Methods and compositions for treating proliferative disorders of the skin and other skin conditions - Patents.com

Abstract

A method for treating a proliferative skin disease includes treating a patient in need of treatment with a topical formulation comprising allopurinol or a pharma- ceutically acceptable salt thereof, and a corresponding formulation is provided. In some embodiments, the topical formulation slows proliferation of cells involved in the proliferative disease of skin cells. A method for treating a skin condition includes treating a patient in need of treatment with a topical formulation comprising allopurinol or a pharma- ceutically acceptable salt thereof. The topical formulation may comprise allopurinol or a pharma- ceutically acceptable salt thereof and glutathione or a pharma- ceutically acceptable salt thereof.

Description

[Technical field]

[0001] [Reference to related application] This application claims priority to and the benefit of U.S. Provisional Patent Application No. 63 / 266,433, filed January 5, 2022, and U.S. Provisional Patent Application No. 63 / 382,008, filed November 2, 2022, which are incorporated by reference in their entireties herein.

[0002] [Field of the Invention] The present embodiments are directed to the field of treatment of skin conditions. More particularly, the present embodiments relate to methods and compositions for the topical treatment of diseases manifested by increased cell proliferation, such as, for example, psoriasis and keratoacanthoma. [Background technology]

[0003] Many skin diseases with various causes appear with an increased proliferation rate of certain cells in the skin. Such skin diseases include psoriasis, keratoacanthoma, rosacea and keloids. Skin psoriasis, also known as plaque psoriasis, is a common disease that affects more than 3 million patients every year in the United States alone. Skin psoriasis is a chronic inflammatory disease that may manifest itself on the skin as a rash, dryness, cracks, flakiness, peeling, small bumps, thickening, and / or redness. Guttate psoriasis is another type of psoriasis that is less common than plaque psoriasis, but is more acute, and typically appears as small, teardrop-shaped red spots.

[0004] Conventional topical treatments for cutaneous psoriasis include corticosteroids, vitamin D analogs, 0.05%-0.1% tazarotene, 3%-10% salicylic acid, methotrexate, and 20% liquor carbonic distillate (LCD) solution. For mild to moderate psoriasis, topical vitamin D is often used in combination with steroids. Vitamin D administration may inhibit cell replication, possibly by increasing cellular calcium uptake, but vitamin D may also increase the incidence of rosacea and is not effective for scalp psoriasis or more aggressive cases of psoriasis. The most common form of psoriasis is plaque psoriasis, which is manifested by plaques with marked thickening of the stratum corneum or keratin crust. The plaques provide a barrier to drug penetration. Unfortunately, methotrexate has a narrow therapeutic window and is toxic in overdoses. There is currently no satisfactory treatment for cutaneous psoriasis.

[0005] Many efforts have been made to treat these conditions by utilizing immune inhibition, including anti-cytokine biologics targeting interleukin-17 (IL-17) or its ligand (IL-17A), interleukin-23 (IL-23), or tumor necrosis factor alpha (TNF-α), or the small molecule drug apremilast (marketed as Otezla by Amgen, Thousand Oaks, Calif.), a selective inhibitor of phosphodiesterase 4 (PDE4), an enzyme that blocks the production of TNF-α by rheumatoid synovial cells. (R) Drugs such as cyclosporine (sold under the brand name cyclosporine) are increasingly being used to treat psoriasis. Although initially effective, these drugs carry a low-intensity risk of serious toxicities, including cancer and activated tuberculosis. Moreover, many of these treatments only attempt to interrupt a single step in a highly complex and redundant replication-stimulating network.

[0006] The inflammatory nature of conditions such as psoriasis has also been the subject of intense research for decades. In 2005, Namazi ("Cannabinoids, loratadine and allopurinol as novel additions to the antisporiatic ammunition", Journal of the European Academy of Dermatology and Venerology, Vol. 19, pp. 319-322, (2005), incorporated herein by reference) suggested the cannabinoids, loratadine and allopurinol, as useful drugs for treating these conditions.

[0007] Allopurinol was mentioned by Namazi as an inhibitor of xanthine oxidase with free radical scavenging activity, and was speculated to have the ability to alleviate inflammation associated with psoriasis. If allopurinol effectively treats psoriasis by interfering with the free radical scavenging activity mediated by xanthine oxidase, it is expected that oxypurinol, the only active metabolite of allopurinol responsible for xanthine oxidase inhibition, will be even more effective in treating psoriasis by acting through the mechanism of xanthine oxidase inhibition. Allopurinol inhibits the production of intercellular adhesion molecule-1 (ICAM-1), P2X, and tumor necrosis factor alpha (TNF-α).

[0008] Allopurinol has been used topically to prevent chemotherapeutic agent skin toxicity, termed "hand-foot" syndrome by Rodemer (see, e.g., U.S. Patent No. 8,623,878), but a subsequent clinical trial with allopurinol by Rodemer, although accrual was completed, did not achieve clinical efficacy endpoints.

[0009] Salim proposed the topical use of allopurinol or oxypurinol in combination with sulfonylmethanesulfonates for a number of diseases (see, e.g., WO94 / 05291 and WO94 / 05293), but Salim's related publications were effectively retracted by the publishers as fraudulent (see Hammerschmidt et al., "Allegations of Impropriety in Manuscripts by Aws S. Salim: Examination and Withdrawal of Journal aegis", J. Lab. Clin. Med., Vol. 123, pp. 795-799 (1994)). Salim did not suggest psoriasis in the list of conditions treated by either drug. With regard to oral use, allopurinol has long been in clinical use as an oral treatment for gout and certain types of kidney stones.

[0010] There have been puzzling and conflicting reports regarding the oral use of allopurinol for the treatment of psoriasis. Viglioglia et al. (see "Allopurinol in Psoriasis", Dermatologica, Vol. 141, pp.203-207 (1970)) reported that the treatment of psoriasis with allopurinol gave excellent results in 50% of cases, good results in 34% of cases, and fair results in 16% of cases.

[0011] However, in a subsequent crossover double-blind study, oral administration of allopurinol showed no improvement over placebo in treating psoriasis (see Feuerman et al., "Allopurinol in psoriasis-a double blind study", British Journal of Dermatology, Vol. 89, pp. 83-86 (1973)).

[0012] As mentioned above, the active species in xanthine oxidase inhibition by allopurinol is oxypurinol. Oxypurinol is the only known metabolite of allopurinol that is responsible for xanthine oxidase inhibition. If Namazi's proposal of xanthine oxidase inhibition is correct, oxypurinol would be superior to allopurinol in treating psoriasis, and allopurinol would not be used as a therapeutic agent.

[0013] Another reason to study oxypurinol rather than allopurinol is that it has long been known that allopurinol, unlike oxypurinol, causes a deleterious depletion of phosphoribosyl pyrophosphate (PRPP), an important energy mediator, in people (see, e.g., Fox et al., “Depletion of Erythrocyte Phosphoribosylpyrophosphate in Man: A Newly Observed Effect of Allopurinol”, The New England Journal of Medicine, Vol. 283, pp. 1177-1182 (1970), incorporated herein by reference). Thus, if true, oxypurinol rather than allopurinol would be a more active and safer drug than allopurinol. After a great deal of time and resources, the present study has shown, quite unexpectedly, that allopurinol is much more active and less toxic than oxypurinol.

[0014] In fact, the above careful review of the published literature regarding oxypurinol and allopurinol made such a compelling case regarding the superiority of oxypurinol over allopurinol as a topical agent that significant time and financial resources were expended to prepare, file, and defend U.S. Patent Application Publication No. 2020 / 0046703, published February 13, 2020, regarding the use of oxypurinol over allopurinol, which was abandoned after clinical trials confirming the superiority of oxypurinol over allopurinol conclusively failed.

[0015] There is a strong need for safe and effective treatments for psoriasis and other skin diseases characterized by increased cell proliferation. Broad inhibition of cell proliferation carries the risk of serious side effects. For one, inhibition of all proliferation after tissue injury would prevent the natural healing process, which is a concern for any cell proliferation inhibition strategy.

[0016] An ideal therapy would reduce cell replication by inhibiting non-redundant pathways essential for cell replication, while at the same time not inhibiting the baseline and salvage functions of cells that are important for maintaining viability.

[0017] This study was the first to attempt to explore the possibility of developing a xanthine oxidase inhibitor therapy for psoriasis by using oxypurinol, an essential metabolite of allopurinol, which inhibits xanthine oxidase. Summary of the Invention

[0018] In some embodiments of the present disclosure, a topical pharmaceutical formulation is provided that includes allopurinol or a pharma- ceutically acceptable salt thereof and a co-agent or a pharma- ceutically acceptable salt thereof.

[0019] In another aspect of the present disclosure, there is provided a method of treatment for a skin disorder comprising applying to a patient in need of treatment a topical formulation comprising allopurinol or a pharma- ceutically acceptable salt thereof and an adjuvant or a pharma- ceutically acceptable salt thereof. [Brief description of the drawings]

[0020] [Figure 1] FIG. 1 shows the number of total HaCaT cells versus the initial number of viable cells in the presence of certain amounts of allopurinol, oxypurinol and methotrexate in the M5 psoriasis cocktail.

[0021] [Diagram 2] FIG. 2 shows the number of dead HaCaT cells relative to the initial number of live cells in the presence of certain amounts of allopurinol, oxypurinol and methotrexate in the M5 psoriasis cocktail.

[0022] [Diagram 3] FIG. 3 shows images of the MatTek human psoriasis tissue culture system (MatTek, Ashland, Mass.) after exposure to 3 mM allopurinol treatment medium for 5 days.

[0023] [Figure 4] FIG. 4 shows images of the MatTek human psoriasis tissue culture system after exposure to 3 mM allopurinol and 3 mM glutathione treatment medium for 5 days.

[0024] [Figure 5A] FIG. 5A shows the initial images of the MatTek human psoriasis tissue culture system after exposure to 100 nM vitamin D treatment medium for 5 days.

[0025] [Figure 5B] FIG. 5B shows a second image of the MatTek human psoriasis tissue culture system after exposure to 100 nM Vitamin D treatment medium for 5 days.

[0026] [Figure 6] FIG. 6 shows dermal thickness in the MatTek human psoriatic tissue culture system treated with allopurinol and with a combination of allopurinol and glutathione versus controls.

[0027] [Figure 7] FIG. 7 is a bar graph of relative levels of Ki-67 in the MatTek human psoriatic tissue culture system at day 6.

[0028] [Figure 8] FIG. 8 shows the relative levels of biomarkers Ki-67, p21, IL-17A, IL-23A, TNF-α, and TGF-β after exposure of MatTek human psoriatic tissue cultures to allopurinol, glutathione, a combination of allopurinol and glutathione, and controls.

[0029] [Figure 9] FIG. 9 shows the ratio of p21 to Ki-67 mRNA abundance in the MatTek human psoriatic tissue culture system.

[0030] [Figure 10A] FIG. 10A shows a psoriatic plaque on the left shoulder before treatment.

[0031] [Figure 10B] FIG. 10B shows the left shoulder of FIG. 10A after one month of treatment.

[0032] [Figure 11A] FIG. 11A shows Dupuytren's contracture of the right fifth finger before treatment.

[0033] [Figure 11B] FIG. 11B shows the right fifth finger of FIG. 11A after one month of treatment.

[0034] [Figure 11C] FIG. 11C shows the right fifth finger of FIG. 11A after two months of treatment.

[0035] [Figure 11D] FIG. 11D shows the right fifth finger of FIG. 11A after 4 months of treatment.

[0036] [Figure 12A] FIG. 12A shows a long-standing allergic reaction in the calves before treatment.

[0037] [Figure 12B] FIG. 12B shows the back of the calf from FIG. 12A after 5 days of treatment.

[0038] [Figure 13A] FIG. 13A shows three areas of guttate psoriasis before treatment.

[0039] [Figure 13B] FIG. 13B shows the three areas of FIG. 13A after one and a half days of treatment.

[0040] [Figure 13C] FIG. 13C shows the three areas of FIG. 13A after two days of treatment.

[0041] [Figure 13D] FIG. 13D shows the three areas of FIG. 13A after two and a half days of treatment.

[0042] [Figure 13E] FIG. 13E shows the three areas of FIG. 13A after 6.5 days of treatment.

[0043] [Figure 14A] FIG. 14A shows the allergic reaction in the shin before treatment.

[0044] [Figure 14B] FIG. 14B shows the shin of FIG. 14A after 7 days of treatment. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0045] [Detailed Description] Surprisingly, it has been found that not only is allopurinol more effective than oxypurinol in reducing psoriasis-induced abnormal cell proliferation in tissue culture, but that allopurinol reduces cell death more effectively than oxypurinol.

[0046] Keratinocytes grow slowly, but not without growth (see, e.g., Squier et al., ed., Human Oral Mucosa: Development, Structure, and Function, Wiley-Blackwell, p. 29 (2011)). Thus, maintaining tissue integrity is crucial. Again, surprisingly, allopurinol was found to cause less cell death than oxypurinol in tissue culture models. At the highest human dose tested, topical allopurinol was both effective and well tolerated.

[0047] In representative embodiments, a co-agent is administered with allopurinol. In many embodiments, the co-agent is an antioxidant. In some embodiments, the co-agent is a skin permeation enhancer. In some embodiments, the co-agent is a sulfur-containing antioxidant. In many embodiments, the co-agent comprises a thiol or prothiol. Suitable thiols may include, but are not limited to, glutathione, N-acetylcysteine, cysteine, theanine, cystine, β-mercaptoethanol, and / or dithiothreitol. In certain embodiments, the co-agent is glutathione.

[0048] The formulations of the present disclosure have been shown to be effective against psoriasis. Psoriasis plaques are a barrier to topical drug penetration. It was expected that agents such as glutathione would be useful for improving allopurinol delivery, but it was surprisingly found that co-administration of glutathione can not only improve drug delivery, but also reduce psoriasis swelling that was not observed when allopurinol was administered alone. Example 2 shows that co-administration of topical allopurinol and glutathione has a synergistic effect in a tissue culture model of psoriasis without plaques, compared to treatment with allopurinol or glutathione individually. Glutathione can significantly reduce the thickness of the important dermis layer that helps control psoriasis.

[0049] In view of the contradictory and confusing prior art reports on the oral administration of allopurinol in the treatment of psoriasis, allopurinol and oxypurinol, a metabolite of allopurinol and a direct inhibitor of xanthine oxidase, were tested for topical application.The problem of reduced GI uptake of oxypurinol compared to allopurinol is not a factor for topical application as it is for oral use.

[0050] As clinically noted in 1970, the deficiency of phosphoribosyl pyrophosphate (PRPP) caused by allopurinol but not oxypurinol would be expected to cause cytotoxicity and adversely affect the efficacy of allopurinol compared to oxypurinol, as implied by Fox et al. ("Depletion of Erythrocyte Phosphoribosylpyrophosphate in Man: A Newly Observed Effect of Allopurinol", The New England Journal of Medicine, Vol. 283, pp. 1177-1182 (1970)).

[0051] Surprisingly, in an in vitro model of human psoriasis, allopurinol control was dramatically more effective than oxypurinol in reducing cell proliferation without increasing cell death. Without being bound by theory, this may be a result of, rather than in spite of, the deficiency of PRPP caused by allopurinol.

[0052] Methotrexate has been taken as a model for a therapeutic composition that inhibits cell replication, and inhibition of cell replication correlates with response to methotrexate in the treatment of psoriasis. Unfortunately, methotrexate has the problem of having a narrow therapeutic window and being toxic when applied in excess.

[0053] Provided herein are methods and compositions for treating skin disorders, such as allergic skin reactions or skin cell proliferative disorders or conditions, e.g., proliferative skin disorders or conditions, e.g., psoriasis, keratoacanthoma, rosacea, keloids, hand-foot syndrome, and cancer of the skin. Additionally, disorders such as Dupuytren's contracture and allergies can be effectively treated with the compositions of the present disclosure.

[0054] Embodiments of the present disclosure, compared to concepts that fail to include one or more of the features disclosed herein, are capable of reducing the proliferation rate of cells associated with a skin cell proliferative disorder, preferentially reducing the proliferation rate of cells of a cell type having a higher proliferation rate in a treatment site that includes other cells of a cell type having a lower proliferation rate, alleviating or preventing one or more symptoms associated with a skin cell proliferative disorder, reducing the likelihood or severity of one or more symptoms associated with a skin cell proliferative disorder, reducing the frequency of recurrence of one or more symptoms associated with a skin cell proliferative disorder, or any combination thereof.

[0055] In a representative embodiment, the protective formulation limits cell proliferation without increasing cell death of targeted skin cells.Methotrexate has been used to treat certain skin diseases, but it limits cell proliferation by increasing cell death of skin cells and thereby limiting the number of viable cells.However, as disclosed herein, it has been observed in vitro that allopurinol limits skin cell proliferation with minimal increase in cell death, much more effectively than methotrexate, in a psoriasis-inducing environment.

[0056] In an exemplary embodiment, the protective formulation comprises a therapeutic amount of allopurinol or a pharma- ceutically acceptable salt thereof. In an exemplary embodiment, the protective formulation is topically applied at the site of the skin condition. In some embodiments, the skin condition is a cutaneous cellular proliferative disorder. In some exemplary embodiments, the therapeutic amount of allopurinol slows proliferation of cells associated with the skin cell proliferative disorder. In some embodiments, the protective formulation treats at least one symptom associated with the skin cell proliferative disorder. In some embodiments, the skin condition is an allergic skin reaction. In some embodiments, the protective formulation is a topical formulation.

[0057] In some embodiments, the protective formulation also includes a therapeutic amount of an adjuvant. In some embodiments, the adjuvant is an antioxidant. In some embodiments, the adjuvant is a skin permeation enhancer. In some embodiments, the adjuvant is a sulfur-containing antioxidant. In many embodiments, the adjuvant includes a thiol or prothiol. Suitable thiols include, but are not limited to, glutathione, N-acetylcysteine, cysteine, theanine, cystine, β-mercaptoethanol, and / or dithiothreitol. In some embodiments, the adjuvant provides a synergistic effect when used in combination with allopurinol, such as in a predetermined amount. In some embodiments, the adjuvant enhances the delivery of allopurinol to the treatment site. In some embodiments, the adjuvant disrupts disulfide bridges of keratin in skin plaques.

[0058] In some embodiments, the auxiliary agent is glutathione or a pharma- ceutically acceptable salt thereof. In some embodiments, glutathione softens the skin, improves oxidative stress in the stratum corneum, dissolves keratin polymers, reduces swelling, and / or promotes penetration of topical preparations into the skin. In some embodiments, glutathione increases the penetration of protective preparations into psoriasis plaques.

[0059] In some embodiments, the protective formulation further comprises one or more agents that promote stabilization and / or preservation of glutathione.

[0060] In some embodiments, the protective formulation comprises a therapeutic amount of glutathione or a pharma- ceutically acceptable salt thereof, and optionally a second active ingredient.

[0061]

[0023] Embodiments of the present disclosure further include pharmaceutical compositions comprising allopurinol and glutathione, and optionally further comprising one or more pharma- ceutically acceptable excipients. Such pharmaceutical compositions may be administered orally or may be configured to be delivered in any effective conventional dosage form, including, for example, immediate release, slow release and sustained release oral formulations, parenteral, topical, nasal, ophthalmic, optical, sublingual, rectal, and vaginal administration. The present disclosure further includes methods of treating a disease, comprising administering allopurinol and glutathione, such as, for example, a pharmaceutical composition comprising allopurinol, glutathione, and one or more pharma- ceutically acceptable excipients, to a patient in need of such treatment.

[0062] As discussed in detail in the Examples section, allopurinol has been observed in vitro to reduce both cell proliferation and cell death in a model of human psoriasis system. Topical treatment with allopurinol and glutathione has been observed in the Examples to treat human psoriatic plaques, human guttate psoriasis, Dupuytren's contracture, and allergic skin reactions. For human psoriatic plaques, treatment was curative. In all other cases, improvement was observed.

[0063] Ki-67 is a cell replication marker, and the lower the value, the lower the replication rate. p21 is a protein that is a marker for inhibition of cell proliferation. Thus, an increase in the ratio of p21 / Ki-67 indicates a decrease in cell replication. Figure 9 illustrates this ratio under several conditions, as described in Example 2.

[0064] Increasing allopurinol exposure by increasing the concentration of topical allopurinol from 0.5 mM to 3.0 mM did not increase the p21 / Ki-67 ratio, and the effect did not achieve statistical significance. However, applying allopurinol together with glutathione increased the p21 / Ki-67 ratio, and the difference also reached statistical significance. Thus, the effect of the combined administration of glutathione and allopurinol synergistically restricted cell replication.

[0065] The combination therapy of allopurinol and glutathione also reduced the thickness of the dermal layer tissue in the psoriasis skin model on the sixth day of treatment, compared to the thickness of the dermal layer on the second day of treatment. The reduction in the thickness of the dermal layer has been used to demonstrate the success of the treatment response in human psoriasis patients (see, for example, Phillips et al., "Dermal Reflectivity Determined by Optical Coherence Tomography is an Indicator of Epidermal Hyperplasia and Dermal Edemawithin Inflamed Skin", Journal of Biomedical Optics, Vol. 16, Art. 040503(2011)). Only the treatment with 3 mM concentration of allopurinol caused an increase in the dermal layer over this period (see Example 2).

[0066] Allopurinol, when compared to the standard of care methotrexate, was better at inhibiting cell replication and better maintaining cell viability in the HaCaT human cell monolayer model of psoriasis, as seen in Example 1, Figures 1 and 2.

[0067] Application of glutathione alone was shown to inhibit Ki-67.

[0068] On this basis, allopurinol, either alone or in combination with glutathione, was expected to be beneficial in the topical protective treatment of all agents that cause toxicity in the hands and feet but not in the relatively slow-growing non-palmar and non-plantar human skin, as well as in the topical protective treatment of any proliferative disease or condition whose treatment is by reducing cell proliferation at the site of disease or condition onset.

[0069] In summary, application of allopurinol unexpectedly inhibited cell replication while preserving cell viability in an in vitro monolayer model of psoriatic HaCaT human cells, i.e., a significant improvement over methotrexate, the standard drug for the treatment of psoriasis. Without wishing to be bound by theory, the inhibition of growth may reflect consumption of PRPP by hypoxanthine xanthine phosphoribiosyltransferase, as originally described by Nelson et al. ("Formation of nucleotides of (6-14C)allopurinol and (6-14C)oxipurinolin rat tissues and effects on uridine nucleotide pools", Biochem. Pharmacol.,Vol. 22, pp. 2003-2022, (1973)). Unexpectedly, in the MatTek tissue model of psoriasis, application of glutathione alone resulted in the strongest inhibition of expression of the cell replication marker Ki-67 compared with allopurinol and comparable to exposure to the standard psoriasis treatment, but inadequate vitamin D. Glutathione also inhibited expression of the inflammatory markers IL-17A, IL-23, TNF-α, and TNF-β, as well as inducing expression of the inhibitor of replication, p21.

[0070] The combination of allopurinol and glutathione caused a decrease in tissue thickness of the PSE dermis layer on day 6 of treatment compared to the dermis thickness on day 2 of treatment. Application of 3.0 mM allopurinol alone paradoxically caused an increase in the dermis layer over this period. This result was consistent with a synergistic effect of allopurinol plus glutathione to improve tissue homeostasis. The decrease in dermis layer thickness has previously been used to monitor successful treatment response in human psoriasis patients. A higher p21 / Ki-67 ratio was observed with the combination of glutathione and allopurinol treatment compared to allopurinol or glutathione treatment alone. Increasing allopurinol exposure from 0.5 mM to 3.0 mM also did not provide evidence of a beneficial response to increasing allopurinol exposure, similar to the results for dermis thickness, again consistent with a synergistic effect of glutathione and allopurinol.

[0071] Both allopurinol and glutathione have been used safely topically in humans, individually and separately. Allopurinol has been widely and safely administered orally in humans for the treatment of gout for over 50 years. Glutathione is an antioxidant and a normal cellular component present in cells at concentrations ranging from 1 to 10 mM. Topical use of allopurinol and glutathione at 3% twice daily for 7 weeks resulted in the disappearance of an area of ​​3 cm of psoriatic plaque that had been present for 2 years.

[0072] In some embodiments, allopurinol is in the form of a pharma- ceutically acceptable salt, which may provide one or more advantageous properties over allopurinol alone.

[0073] In some embodiments, the glutathione is in the form of a pharma- ceutically acceptable salt, which may provide one or more advantageous properties over glutathione alone.

[0074] The topical efficacy and superiority of allopurinol over oxypurinol is surprising and unexpected. Applicants are unaware of any references or rationale that suggests a topical efficacy superiority for allopurinol over oxypurinol.

[0075] Allopurinol has not been tested for the topical treatment of psoriasis, but it has been commonly observed with other treatments that allopurinol applied topically produces only about 2% to 3% incidence of rash. In the literature, it is almost unheard of for any drug applied to intact non-mucosal skin to cause severe skin reactions (see, for example, Sachs et al., "Anaphylaxis and toxic epidermal necrolysis or Stevens-Johnson syndrome after nonmucosal topical drug application: fact or fiction?", Allergy, Vol. 62, pp. 877-883 (2007)), suggesting that topical compositions carry little risk of severe systemic complications.

[0076] In some embodiments, the compositions and methods include allopurinol as an active ingredient in a protective formulation. In some embodiments, the compositions and methods include allopurinol as an active ingredient in a protective formulation in combination with the adjunct glutathione.

[0077] In some embodiments, the protective formulation is formulated for topical application to the skin, and the protective formulation may be usefully formulated as a topical formulation. Suitable topical formulations may include, but are not limited to, ointments, creams, lotions, pastes, aerosol sprays, roll-on liquids, sticks or pads, or aerosol foam (mousse) compositions. In some embodiments, the compositions and methods include topical delivery of protective formulations containing allopurinol as an active ingredient, as described in U.S. Patent No. 9,084,788, issued to Ford on July 21, 2015, entitled "Compositions and methods for treating and preventing dermatoses," which discloses compositions and methods for topical administration.

[0078] In some embodiments, the treatment comprises topically applying the protective formulation to the skin at least once a day, such as twice a day, In some embodiments, the treatment comprises continuing the topical application for at least 2 days, such as 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, or for a period of more than 3 months.

[0079] The preferred clinical composition of the protective formulation may depend on the tissue that is desired to be protected. Pharmaceutical formulation is a well-established technology in that therapeutic agents must be formulated and it is known that there are a myriad of variables involved in such formulation (see, for example, Allen ed., Remington: The Science and Practice of Pharmacy, 22nd ed., Pharmaceutical Press (2012); Allen, Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems, 11th ed., Wolters Kluwer (2017); Rowe et al., ed., Handbook of Pharmaceutical Excipients, 6th ed., Pharmaceutical Press (2009)). Therefore, it often remains a challenge to prepare clinical and commercial formulations and formulations with specific functionality.

[0080] Suitable topical formulations may be, for example, anhydrous, aqueous, or water-in-oil or oil-in-water emulsions. Suitable topical formulations may further comprise one or more pharma- ceutically acceptable carriers and various skin active agents. The amount of carrier may range from about 1% to about 99% by weight, preferably from about 5% to about 70% by weight, and optimally from about 10% to about 40% by weight. Among the useful carriers are emollients, water, inorganic powders, foaming agents, emulsifiers, fatty alcohols, fatty acids, or combinations thereof.

[0081] Emollients may be selected from polyols, esters and hydrocarbons. Suitable polyols for topical formulations may include propylene glycol, dipropylene glycol, polypropylene glycol, polyethylene glycol, sorbitol, hydroxypropyl sorbitol, hexylene glycol, 1,3-butylene glycol, 1,2,6-hexanetriol, glycerin, ethoxylated glycerin, propoxylated glycerin, xylitol or mixtures thereof.

[0082] Esters useful as emollients include alkyl esters of fatty acids having 10 to 20 carbon atoms. Methyl, isopropyl and butyl esters of fatty acids are possibly useful. Examples include hexyl laurate, isohexyl laurate, isohexyl palmitate, isopropyl palmitate, decyl oleate, isodecyl oleate, hexadecyl stearate, decyl stearate, isopropyl isostearate, diisopropyl adipate, diisohexyl adipate, dihexyldecyl adipate, diisopropyl sebacate, lauryl lactate, myristyl lactate and cetyl lactate. Particularly preferred are C 12 ~C 15 It is an alcohol benzoate ester.

[0083] Esters useful as emollients may also include alkenyl esters of fatty acids having from 10 to 20 carbon atoms, such as oleyl myristate, oleyl stearate, and oleyl oleate. Esters useful as emollients may also include ether esters, such as fatty acid esters of ethoxylated fatty alcohols.

[0084] Esters useful as emollients may include polyhydric alcohol esters, such as mono- and di-fatty acid esters of ethylene glycol, mono- and di-fatty acid esters of diethylene glycol, mono- and di-fatty acid esters of polyethylene glycol (200-6,000), polyfatty acid esters of polyglycerol, ethoxylated glyceryl monostearate, 1,3-butylene glycol monostearate, 1,3-butylene glycol distearate, polyoxyethylene polyol fatty acid esters, sorbitan fatty acid esters, and / or polyoxyethylene sorbitan fatty acid esters.

[0085] Esters useful as emollients may further include wax esters (such as, for example, beeswax, spermaceti, myristyl myristate, and / or stearyl stearate) and sterol esters (such as, for example, cholesterol fatty acid esters).

[0086] Suitable hydrocarbon carriers may include mineral oil, polyalphaolefins, petrolatum, isoparaffins, polybutenes, and / or mixtures thereof.

[0087] Inorganic powders may also be useful as carriers in topical formulations. Examples may include clay (such as montmorillonite, hectorite, laponite, and bentonite), talc, mica, silica, alumina, zeolites, sodium sulfate, sodium bicarbonate, sodium carbonate, calcium sulfate, and / or mixtures thereof.

[0088] Suitable topical formulations may contain aerosol propellants as a carrier or in addition to a carrier. Propellants may be based on volatile hydrocarbons such as propane, butane, isobutene, pentane, isopropane and mixtures thereof. PhillipsPetroleum (Bartlesville, Oklahoma) may be a source of such propellants under trademarks including A3, A32, A51, and / or A70. Halogenated carbons, including fluorocarbons, may be more widely employed as propellants.

[0089] Suitable topical formulations for administration to the skin may include an emulsifying agent as, or in addition to, a carrier.

[0090] Suitable emulsifiers may be selected from nonionic, anionic, cationic, and / or amphoteric emulsifiers. Suitable emulsifiers may be in an amount ranging from about 0.1% to about 20% by weight.

[0091] Suitable non-ionic emulsifiers are C 10 ~C 22Alkoxylated compounds based on fatty alcohols and acids and sorbitan may be included. Suitable raw materials may be available, for example, as polyoxypropylene polyoxyethylene copolymers sold under the trademark Neodol (ShellOil, Houston, TX), Pluronic (BASF, Ludwigshafen, Germany), and / or alkyl polyglycosides available from Henkel (Düsseldorf, Germany).

[0092] Suitable anionic emulsifiers may include fatty acid soaps, sodium lauryl sulfate, sodium lauryl ether sulfate, alkyl benzene sulfonates, mono- and di-alkyl acid phosphates, sarcosinates, taurates, and / or sodium fatty acyl isethionates.

[0093] Suitable amphoteric emulsifiers may include dialkylamine oxides and various types of betaines, such as cocamidopropyl betaine.

[0094] Suitable topical formulations may also contain preservatives, such as methylparaben and propylparaben, which are useful for preventing microbial contamination.

[0095] In some embodiments, the compositions and methods include oral delivery of a protective formulation comprising oxypurinol as an active ingredient, as described in U.S. Pat. No. 9,119,855, issued to Ford on Sep. 1, 2015, entitled "Compositions and methods for treatment of the side-effects associated with administration of cancer chemotherapeutic agents," which discloses compositions and methods for oral administration.

[0096] In some embodiments, each active ingredient may be present in the protective formulation in a weight / weight percentage of at least 0.01%, 0.05%, 1.0%, 1.5%, 2.0%, 2.5%, 3.5%, 4.0%, 4.5%, 5.0%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or even 50% or more, with intermediate values ​​being acceptable, but typically about 50%, 45%, 40%, 30%, 35%, 30%, 25%, 20%, 15%, 10%, 5%, 45%, 4.0%, 3.5%, 3.0%, 2.5%, 2.0%, 1.5%, 1.0%, and sometimes about 0.05% or less, or even 0.01% or less, with intermediate values ​​being acceptable.

[0097] In some embodiments, allopurinol is present in the composition in an amount by weight ranging from about 1% to about 5%, alternatively from about 2% to about 4%, alternatively from about 3%, or any value, range or subrange therebetween.

[0098] In some embodiments, glutathione is present in the composition in an amount by weight in the range of about 0.1% to about 5%, alternatively in the range of about 1% to about 5%, alternatively in the range of about 2% to about 4%, alternatively in the range of about 3%, or any value, range or subrange therebetween.

[0099] In some embodiments, the protective formulation is applied or delivered to the protective site to provide a sustained concentration of the active ingredient of at least 0.03 mM, alternatively 0.03 mM to 30 mM, alternatively 0.03 mM to 3 mM, alternatively 0.03 mM to 0.1 mM, alternatively at least 0.1 mM, alternatively 0.1 mM to 0.3 mM, alternatively at least 0.3 mM, alternatively 0.3 mM to 1 mM, alternatively at least 1 mM, alternatively 1 mM to 30 mM, alternatively 1 mM to 10 mM, alternatively 1 mM to 3 mM, alternatively at least 3 mM, alternatively 3 mM or less, alternatively 1 mM or less, alternatively 0.3 mM or less, alternatively 0.1 mM or less, or any value, range or subrange therebetween.

[0100] In some embodiments, the active ingredient comprises allopurinol or a pharma- ceutically acceptable salt thereof. In some embodiments, allopurinol is the primary active ingredient in the protective formulation. In some embodiments, allopurinol is the only active ingredient in the protective formulation. In some embodiments, the protective formulation is free or substantially free of other active ingredients. As used herein, substantially free refers to the amount of compound in the composition being less than 0.001 mM.

[0101] In these and other embodiments, the protective formulation may also include an adjuvant as an active ingredient. In some embodiments, the adjuvant provides a synergistic effect in combination with allopurinol. An example of such an adjuvant is glutathione.

[0102] The negligible toxicity of local therapeutic doses of allopurinol suggested significant benefits of topically and locally administering protective formulations containing therapeutic amounts of allopurinol to the relevant tissues of patients susceptible to skin cell proliferative disorders, the therapeutic dose comprising a sufficient therapeutic amount of allopurinol to slow cell proliferation in the relevant tissues after onset of the skin cell proliferative disorder.

[0103] In some embodiments, a protective formulation comprising a therapeutic amount of allopurinol is administered topically and locally to the relevant tissue of a patient to reduce and treat the symptoms of a skin cell proliferative disorder, the therapeutic dose comprising a therapeutic amount of allopurinol sufficient to slow cell proliferation in the relevant tissue and treat the skin cell proliferative disorder.

[0104] In some embodiments, the method of site-specifically reducing the rate of skin cell replication in a patient comprises topically applying or locally delivering an effective amount of allopurinol to the site on the patient's skin where reduced skin cell replication is desired.

[0105] Representative, non-limiting embodiments of the present disclosure are set forth in the enumerated sections.

[0106] Item 1. A topical pharmaceutical formulation comprising allopurinol or a pharma- ceutically acceptable salt thereof.

[0107] Item 2. A topical preparation comprising allopurinol or a pharma- ceutically acceptable salt thereof and a co-agent or a pharma- ceutically acceptable salt thereof.

[0108] Clause 3. A topical formulation as described in clause 2, wherein the adjuvant is a thiol.

[0109] Item 4. The topical formulation according to item 3, wherein the thiol is selected from the group consisting of glutathione, N-acetylcysteine, cysteine, theanine, cystine, β-mercaptoethanol and dithiothreitol.

[0110] Item 5. The topical formulation described in Item 4, wherein the thiol is glutathione.

[0111] Item 6. An external preparation according to any one of items 1 to 5, further comprising a carrier.

[0112] Item 7. The topical formulation according to any one of items 1 to 5, wherein the amount of allopurinol or a pharma- ceutically acceptable salt thereof is from about 1% by weight to about 10% by weight.

[0113] Item 8. The topical formulation described in item 7, wherein the thiol is glutathione or a pharma- ceutically acceptable salt thereof and is present in an amount of about 1% to about 10% by weight.

[0114] Item 9. The topical preparation according to any one of items 1 to 8, wherein the preparation is anhydrous, aqueous or emulsion.

[0115] Item 10. An external preparation according to any one of items 5 to 9, wherein the active ingredients are allopurinol and glutathione or pharma- ceutically acceptable salts thereof.

[0116] Item 11. An external preparation according to any one of items 1 to 9, further comprising one or more additional active ingredients.

[0117] Item 12. A method for treating a skin disease, comprising treating a patient in need of treatment with an topical preparation containing allopurinol or a pharma- ceutical acceptable salt thereof.

[0118] Item 13. A method for treating a skin disease, comprising treating a patient in need of treatment with the topical preparation described in any one of items 1 to 11.

[0119] Item 14. The method described in item 12 or 13, wherein the skin disease is a proliferative skin disease.

[0120] Item 15. The method of item 14, wherein the proliferative skin disease is psoriasis.

[0121] Clause 16. The method of clause 14, wherein the proliferative skin disease is selected from the group consisting of psoriasis, keratoacanthoma, rosacea, keloids, hand-foot syndrome and cancer of the skin.

[0122] Item 17. The method of item 13, wherein the skin disease is Dupuytren's contracture.

[0123] Item 18. The method according to item 13, wherein the skin disease is an allergic reaction.

[0124] Item 19. The method of item 18, wherein the allergic reaction is a skin allergic reaction.

[0125] Item 20. The method of item 14, wherein the proliferative skin disease is a proliferative disease of skin cells.

[0126] Item 21. The method of item 20, wherein the topical preparation slows proliferation of cells involved in a proliferative disorder of skin cells.

[0127] Item 22. The method of item 21, wherein the topical preparation alleviates at least one symptom of the skin cell proliferative disorder.

[0128] Clause 23. The method of any one of clauses 12 to 22, wherein the treatment is administered at least once per day.

[0129] Item 24. The method of item 23, wherein said treatment is administered for 30 days or more.

[0130] Item 25. A pharmaceutical composition comprising allopurinol and glutathione.

[0131] Item 26. The pharmaceutical composition of item 25, further comprising one or more pharma- ceutically acceptable excipients.

[0132] Section 27. A method for treating a disease comprising administering allopurinol and glutathione to a patient in need of treatment.

[0133] Item 28. A method for treating a disease, comprising administering the pharmaceutical composition described in item 25 or 26 to a patient in need of treatment. EXAMPLES

[0134] The present invention is further described in the context of the following examples, which are offered by way of illustration and not by way of limitation.

[0135] [Example 1] Human immortalized keratinocytes (HaCaT cells) purchased from AddexBio (San Diego, CA) were maintained in vitro in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and antibiotics (100 U / mL penicillin and 100 mg / mL streptomycin) and cultured in an incubator at 37°C with 5% CO2.

[0136] A psoriasis-like keratinocyte model was established by adding the M5 cocktail of cytokines known to be involved in the formation of the psoriatic phenotype (IL-17A, IL-22, Oncostatin M, IL-1α and TNF-α, each at a final concentration of 2.5 ng / mL) to the culture medium of HaCaT keratinocytes. This mixture has been shown to induce many characteristics of the psoriatic phenotype. As a control, this mixture was incubated for 72 hours.

[0137] HaCaT cells were cultured at a density of approximately 50% (1 × 10 4 Cells were seeded with 100 μl of 10 ...

[0138] Other HaCaT cells were incubated in vitro for 72 hours in the presence of M5 compositions that also contained 1.0 μM methotrexate, 0.5 mM allopurinol, 3.0 mM allopurinol, 0.5 mM oxypurinol, or 3.0 mM oxypurinol. Allopurinol, oxypurinol, and methotrexate were all obtained from Sigma (St. Louis, MO). Each was taken up in 1N NaOH solution and then diluted to the appropriate concentration in cell culture medium.

[0139] The number of live and dead cells was quantified by the MultiTox-Glo multiplex cytotoxicity assay (Promega, Madison, WI) according to the manufacturer's protocol. A dilution series of cells was used to generate a standard curve to calculate the number of cells in each well. The total cell number is the sum of live and dead cells.

[0140] Table 1 shows the live and dead cell counts of HaCaT cells at 0 hours, and the total and dead cell counts at 48 and 72 hours, with the standard deviations also listed. The total cell count is the sum of the live and dead cell counts. Methotrexate was the control drug therapy for topical psoriasis treatment at the standard 1 μM dose. The ideal outcome of cell culture is that the total cell count neither increases nor decreases over time. An increase in the total cell count suggests that the psoriasis phenotype is not suppressed. A decrease in the total cell count suggests cytotoxicity resulting from the treatment.

[0141] With regard to dead cells, the ideal outcome would be as close to zero dead cells as possible. An increase in the number of dead cells indicates cytotoxicity. Note that the lowest total cell count was observed for 3.0 mM allopurinol. Note that at 72 hours, 3.0 mM allopurinol had the lowest both total and dead cell counts. [Table 1]

[0142] The raw data in Table 1 was converted to the relative cell ratio data in Table 2. The cell ratios in Table 2 were obtained by dividing the total and dead cell counts in Table 1 by the live cell count at time zero, which was 9177. The ideal result for the total cell ratio is one that is close to one and does not increase or decrease over time. The ideal result for the dead cell ratio is one that is as close to zero as possible and does not increase over time.

[0143] The values ​​of total cell ratio in Table 2 are plotted in FIG. 1. For the control M5 treated cells 10, the total cell ratio was 1.89, i.e., an increase of 89% over the initial viable cell number during the 72-hour culture period. For 0.5 mM oxypurinol 20, the total cell ratio after 72 hours was also 1.86 of the initial viable cell number. For methotrexate 40, the total cell ratio after 72 hours was 1.26 of the initial viable cell number. For 3.0 mM oxypurinol 25, the total cell number after 72 hours was 1.55 of the initial viable cell number. For 0.5 mM allopurinol 30, the total cell number after 72 hours was 1.46 of the initial viable cell number. For 3.0 mM allopurinol 35, the total cell number after 72 hours was only 1.02 of the initial viable cell number, i.e., a negligible increase. [Table 2]

[0144] The values ​​of the dead cell ratio from Table 2 are plotted in FIG. 2. The value 50 of the control for the dead cell ratio after 72 hours is 0.56, i.e. 56% of the input number of live cells. Methotrexate 80 gives a higher ratio of 0.69, i.e. 69% of the input live cells are dead. Surprisingly, the percentage of dead cells is lower for allopurinol than for the control, oxypurinol and methotrexate. This percentage is even lower when the concentration of allopurinol is increased from 0.5 mM to 3.0 mM. For 0.5 mM allopurinol 70, the dead cell ratio is 0.44, and for 3 mM allopurinol 75, the dead cell ratio is only 0.22. Also, quite unexpectedly, the advantage of the active species oxypurinol is reduced, with a dead cell ratio of 0.52 for 0.5 mM oxypurinol 60 and 0.40 for 3.0 mM oxypurinol 65. Importantly, in contrast to methotrexate, allopurinol at 3.0 mM reduced cell proliferation without increasing cell death.

[0145] In summary, the results in Figures 1 and 2 for this widely used psoriasis model show that allopurinol can control cell proliferation much more effectively than both its metabolite oxypurinol and methotrexate, the standard drug in clinical use, making allopurinol a therapeutic option that can halt the psoriatic process.

[0146] [Example 2] The proliferative effects of allopurinol and / or glutathione, including vitamin D as a control, were tested in a MatTek human PSE culture system (SOR-300-FT) containing the stratum corneum layer, the cellular epidermis layer, and the dermis layer. Growth conditions for psoriatic tissue were in accordance with the proprietary conditions of the manufacturer, MatTek. The system contained a dermal layer of psoriasis origin covered by a layer of normal epidermal cells. The stratum corneum is a superficial keratinous layer that is not composed of cells. The cellular epidermis underlies the stratum corneum and covers the deep dermal layer. Only the dermal layer in this system was of psoriasis origin. Psoriasis was characterized by an increase in the thickness of all three layers.

[0147] Six MatTek human PSE cultures were exposed to four different treatment compositions for five days. The first treatment composition was a negative control. The second treatment composition contained 0.5 mM allopurinol. The third treatment composition contained 3.0 mM allopurinol. The fourth treatment composition contained 3.0 mM glutathione. The fifth treatment composition contained a combination of 3.0 mM allopurinol and 3.0 mM glutathione. The sixth treatment composition contained 100 nM vitamin D as a control. Allopurinol, glutathione, and vitamin D were all obtained from Sigma (St. Louis, MO). Each was taken up in 1N NaOH solution and then diluted to the appropriate concentration in cell culture medium.

[0148] FIG. 3 shows the MatTek Human PSE culture system after 5 days of exposure to the third therapeutic composition. FIG. 4 shows the MatTek Human PSE culture system after 5 days of exposure to the fifth therapeutic composition. FIG. 5A and FIG. 5B show the MatTek Human PSE culture system after 5 days of exposure to the sixth therapeutic composition. The images in FIG. 3, FIG. 4, FIG. 5A and FIG. 5B are at the same magnification. The dermis layer is the lighter layer and the epidermis layer is the darker layer. The photographs show that the thickness of the dermis layer is reduced in the presence of both glutathione and allopurinol (FIG. 4) compared to allopurinol alone (FIG. 3). FIG. 5A and FIG. 5B show the separation of the dermis and epidermis layers resulting from vitamin D treatment, possibly reflecting the known toxicity of vitamin D treatment.

[0149] The thickness of each of the three tissue layers of the skin was monitored and measured in a blinded manner by a board-certified and nationally respected pathologist after 2 days and 6 days of exposure to the treatment medium. The measurements obtained are shown in Table 3 and Figure 6.

[0150] Increased dermal thickness is a hallmark of psoriasis (see, for example, Phillips et al., "Dermal Reflectivity Determined by Optical Coherence Tomography is an Indicator of Epidermal Hyperplasia and Dermal Edema within Inflamed Skin", Journal of Biomedical Optics, Vol. 16, Art. 040503 (2011)) and is an unexpected result of glutathione treatment in Table 3. Table 3 shows that treatment with a combination of allopurinol and glutathione actually reduced dermal thickness from days 2 to 6. It is interesting that the effects of allopurinol and glutathione are involved in the specific reversal of the thickness of the psoriatic dermal layer. Furthermore, treatment with both allopurinol and glutathione preserved the integrity of the tissue. Treatment with the active agent vitamin D resulted in separation of the thinned dermal layer from the epidermal layer, which may be evidence of toxicity, a known side effect of vitamin D therapy. [Table 3]

[0151] Figure 6 shows that 3.0 mM allopurinol alone 100 caused a 1.4-fold increase in dermal thickness during this time interval from day 2 to day 6 of treatment. In contrast, treatment with a combination of 3.0 mM allopurinol and 3.0 mM glutathione 110 reduced dermal thickness to 0.83-fold during the same time interval. The control 90 showed no dermal thickening. As seen in Figure 6, increasing from 0.5 mM allopurinol 105 to 3 mM allopurinol 110 significantly increased dermal thickening on day 6. Thus, there is a specific effect of the combination of allopurinol and glutathione to reverse the psoriasis dermal thickening seen with 3.0 mM allopurinol treatment alone.

[0152] Furthermore, tissue integrity was maintained by treatment with both allopurinol and glutathione: combined treatment with 3 mM allopurinol and glutathione resulted in no toxicity to keratinocytes (dyskeratosis) and reduced presence of cells in the keratin layer (parakeratosis) after 6 days compared to treatment with allopurinol alone.

[0153] The amount of Ki-67 in the MatTek tissue culture system was also measured. Ki-67 is a nuclear protein and a replication marker. Ki-67 levels were measured by polymerase chain reaction (PCR) analysis using standard protocols. The relative levels of Ki-67 measured on day 6 are shown in Table 4 and Figure 7. The relative levels are normalized to the control Ki-67 measured levels on day 0, which were adjusted to 1.0.

[0154] Ki-67 levels are an indicator of the cell's replication rate, with lower values ​​indicating a slower replication rate. Table 4 shows that treatment with 3 mM allopurinol and a combination of 3 mM allopurinol and 3 mM glutathione reduced Ki-67 levels relative to the control, with the combination performing slightly better than 3 mM allopurinol alone and much better than 100 nM vitamin D. [Table 4]

[0155] Other markers of cell proliferation, namely (protein p21), and four markers of inflammatory signaling (IL-17A, IL-23A, TNF-α, and transforming growth factor beta (TGF-β)) were also measured in the MatTekPSE cultures after 2 days of treatment. Ki-67 is a nuclear protein and a replication marker. The levels of markers were measured by isolating total RNA with the Promega SV96 Total RNA Isolation Vacuum System (Promega, Madison, WI) according to the manufacturer's instructions. Total RNA was reverse transcribed using the ABI High Capacity cDNA Reverse Transcription Kit (Applied Biosystems, Foster City, CA). Quantitative real-time polymerase chain reaction (qPCR) was performed using SYBR Green PCR Master Mix (Applied Biosystems) according to the manufacturer's protocol and amplified with a StepOnePlus real-time PCR system.

[0156] The relative levels of the markers measured after two days of treatment are shown in Table 5 and in FIG. [Table 5]

[0157] Unexpectedly, as shown in Table 5 and Figure 8, after 2 days of treatment, the mRNA levels of Ki-67, IL-17A, IL-23A and TNF-α were significantly decreased in PSE cells treated with allopurinol or glutathione individually, while the expression of p21, a gene that inhibits cell replication, was increased. To highlight the effect on cell proliferation, the ratio of p21 to Ki-67 mRNA levels was calculated, and the results are shown in Figure 9. After 2 days of treatment, allopurinol alone, glutathione alone, the synergistic combination of allopurinol and glutathione, and vitamin D in parallel treatment all demonstrated a significant increase in the p21 / Ki-67 ratio, suggesting a decrease in cell replication.

[0158] [Example 3] The psoriatic plaque on the left shoulder had persisted for over two years. Figure 10A shows the plaque before treatment.

[0159] Approximately 200 μL of a treatment composition of 3% allopurinol and 3% glutathione suspended in a petrolatum-based ointment was applied twice daily to the psoriatic plaques for one month. Both allopurinol and glutathione were United States Pharmacopeia (USP) grade. Allopurinol was in the form of finely crushed tablets and mixed to the appropriate concentration in the petrolatum-based ointment. Glutathione was in the form of a powder that was first mixed sparingly with mineral oil and then mixed to the appropriate concentration in the petrolatum-based ointment. The petrolatum-based ointment also contained water, mineral oil, ceresin, lanolin alcohol, panthenol, glycerin, and bisabolol as inactive ingredients. Figure 10B shows that the left shoulder responded completely to the treatment and the plaques were no longer visible. No side effects of the treatment were observed or reported.

[0160] [Example 4] Dupuytren's contracture is typically an abnormal thickening of the skin on the palm of the hand at the base of the fingers. This thickening may cause the fingers to bend or turn laterally or palmarly. Figure 11A shows Dupuytren's contracture on the right fifth finger of a volunteer before treatment. Figure 11B shows the middle phalanx 120 of the little finger with thickened overlying skin 122, thickened skin overlying tissue of the distal head of the fifth metacarpal 124, and thickened skin on the proximal phalanx 126, characteristic of Dupuytren's contracture.

[0161] A therapeutic composition of about 200 μL of 3% allopurinol suspended in a petrolatum-based ointment was applied twice daily to the thickened skin for one month. The allopurinol was USP grade. The allopurinol was in the form of finely crushed tablets and mixed at the appropriate concentration in the petrolatum-based ointment. Figure 11B shows that the finger responded with some improved movement.

[0162] Thereafter, a therapeutic composition of about 200 μL of 3% allopurinol and 3% glutathione suspended in a petrolatum-based ointment was applied twice daily to the psoriasis plaque for another month. The glutathione was USP grade. The glutathione was in powder form, first mixed sparingly with mineral oil, and then mixed to the appropriate concentration in the petrolatum-based ointment. Figure 11C shows that after two months, the right fifth finger had reduced swelling and was functionally nearly normal. The right fifth finger was fully mobile, but remained shortened by about 1 cm compared to the left fifth finger after more than two months of treatment. No side effects of the treatment were observed or reported.

[0163] Progressive changes over four months of daily topical human use of 10% allopurinol and 10% glutathione suspended in a petrolatum-based ointment in the treatment of Dupuytren's contracture are manifested in a gradual but progressive decrease in tissue thickness over time. FIG. 11D shows that the middle phalanx 120 of the little finger shows continued contraction of the overlying skin 122. The skin overlying the tissue at the distal head of the fifth metacarpal 124 also shows skin contraction, as does the skin over the proximal phalanx 126. Mobility of the right fifth finger gradually improved, and it became at least as strong as the left (dominant) fifth finger. No side effects were noted.

[0164] [Example 5] FIG. 12A shows a widespread rash on the back of the calf leg that began as an allergic reaction approximately 2 months ago.

[0165] A therapeutic composition consisting of 3% allopurinol suspended in a petrolatum-based ointment was applied twice daily to the left calf, and a therapeutic composition consisting of 3% allopurinol and 3% glutathione suspended in a petrolatum-based ointment was applied twice daily to the right calf. Both allopurinol and glutathione were USP grade. Allopurinol was in the form of finely ground tablets and mixed to the appropriate concentration in the petrolatum-based ointment. Glutathione was in the form of a powder, sparingly mixed with mineral oil and then mixed to the appropriate concentration in the petrolatum-based ointment.

[0166] Figure 12B shows the calves after 5 days of treatment. The right calf treated with a combination of 3% allopurinol and 3% glutathione shows more improvement than the left calf treated with only 3% allopurinol.

[0167] The recurrent rash responded rapidly to further treatment with 3% allopurinol and 3% glutathione applied twice daily: the itching resolved within 24 hours after further treatment and the swelling began to decrease within 48 hours and remained responsive thereafter.

[0168] [Example 6] FIG. 13A shows three areas of guttate psoriasis, a more acute form of psoriasis than plaque psoriasis, before treatment.

[0169] No treatment was applied to the leftmost area of ​​guttate psoriasis. A treatment composition of 3% allopurinol suspended in a petrolatum-based ointment was applied twice daily to the central area of ​​guttate psoriasis. A treatment composition of 3% allopurinol and 3% glutathione suspended in a petrolatum-based ointment was applied twice daily to the rightmost area of ​​guttate psoriasis. Both allopurinol and glutathione were USP grade. Allopurinol was in the form of finely crushed tablets and mixed to the appropriate concentration in a petrolatum-based ointment. Glutathione was in the form of a powder and mixed sparingly with mineral oil and then mixed to the appropriate concentration in a petrolatum-based ointment.

[0170] Figures 13B, 13C, 13D and 13E show the three regions 1.5 days, 2 days, 2.5 days and 6.5 days after the start of treatment.

[0171] FIG. 13C shows that after two days of treatment, the lesions in the central area treated with both allopurinol and glutathione had improved significantly and were nearly flattened.

[0172] [Example 7] FIG. 14A shows an allergic reaction with fluid dripping from both legs prior to treatment.

[0173] A therapeutic composition of 3% allopurinol suspended in a petrolatum-based ointment was applied twice daily to the individual's left foot, and a therapeutic composition of 3% allopurinol and 3% glutathione suspended in a petrolatum-based ointment was applied twice daily to the individual's right foot for seven days. Both allopurinol and glutathione were USP grade. Allopurinol was in the form of finely crushed tablets and mixed to the appropriate concentration in the petrolatum-based ointment. Glutathione was in the form of a powder, mixed sparingly with mineral oil, and then mixed to the appropriate concentration in the petrolatum-based ointment.

[0174] Figure 14B shows that after 7 days of treatment, the treated skin on both legs showed significant improvement. Although not quantitatively measured, the treated right leg that received a treatment composition containing both allopurinol and glutathione showed a dramatic improvement in the definition of the right tibial bone compared to pre-treatment.

[0175] All documents mentioned herein are hereby incorporated by reference.

[0176] Although the present invention has been described with reference to one or more embodiments, those skilled in the art will recognize that various modifications may be made and equivalents may be substituted for elements thereof without departing from the scope of the present invention. In addition, many modifications may be made to adapt a particular situation or material to the teachings of the present invention without departing from the essential scope of the invention. Therefore, it is not intended that the invention be limited to the particular embodiment disclosed as the best mode contemplated for carrying out the invention, but the invention is intended to include all embodiments falling within the scope of the appended claims. In addition, all numerical values ​​specified in the detailed description should be interpreted as if both the exact value and the approximate value were explicitly specified.

Claims

1. A topical preparation comprising allopurinol or a pharmaceutically acceptable salt thereof, and an adjuvant or a pharmaceutically acceptable salt thereof.

2. 2. The topical preparation according to claim 1, wherein the adjuvant is a thiol or a pharmaceutically acceptable salt thereof.

3. 3. The topical preparation according to claim 2, wherein the thiol is selected from the group consisting of glutathione, N-acetylcysteine, cysteine, theanine, cystine, β-mercaptoethanol, and dithiothreitol.

4. 4. The topical preparation according to claim 3, wherein the thiol is glutathione.

5. The topical formulation of claim 1, further comprising a carrier.

6. 2. The topical formulation according to claim 1, wherein the amount of allopurinol or a pharmaceutically acceptable salt thereof is from about 1% to about 10% by weight.

7. 7. The topical formulation of claim 6, wherein the thiol is glutathione or a pharmaceutically acceptable salt thereof and is present in an amount of about 1% to about 10% by weight.

8. 2. The topical formulation according to claim 1, wherein the formulation is anhydrous, aqueous or emulsion.

9. 5. The topical preparation according to claim 4, wherein the active ingredients are allopurinol and glutathione or pharmaceutically acceptable salts thereof.

10. 10. The topical formulation of claim 1, further comprising one or more additional active ingredients.

11. The topical preparation according to claim 1, wherein the topical preparation is for treating a skin disease.

12. The topical preparation according to claim 11, wherein the skin disease is a proliferative skin disease.

13. The topical preparation according to claim 12, wherein the proliferative skin disease is psoriasis.

14. 13. The topical preparation according to claim 12, wherein the proliferative skin disease is selected from the group consisting of psoriasis, keratoacanthoma, rosacea, keloids, hand-foot syndrome and cancer of the skin.

15. The topical preparation according to claim 11, wherein the skin disease is Dupuytren's contracture.

16. The topical preparation according to claim 11, wherein the skin disease is an allergic reaction.

17. The topical preparation according to claim 16, wherein the allergic reaction is a skin allergic reaction.

18. The topical preparation according to claim 12, wherein the proliferative skin disease is a proliferative disease of skin cells.

19. 19. The topical preparation according to claim 18, wherein the topical preparation slows the proliferation of cells involved in the skin cell proliferative disorder.

20. 20. The topical preparation of claim 19, wherein the topical preparation alleviates at least one symptom of the skin cell proliferative disorder.

21. The topical preparation according to claim 11, wherein the treatment is administered at least once per day.

22. 22. The topical preparation according to claim 21, wherein the treatment is administered for 30 days or more.

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