used to treat cancer.  
      It is also worth noting that free radicals seem to be involved in some specific skin diseases, like vitiligo, and these diseases may respond well to treatment with thioredoxin.  
      Having described the invention with reference to particular compositions, theories of effectiveness, it will be apparent to those of skill in the art that it is not intended that the invention be limited by such illustrative embodiments or mechanisms, and that modifications can be made without departing from the scope or spirit of the invention, as defined by the appended claims. It is intended that all modifications and variations be included within the scope of the invention. The claims are meant to cover the claimed components and steps in any sequence which is effective to meet the objectives there intended, unless the context specifically indicates the contrary.