Pharmaceutical composition for hailey-hailey disease and use thereof

Abstract

The present invention provides a novel pharmaceutical composition for Hailey-Hailey disease.　The pharmaceutical composition for Hailey-Hailey disease according to the present invention contains 5-methyl-2-(1-piperazinyl)benzenesulfonic acid. The compound is used as an active ingredient in the form of an anhydride thereof, a salt thereof, a hydrate or solvate thereof, or a hydrate or solvate of the salt thereof. The hydrate is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate, for example. The pharmaceutical composition according to the present invention enables the treatment or prevention of Hailey-Hailey disease.

Description

Pharmaceutical compositions for Hailey-Hailey disease and uses thereof

[0001] The present invention relates to a pharmaceutical composition for Hailey-Hailey disease and a method for treating or preventing Hailey-Hailey disease.

[0002] Hailey-Hailey disease is a skin disorder characterized by epidermal acantholysis, which frequently causes blister-like eruptions (vesicles) in intertriginous areas such as the neck, axillae, and groin (Non-Patent Document 1). The blisters are accompanied by itching, and when the condition worsens, they rupture, exposing the mucous membrane and forming erosions. These conditions are often accompanied by crusts, pustules, pigmentation, or secondary infection, resulting in a condition similar to impetigo and producing a foul odor. The risk of chronicity and recurrence is high, leading to a decline in patients' quality of life (QOL) and potential social handicaps.

[0003] Currently, there is no fundamental treatment for Hailey-Hailey disease recommended by guidelines, and treatment focuses on disease management and symptom management by avoiding triggers that exacerbate the condition. For example, patients may receive lifestyle advice to reduce factors that worsen symptoms, such as friction, sweating, and exposure to sunlight. In mild cases, treatment involves the administration of steroid ointments or vitamin D preparations, while in severe cases, treatment involves the administration of immunosuppressants or retinoids. Laser treatment and surgical exfoliation may also be performed.

[0004] Treatments such as retinoids, steroids, and vitamin D preparations have been reported to have side effects, and none are safe and consistently effective. Therefore, there remains a high unmet medical need for safe and effective treatments for Hailey-Hailey disease.

[0005] Guidelines for the treatment of familial benign chronic pemphigus 2023, Journal of the Japanese Dermatological Association: 134(2), 273-287, 2024

[0006] Therefore, an object of the present invention is to provide a novel pharmaceutical composition and a method for treating or preventing Hailey-Hailey disease.

[0007] The pharmaceutical composition for Hailey-Hailey disease of the present invention comprises 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (5-methyl-2-(piperazin-1-yl)benzenesulfonic acid).

[0008] The method of the present invention for treating or preventing Hailey-Hailey disease comprises the step of administering 5-methyl-2-(1-piperazinyl)benzenesulfonic acid to a subject.

[0009] The present invention is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid for use in the treatment or prevention of Hailey-Hailey disease.

[0010] The present invention is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid in the preparation of a pharmaceutical composition for Hailey-Hailey disease.

[0011] According to the pharmaceutical composition of the present invention, by containing 5-methyl-2-(1-piperazinyl)benzenesulfonic acid, it is possible to treat or prevent Hailey-Hailey disease.

[0012] Figure 1A is a graph showing the relationship between administration of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate and expression of differentiation marker molecules in an in vitro HHD three-dimensional cultured epidermal model in Example 1, with the vertical axis showing relative gene expression levels based on the gene expression levels in keratinocytes on Day 0. Figure 1B is a graph showing the relationship between administration of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate and expression of differentiation marker molecules in an in vitro HHD three-dimensional cultured epidermal model in Example 1, with the vertical axis showing the rate of suppression of increased gene expression of differentiation markers by ATP2C1 siRNA treatment. Figure 2 is a graph showing the relationship between administration of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate and intercellular adhesion in an in vitro epidermal cell-cell adhesion disorder model in Example 2.

[0013] The present invention can be exemplified by the following embodiments. [1] A pharmaceutical composition for Hailey-Hailey disease, comprising 5-methyl-2-(1-piperazinyl)benzenesulfonic acid. [2] The pharmaceutical composition according to [1], wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a solvate of the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid, or a hydrate or solvate of the salt thereof. [3] The pharmaceutical composition according to [1], wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a hydrate of the salt thereof. [4] The pharmaceutical composition according to [2] or [3], wherein the hydrate is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate. [5] The pharmaceutical composition according to any one of [1] to [4], which suppresses acantholysis in Hailey-Hailey disease. [6] The pharmaceutical composition according to any one of [1] to [5], which improves the IGA score in patients with Hailey-Hailey disease. [7] The pharmaceutical composition according to any one of [1] to [6], which suppresses or improves at least one symptom selected from the group consisting of lesion skin area, itching, pain, and odor in patients with Hailey-Hailey disease. [8] The pharmaceutical composition according to any one of [1] to [7], wherein the dosage of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is 1 to 1000 mg / day, and the dosage is calculated as the amount of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid anhydride.

[0014] [9] A method for treating or preventing Hailey-Hailey disease, comprising a step of administering 5-methyl-2-(1-piperazinyl)benzenesulfonic acid to a subject.

[10] The method according to [9], wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate or solvate thereof, or a hydrate or solvate of a salt thereof.

[11] The method according to [9], wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a hydrate of a salt thereof.

[12] The pharmaceutical composition according to

[10] or

[11] , wherein the hydrate is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate.

[13] The method according to any one of [9] to

[12] , wherein the administration method is oral administration.

[14] The method according to any one of [9] to

[13] , wherein the daily dose is 1 to 1000 mg / day, and the dose is calculated as the anhydride of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid.

[15] The method according to any one of [9] to

[14] , wherein the number of times of administration per day is 1 to 3 times.

[16] The method according to any one of [9] to

[15] , wherein the administration interval is every day.

[0015]

[17] 5-methyl-2-(1-piperazinyl)benzenesulfonic acid for use in the treatment or prevention of Hailey-Hailey disease.

[18] The 5-methyl-2-(1-piperazinyl)benzenesulfonic acid according to

[17] , wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a solvate of the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid, or a hydrate or solvate of the salt thereof.

[19] The 5-methyl-2-(1-piperazinyl)benzenesulfonic acid according to

[17] , wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a hydrate of the salt thereof.

[20] The 5-methyl-2-(1-piperazinyl)benzenesulfonic acid according to

[18] or

[19] , wherein the hydrate is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate.

[21] 5-methyl-2-(1-piperazinyl)benzenesulfonic acid for use in the manufacture of a pharmaceutical composition for Hailey-Hailey disease.

[22] The 5-methyl-2-(1-piperazinyl)benzenesulfonic acid according to

[21] , wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a solvate of the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid, or a hydrate or solvate of the salt.

[23] The 5-methyl-2-(1-piperazinyl)benzenesulfonic acid according to

[21] , wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a hydrate of the salt.

[24] The 5-methyl-2-(1-piperazinyl)benzenesulfonic acid according to

[22] or

[23] , wherein the hydrate is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate.

[0016]

[25] A pharmaceutical composition for acantholysis comprising 5-methyl-2-(1-piperazinyl)benzenesulfonic acid.

[26] The pharmaceutical composition according to

[25] , wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate or solvate thereof, or a hydrate or solvate of a salt thereof.

[27] The pharmaceutical composition according to

[25] , wherein the 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is an anhydride, a salt, a hydrate, or a hydrate of a salt thereof.

[28] The pharmaceutical composition according to

[26] or

[27] , wherein the hydrate is 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate.

[0017] As used herein, "treatment of a disease" includes, for example, curing a disease, remission of a disease, alleviating a disease, or suppressing the progression of a disease, and "treatment of a disease" also includes, for example, treating symptoms caused by the disease (e.g., curing, remission, alleviating, or suppressing symptoms). "Prevention of a disease" includes, for example, preventing contraction of a disease, preventing the onset of a disease, and preventing the recurrence of a disease, and "prevention of a disease" also includes, for example, preventing symptoms caused by the disease (e.g., preventing the onset of symptoms or preventing the recurrence of symptoms). As used herein, "treatment or prevention of a disease" can also be referred to, for example, as suppression of a disease, and "treatment or prevention of symptoms caused by a disease" can also be referred to, for example, as suppression of a disease.

[0018] The "epidermis" of the skin is usually composed of the stratum corneum, stratum granulosum, stratum spinosum, and stratum basale.

[0019] As used herein, treatment refers to, for example, treatment for a subject diagnosed by a physician as having a disease or a symptom of a disease. Furthermore, as used herein, prevention refers to, for example, treatment for a subject who does not have a disease or a symptom of a disease, and refers to treatment aimed at preventing the onset of a disease or a symptom of a disease. The subject may also be referred to as, for example, a test subject or a subject, and may be a human (patient) or a non-human animal (animal patient). As used herein, the term "patient" may also include, for example, the meaning of a non-human animal patient, and may be interpreted as "animal patient." Furthermore, if the subject does not have the disease or its symptoms, they are also referred to as a healthy subject (a healthy person or a healthy non-human animal) with respect to the disease or symptom of interest.

[0020] The present invention will be described below with reference to specific examples, but the present invention is not limited to these examples. The embodiments exemplified below can be mutually incorporated unless otherwise specified. Hereinafter, Hailey-Hailey disease will also be referred to as HHD.

[0021] [Pharmaceutical Composition for HHD] The pharmaceutical composition for HHD of the present invention is characterized by containing 5-methyl-2-(1-piperazinyl)benzenesulfonic acid. The pharmaceutical composition for HHD of the present invention is characterized by containing the compound, and other components and conditions are not particularly limited.

[0022] As described above, the pharmaceutical composition for HHD of the present invention can treat or prevent HHD. The pharmaceutical composition for HHD of the present invention can be used, for example, for the purpose of treatment, for the purpose of prevention, or for both the purpose of treatment and prevention. Hereinafter, in this specification, the term "treatment / prevention" can be interpreted as meaning either treatment, prevention, or both treatment and prevention. The pharmaceutical composition for HHD of the present invention can, for example, provide safe treatment / prevention with few side effects.

[0023] Hereinafter, 5-methyl-2-(1-piperazinyl)benzenesulfonic acid will be referred to as "MPBS." The form of MPBS is not limited, and may include an anhydrous MPBS (also referred to as "MPBS anhydrous"), a salt of MPBS (also referred to as "MPBS salt"), a hydrate of MPBS or a hydrate of the MPBS salt (also referred to as "MPBS hydrate"), and a solvate of MPBS or a solvate of the MPBS salt (also referred to as "MPBS solvate"). These are collectively referred to as the "MPBSs" of the present invention. The MPBSs contained in the pharmaceutical composition of the present invention may be, for example, the MPBS anhydrous, the MPBS salt, the MPBS hydrate, or the MPBS solvate. It may contain only one of these, or two or more of these. The pharmaceutical composition of the present invention preferably contains the MPBSs as an active ingredient.

[0024] The type of MPBS salt is not particularly limited and may include, for example, alkali metal salts, alkaline earth metal salts, amphoteric element salts, amine salts, inorganic acid salts, or organic acid salts. Examples of the alkali metal salts include sodium salts or potassium salts, examples of the alkaline earth metal salts include magnesium salts or calcium salts, and examples of the amphoteric element salts include aluminum salts. Examples of the amine salts include lower alkylamine salts such as triethylamine salts; hydroxy lower alkylamine salts such as 2-hydroxyethylamine salts, bis-(2-hydroxyethyl)amine salts, tris(hydroxymethyl)aminomethane salts, and N-methyl-D-glucamine salts; cycloalkylamine salts such as dicyclohexylamine salts; benzylamine salts such as N,N-dibenzylethylenediamine salts; and dibenzylamine salts. Examples of the inorganic acid salts include hydrochloride, hydrobromide, sulfate, and phosphate, and examples of the organic acid salts include fumarate, succinate, oxalate, and lactate.

[0025] The type of MPBS hydrate is not particularly limited and may be, for example, a monohydrate. The type of MPBS solvate is not particularly limited. The solvent that can form the MPBS solvate is not particularly limited and may be, for example, a non-aqueous solvent, specific examples of which include alcohols such as methanol, ethanol, and isopropyl alcohol, acetone, ethyl acetate, and methylene chloride.

[0026] In the pharmaceutical composition for HHD of the present invention, the MPBS is preferably the anhydrous MPBS or the hydrated MPBS, more preferably the hydrated MPBS, and specifically the monohydrate MPBS.

[0027] The MPBSs are known compounds that can be synthesized by the methods described in, for example, JP-A-3-7263, JP-A-9-221479, EP-A-390654, EP-A-779283, U.S. Pat. No. 5,053,409, and U.S. Pat. No. 5,990,113, and are readily available to those skilled in the art.

[0028] The MPBSs can be substituted with a compound represented by the following general formula (I), or a salt thereof, or a hydrate or solvate thereof, which is described in International Publication WO03 / 011296.

[0029] In the formula, R １ is a hydrogen atom, C １ -C ６ alkyl group of C ３ -C ７ a cycloalkyl group of C １ -C ４ a halogenated alkyl group, a halogen atom, or C ６ -C １２ represents an aryl group represented by the formula: R ２ is a hydrogen atom, C １ -C ６ or an alkyl group of C ７ -C １２ The aralkyl group is an aralkyl group represented by the formula: １ -C ６ an alkoxy group, a halogen atom, C１ -C ６ and an amino group; n represents an integer of 1 to 4.

[0030] In the general formula (I), R １ C defined as １ -C ６ Examples of the alkyl group include a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, a sec-butyl group, a tert-butyl group, a pentyl group, an isopentyl group, a neopentyl group, a tert-pentyl group, a hexyl group, and an isohexyl group. ３ -C ７ Examples of the cycloalkyl group include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group, and a cycloheptyl group. １ -C ４ Examples of the halogenated alkyl group include a trifluoromethyl group, a trifluoroethyl group, and a pentafluoroethyl group. Examples of the halogen atom include a fluorine atom, a chlorine atom, and a bromine atom. ６ -C １２ Examples of the aryl group include a phenyl group and a naphthyl group.

[0031] R １ Preferred examples of the group include a hydrogen atom, C １ -C ６ alkyl group of C ５ -C ６ A cycloalkyl group represented by the formula (I), a trifluoromethyl group, a halogen atom, or a phenyl group is preferred. １ -C ３ Examples of the alkyl group include the above-mentioned groups, a cyclohexyl group, a trifluoromethyl group, a chlorine atom, a bromine atom, and a phenyl group. More preferred examples include a methyl group or a propyl group, and a methyl group is particularly preferred.

[0032] R ２ C defined as １ -C ６ Examples of the alkyl group in the above R １Examples of the alkyl group include those defined in ７ -C １２ Examples of the aralkyl group include a benzyl group, a phenethyl group, and a naphthylmethyl group. Examples of the aralkyl group include a C aryl group such as a cyano group, a nitro group, a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, a butoxy group, an isobutoxy group, a tert-butoxy group, a pentyloxy group, an isopentyloxy group, a tert-pentyloxy group, and a hexyloxy group. １ -C ６ an alkoxy group of the above R １ a halogen atom as defined above; １ and an amino group.

[0033] R ２ Preferred examples of the group include a hydrogen atom, C １ -C ３ and alkyl groups of C ７ -C １２ The aralkyl group is an aralkyl group represented by the formula: １ -C ３ alkyl group of C １ -C ３ and a halogen atom. More preferred examples include a hydrogen atom, ７ -C １２ The aralkyl group is an aralkyl group represented by the formula: １ -C ３ In particular, a hydrogen atom is preferred. In the above general formula (I), n is preferably 2.

[0034] In the pharmaceutical composition for HHD of the present invention, the MPBSs may be, for example, in an ionized form. When the pharmaceutical composition for HHD of the present invention is, for example, a liquid and contains an aqueous solvent, a non-aqueous solvent, or a mixture thereof, as described below, the MPBSs may be ionized regardless of the type of anhydrate, salt, hydrate, or solvate. The molecular form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is, for example, represented by the following formula (II), and can become an ionized molecular species in a protic solvent such as water. Specific examples of the molecular species include a monovalent cation represented by the following formula (IIIa), a zwitterion (zwitterion) having a positive charge and a negative charge represented by the following formula (IIIb), and a monovalent anion represented by the following formula (IIIc). The monovalent cation molecular species can form a salt with, for example, an acid (anion), and the monovalent anion can form a salt with, for example, a base (cation).

[0035]

[0036] The pharmaceutical composition for HHD of the present invention can be used, for example, for the treatment or prevention of HHD, and the treatment and prevention are, for example, as described above. The pharmaceutical composition for HHD of the present invention can be interpreted, for example, as a pharmaceutical composition used for the symptoms of HHD, and specifically, as a pharmaceutical composition used for the treatment or prevention of the symptoms of HHD. Furthermore, the treatment or prevention of HHD can also be said, for example, to be the suppression of HHD.

[0037] HHD, the subject of the present invention, is a disease in which acantholysis occurs as a pathological symptom. Acantholysis is a condition in which keratinocytes lose intercellular adhesion, for example, in the spinous layer of the skin, and can be observed, for example, as intraepidermal fissures or blister formation. Furthermore, acantholytic cells (Tzanck cells), which are floating, round keratinocytes with pyknotic nuclei, can be observed within the blisters. It is known that, for example, abnormal differentiation of keratinocytes causes acantholysis in the lesions of HHD patients. Acantholysis can be observed, for example, by collecting lesioned skin from a patient and following methods known to those skilled in the art.

[0038] The pharmaceutical composition for HHD of the present invention can, for example, suppress the pathological symptoms of HHD. The pathological symptoms to be suppressed are, for example, acantholysis, and may further include dyskeratinization in addition to acantholysis. That is, the pharmaceutical composition for HHD of the present invention may, for example, suppress acantholysis, or may suppress acantholysis and dyskeratinization. The pharmaceutical composition for HHD of the present invention can, for example, suppress acantholysis, and can suppress acantholysis and dyskeratinization.

[0039] Dyskeratinization refers to the individual keratinization of keratinocytes in the spinous layer of the skin, and in the case of HHD, these can be observed as granular bodies with pyknotic nuclei or acantholytic cells. Dyskeratinization can be observed, for example, by taking a sample of lesional skin from a patient and using methods known to those skilled in the art.

[0040] The pharmaceutical composition for HHD of the present invention can improve, for example, the patient's IGA (Investigator's Global Assessment) score. The IGA score is not particularly limited. IGA scores are generally classified into multiple stages, with higher scores indicating more severe symptoms.

[0041] The pharmaceutical composition for HHD of the present invention can suppress or improve, for example, clinical physical symptoms caused by HHD, specifically, for example, at least one selected from the group consisting of lesion skin area, pruritus (itching), pain (pain), and odor in HHD. The pharmaceutical composition for HHD of the present invention can suppress or improve, for example, the aforementioned pathological symptoms, thereby suppressing or improving the clinical physical symptoms exemplified herein.

[0042] The lesional skin area is, for example, the area where erythema consisting of clusters of small blisters is observed. This erythema resembles impetigo, in which the blisters break and become erosions, and are accompanied by crusts, pustules, pigmentation, or secondary infection. The pharmaceutical composition for HHD of the present invention can suppress or improve the lesional skin area to, for example, 90% or less, 80% or less, 70% or less, 60% or less, 50% or less, 40% or less, 30% or less, 20% or less, or 10% or less compared to before administration.

[0043] Itching can be evaluated, for example, by measuring the itching felt by a subject using a numerical rating scale for itching. For example, 0 indicates no itching and 10 indicates the worst itching imaginable to the subject, and the subject evaluates the itching using this numerical rating scale. The pharmaceutical composition for HHD of the present invention can reduce the value on the numerical rating scale for itching after administration by, for example, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, or 9 or more, compared to the value before administration.

[0044] Pain can be evaluated, for example, by using a numerical rating scale for pain, where the pain felt by the subject is evaluated using a numerical rating scale, where 0 indicates no skin pain and 10 indicates the worst skin pain the subject can imagine. The pharmaceutical composition for HHD of the present invention can reduce the value of the numerical rating scale for pain after administration by, for example, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, or 9 or more, compared to before administration.

[0045] The odor can be evaluated, for example, by using a numerical rating scale for odor, where the odor perceived by the subject is evaluated by the subject using a numerical rating scale, where 0 indicates no odor and 10 indicates the worst odor the subject can imagine. The pharmaceutical composition for HHD of the present invention can reduce the value on the numerical rating scale for odor after administration by, for example, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, or 9 or more, compared to the value before administration.

[0046] The pharmaceutical composition for HHD of the present invention improves, for example, at least one selected from the group consisting of Patient Global Impression of Severity (PGIS), Patient Global Impression of Change (PGIC), Clinician Global Impression of Severity (CGIS), Clinician Global Impression of Change (CGIC), Dermatology Life Quality Index (DLQI), and Skindex-29 in patients with HHD.

[0047] (1) The PGIS is a single questionnaire that evaluates a subject's overall impression of the severity of their condition using a score, and the pharmaceutical composition for HHD of the present invention can improve the PGIS and improve the score. (2) The PGIC is a single questionnaire that evaluates a subject's overall improvement in health status using a score, and the pharmaceutical composition for HHD of the present invention can improve the PGIC and improve the score. (3) The CGIS is a single questionnaire that evaluates a subject's overall impression of the severity of their condition using a score, and the pharmaceutical composition for HHD of the present invention can improve the CGIS and improve the score. (3) The CGIC is a single questionnaire that evaluates a subject's overall improvement in health status using a score, and the pharmaceutical composition for HHD of the present invention can improve the CGIS and improve the score. (4) The DLQI is a questionnaire that evaluates each question related to QOL using a score. The pharmaceutical composition for HHD of the present invention can improve the DLQI and improve the score. (5) Skindex-29 is a questionnaire in which subjects answer questions about their QOL and are evaluated by a score. The pharmaceutical composition for HHD of the present invention can improve Skindex-29 and thereby improve the score.

[0048] The method of administration of the pharmaceutical composition for HHD of the present invention is not particularly limited, and may be oral or parenteral administration, including, for example, transdermal, subcutaneous, intravenous, intraarterial, intraperitoneal, intranasal, and intraintestinal administration.

[0049] The dosage form of the pharmaceutical composition for HHD of the present invention is not particularly limited and can be appropriately determined depending on, for example, the method of administration. Examples of dosage forms include liquid, gel, cream, and solid. Examples of oral dosage forms include granules, fine granules, powders, tablets, capsules (e.g., hard capsules and soft capsules), syrups, emulsions, suspensions, liquids, and jellies. Examples of parenteral dosage forms include injections, suppositories, and transdermal agents.

[0050] The pharmaceutical composition for HHD of the present invention is not particularly limited in other components as long as it contains the MPBS. The pharmaceutical composition for HHD of the present invention may, for example, contain only the MPBS as an active ingredient, or may contain other active ingredients for HHD in addition to the MPBS.

[0051] The pharmaceutical composition for HHD of the present invention may contain, for example, only the active ingredient, or may further contain additives in addition to the active ingredient. The additives are preferably, for example, pharmaceutically acceptable substances. The type of additive is not particularly limited and can be appropriately selected depending on, for example, the dosage form. Examples of the additives include carriers, excipients, stabilizers, lubricants, sweeteners, preservatives, suspending agents, dispersants, thickeners, pH adjusters, antifoaming agents, and flavorings. Examples of the carriers include liquids, solids, gels, and creams.

[0052] The subject to which the pharmaceutical composition for HHD of the present invention is administered is not particularly limited and may be, for example, a human or a non-human animal, preferably a human. Non-human animals include non-human mammals such as mice, rats, rabbits, and horses.

[0053] The conditions for administering the pharmaceutical composition for HHD of the present invention are not particularly limited and can be determined appropriately depending on, for example, the administration method and patient information, such as age, sex, weight, presence or absence of HHD and its symptoms, the severity of HHD and its symptoms, and medical history.

[0054] When the pharmaceutical composition for HHD of the present invention is orally administered, the following conditions can be exemplified. The following examples can be adjusted appropriately for infants, young children, children, adults, elderly people, etc., depending on the patient's information, etc. The dose of the MPBSs can be expressed, for example, as a dose converted into the anhydrous MPBS. The doses exemplified below are doses of the MPBSs converted into the anhydrous MPBS, and as a specific example, for the MPBS monohydrate, they are doses converted into the anhydrous MPBS.

[0055] Daily dose of the MPBS (converted to anhydrous MPBS) Lower limit: For example, 1 mg, 10 mg, 20 mg, 25 mg, 30 mg, 50 mg, 60 mg, 100 mg, 150 mg, 200 mg, or 300 mg Upper limit: For example, 300 mg, 400 mg, 500 mg, 600 mg, 800 mg, 1000 mg, or 1200 mg Range: For example, 1 to 1000 mg, 10 to 1000 mg, 20 to 1000 mg, 25 to 1000 mg, 30 to 1000 mg, 50 to 1000 mg, 60 to 1000 mg, 100 to 1000 mg, 1 to 600 mg, 10 to 600 mg, 25 to 600 mg, 30 to 600 mg, 50 to 600 mg, 100 to 600 mg, 10 to 500 mg, 20 to 500 mg, 25 to 500 mg , 30-500mg, 50-500mg, 60-500mg, 100-500mg, 200-500mg, 10-400mg, 20-400mg, 25-400mg, 30-400m g, 50-400mg, 60-400mg, 100-400mg, 200-400mg, 10-300mg, 20-300mg, 25-300mg, 30-300mg, 50-300m g, 60-300mg, 100-300mg, 200-300mg, 10-200mg, 20-200mg, 25-200mg, 30-200mg, 50-200mg, 60-200 mg, 100-200mg, 10-100mg, 20-100mg, 25-100mg, 30-100mg, 50-100mg, 60-100mg, 300-400mg, 300-5 00 mg, 300-600 mg, 300-800 mg, 300-1000 mg, 300-1200 mg, 400-600 mg, 400-800 mg, 400-1000 mg, 400-1200 mg, 500-600 mg, 500-800 mg, 500-1000 mg, 500-1200 mg, 600-800 mg, 600-1000 mg, or 600-1200 mg Examples of specific dosages: for example, 1 mg, 10 mg, 20 mg, 25 mg, 30 mg, 40 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 700 mg, 800 mg, 900 mg, 1000 mg, or 1200 mg.Number of administrations per day: for example, 1 to 3 times, preferably 1 or 2 times, more preferably 2 times. Frequency of administration (interval): for example, daily.

[0056] When the pharmaceutical composition for HHD of the present invention is orally administered, the timing of administration can be freely set, for example, before meals, during meals, immediately after meals, after meals, between meals, upon waking up, or before bedtime, and administration between meals is preferred. Administration between meals of the pharmaceutical composition for HHD of the present invention means, for example, administration at least 1 hour, preferably at least 2 hours, after ingestion at mealtime, and / or, for example, administration at least 1 hour, 2 hours, 3 hours, 4 hours, or 4.5 hours, preferably 1 hour, after ingestion, before the next meal. That is, the pharmaceutical composition for HHD of the present invention is administered, for example, between 1 or 2 hours after ingestion of a meal and 1, 2, 3, 4, or 4.5 hours before ingestion of the next meal.

[0057] As described above, the pharmaceutical composition for HHD of the present invention can be used to treat or prevent the symptoms of HHD, and HHD is a disease that causes acantholysis as described above. Therefore, the present invention can also be referred to as, for example, a pharmaceutical composition for acantholysis that treats or prevents acantholysis. Furthermore, the present invention can also be referred to as, for example, a pharmaceutical composition for acantholysis and dyskeratosis that simultaneously treats or prevents acantholysis and dyskeratosis.

[0058] [Method for Treating or Preventing HHD] The method for treating or preventing HHD of the present invention comprises the step of administering 5-methyl-2-(1-piperazinyl)benzenesulfonic acid to a subject. As described above, the form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited, and may be any of the MPBSs, i.e., the MPBS anhydrate, the MPBS salt, the MPBS hydrate, and / or the MPBS solvate. Unless otherwise specified, the method for treating or preventing HHD of the present invention will hereinafter be referred to as the HHD treatment / prevention method of the present invention. The treatment / prevention method of the present invention is characterized by administering the MPBSs, and other conditions and steps are not particularly limited.

[0059] In the treatment and prevention methods of the present invention, the subject is a patient, and the patient may be, for example, a patient who has developed HHD or a patient who has not developed HHD.

[0060] In the treatment and prevention methods of the present invention, the administration of the MPBSs is, for example, the administration of the pharmaceutical composition for HHD of the present invention. In the present invention, the MPBSs, their compositions, and administration methods can be referenced from the descriptions of the pharmaceutical composition for HHD of the present invention.

[0061] [Use of MPBSs] The present invention relates to 5-methyl-2-(1-piperazinyl)benzenesulfonic acid for use in the treatment or prevention of HHD. As described above, the form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited, and it may be any of the MPBSs, i.e., the MPBS anhydrate, the MPBS salt, the MPBS hydrate, and / or the MPBS solvate. In the present invention, the descriptions of the pharmaceutical composition for HHD of the present invention can be used to refer to the MPBSs, compositions thereof, and methods of use thereof.

[0062] The present invention also relates to the use of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid in the manufacture of a pharmaceutical composition for HHD. As described above, the form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited, and it may be any of the MPBSs, i.e., the MPBS anhydrate, the MPBS salt, the MPBS hydrate, and / or the MPBS solvate. In the present invention, the descriptions of the pharmaceutical composition for HHD of the present invention can be used to refer to the MPBSs, compositions thereof, and methods of use thereof.

[0063] [Pharmaceutical Composition for Acantholysis and Its Use] The pharmaceutical composition for acantholysis of the present invention contains 5-methyl-2-(1-piperazinyl)benzenesulfonic acid. As described above, the form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited, and may be any of the MPBSs, i.e., the MPBS anhydrate, the MPBS salt, the MPBS hydrate, and / or the MPBS solvate. The pharmaceutical composition for acantholysis of the present invention may contain, for example, only one of the MPBSs or two or more of the MPBSs. The MPBSs can be used for the treatment or prevention of acantholysis. The pharmaceutical composition for acantholysis of the present invention is characterized by containing the MPBSs, and other conditions are not limited in any way. The pharmaceutical composition for acantholysis of the present invention preferably contains the MPBSs as an active ingredient. When the pharmaceutical composition for acantholysis of the present invention is used for the treatment or prevention of acantholysis, the type of disease that causes acantholysis as a pathological symptom is not particularly limited.

[0064] As described above, the pharmaceutical composition for acantholysis of the present invention can treat or prevent acantholysis. The pharmaceutical composition for acantholysis of the present invention can be used, for example, for treatment, prevention, or both treatment and prevention. Hereinafter, the term "treatment / prevention" can be interpreted as meaning either treatment, prevention, or both treatment and prevention. The pharmaceutical composition for acantholysis of the present invention can, for example, provide safe treatment / prevention with few side effects. An example of a disease that causes acantholysis is Hailey-Hailey disease.

[0065] In the pharmaceutical composition for acantholysis of the present invention, the MPBSs, their composition, and method of use are not particularly limited, and for example, the description of the pharmaceutical composition for HHD of the present invention can be used. When using this description, "HHD" and "symptoms of HHD" can be read as "acantholysis."

[0066] The method for treating or preventing acantholysis of the present invention includes administering 5-methyl-2-(1-piperazinyl)benzenesulfonic acid to a subject. As described above, the form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited, and may be any of the MPBSs, i.e., the MPBS anhydrate, the MPBS salt, the MPBS hydrate, and / or the MPBS solvate. Unless otherwise specified, the method for treating or preventing acantholysis of the present invention will hereinafter be referred to as the method for treating or preventing acantholysis of the present invention. The method for treating or preventing acantholysis of the present invention is characterized by administering the MPBSs; other conditions and steps are not particularly limited. The subject of the present invention may be, for example, a patient who has or may develop acantholysis as a pathological symptom, and the type of disease that causes acantholysis as a pathological symptom is not particularly limited.

[0067] In the present invention, the MPBSs, their compositions, and their administration methods are not particularly limited, and the descriptions of the pharmaceutical composition for HHD of the present invention can be used. When using these descriptions, "HHD" and "symptoms of HHD" can be read as "acantholysis."

[0068] The present invention relates to 5-methyl-2-(1-piperazinyl)benzenesulfonic acid for use in the treatment or prevention of acantholysis. As described above, 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited in form and may be any of the MPBSs, i.e., MPBS anhydrate, MPBS salt, MPBS hydrate, and / or MPBS solvate. In the present invention, the MPBSs, compositions thereof, and methods of use can be incorporated by reference to the description of the pharmaceutical composition for HHD of the present invention. When incorporated by reference, "HHD" and "symptoms of HHD" can be read as "acantholysis."

[0069] The present invention also relates to the use of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid in the manufacture of a pharmaceutical composition for acantholysis. As described above, the form of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid (MPBS) is not limited, and may be any of the MPBSs, i.e., the MPBS anhydrate, the MPBS salt, the MPBS hydrate, and / or the MPBS solvate. In the present invention, the MPBSs, their compositions, and methods of use can be referenced from the description of the pharmaceutical composition for HHD of the present invention. When referenced, "HHD" and "symptoms of HHD" can be replaced with "acantholysis."

[0070] In the following examples, 5-methyl-2-(1-piperazinyl)benzenesulfonic acid monohydrate (hereinafter, sometimes referred to as Compound A in the examples) was used as the MPBS.

[0071] Example 1: Suppression of differentiation abnormalities in an in vitro HHD three-dimensional cultured epidermal model Compound A was evaluated for its inhibitory effect on differentiation abnormalities in an HHD three-dimensional cultured epidermal model. It is known that abnormal differentiation of keratinocytes causes acantholysis in the skin lesions of HHD patients. In this model, the effect of Compound A on gene expression changes of differentiation markers was examined.

[0072] <Test Cells and Test Design> Because HHD is known to be associated with mutations in the ATP2C1 gene, an ATP2C1 siRNA-induced in vitro HHD 3D cultured epidermal model was prepared by the following method. The cells and culture medium used were from the LabCyte EPI-KIT (Japan Tissue Engineering Co., Ltd., 401810), a human epidermal model preparation kit. siRNA was transfected into human primary keratinocytes in the human epidermal model preparation kit. For transfection, a cationic lipid formulation, Lipofectamine® RNAiMAX Transfection Reagent (Thermo Fisher Scientific, 13778), was used as the transfection reagent, and Opti-MEM® Reduced Serum Medium (Thermo Fisher Scientific, 31985-062) was used as the medium. As an example, ATP2C1 siRNA (Thermo Fisher Scientific, J-006119-05-0005) was used as the siRNA, and non-targeting control siRNA (Thermo Fisher Scientific, D-001810-01-05) was used as a control. The final concentration of the siRNA was 100 nmol / L. In this specification, the "final concentration" of each test substance solution, reagent, etc. refers to the concentration in the medium at the final stage after all test substance solutions, reagents, etc. have been added.

[0073] The transfected cells into which the siRNA had been introduced were cultured in three dimensions using air-liquid interface culture inserts and 24-well plates (Day 0), and in vitro epidermal tissue was prepared by culturing for 8 days (Day 0 to Day 8). The culture environment conditions were 37°C, 5% CO ２The culture medium was maintained at 100 μmol / L. From the start of the three-dimensional culture (Day 0), an aqueous solution of Compound A (solvent: phosphate-buffered saline) was added to the culture medium of the in vitro epidermal tissue so that the final concentration of Compound A was 10 μmol / L. The culture medium was replaced daily, and the aqueous solution of Compound A was also added at each replacement. Eight days after the start of the three-dimensional culture (Day 8), the in vitro epidermal tissue was sampled and used for evaluation. Three groups were set: a control siRNA treatment group, an ATP2C1 siRNA treatment group, and an ATP2C1 siRNA + Compound A treatment group (an example group) treated with ATP2C1 siRNA and Compound A. The number of samples per group was nine epidermal tissues.

[0074] <Evaluation Method> RNA was extracted from the sampled in vitro epidermal tissue using an RNeasy® Mini QIAcube Kit (QIAGEN, 74116). The RNA concentration was measured using a microspectrophotometer (DropSense 96™ (Trinean) or LUNATIC (Unchained Labs)), and then reverse transcribed into cDNA. For reverse transcription, SuperScript™ IV VILO™ Master Mix (Thermo Fisher Scientific, 11756050) or High-capacity RNA to cDNA™ kit (Thermo Fisher Scientific, 4387406) was used. RT-qPCR analysis was performed using the cDNA as a template with the QuantStudio™ 6 Flex Real-Time PCR System (Applied Biosystems). For the RT-qPCR reaction, TaqMan® Fast Advanced Master Mix (Thermo Fisher Scientific, 4444557) or TaqMan® Fast Universal PCR Master Mix (Thermo Fisher Scientific, 4352042) was used as the master mix, and the TaqMan® probes (Thermo Fisher Scientific) listed in Table 1 were used. The differentiation markers used were the basal layer markers KRT5 and KRT14, and the granular layer marker IVL.

[0075]

[0076] In the RT-qPCR analysis, RPLP0 or HPRT1 was used as an internal standard gene. Then, using the ΔΔCt method, the relative gene expression level, based on the average value of the gene expression level in keratinocytes on Day 0, and the average value and standard error thereof were calculated. The inhibition rate was calculated as follows. Inhibition rate (%) = 100 × (A - B) / (A - C) A: Average relative gene expression level in the ATP2C1 siRNA treatment group B: Relative gene expression level in the ATP2C1 siRNA + Compound A treatment group C: Average relative gene expression level in the control siRNA treatment group

[0077] <Statistical Analysis> Statistical analysis was performed using SAS, with a two-sided significance level of 5%. The success of induction of the in vitro HHD three-dimensional cultured epidermal model was assessed by comparing the control siRNA-treated group with the ATP2C1 siRNA-treated group, and the pharmacological action of Compound A was assessed by comparing the ATP2C1 siRNA-treated group with the example group (ATP2C1 siRNA + Compound A-treated group). Student's t-test was used for all tests.

[0078] 1A and 1B are graphs showing the results of gene expression analysis, specifically, the relationship between administration of MPBS monohydrate and expression of differentiation marker molecules in an in vitro HHD 3D cultured epidermal model. The vertical axis of Fig. 1A represents relative gene expression levels based on the gene expression levels in keratinocytes on Day 0, and the vertical axis of Fig. 1B represents the rate of suppression of increased gene expression of differentiation markers by ATP2C1 siRNA treatment.

[0079] In the ATP2C1 siRNA treatment group, statistically significant increases in expression were observed for the basal layer markers KRT5 and KRT14, and the granular layer marker IVL, whose expression has been reported to be abnormal in HHD patients, compared to the control siRNA treatment group.In contrast, in the example group, the addition of Compound A suppressed the increase in expression of these differentiation markers, correcting the abnormal expression of these marker genes.

[0080] Example 2: Improvement of intercellular adhesion disorders in an in vitro epidermal intercellular adhesion disorder model Compound A was evaluated for its ability to improve intercellular adhesion in an epidermal intercellular adhesion disorder model. It is known that damage to intercellular adhesion can lead to acantholysis. The damage to intercellular adhesion was evaluated by measuring changes in electrical resistance (cell index) using a real-time cell analyzer.

[0081] <Test Cells and Test Design> An in vitro epidermal cell-cell adhesion disorder model was created using 2,2'-methylenebis(6-t-butyl-4-methylphenol) (hereinafter referred to as bisphenol) by the following method. Bisphenol has been reported to selectively inhibit the ATPase activity of SPCA1, the molecule responsible for HHD. Primary human keratinocytes (KURABO, KK-4109) derived from a Caucasian woman were used as cells. HuMedia-KG2 medium (KURABO, KK-2150S) was used to culture the primary human keratinocytes. The real-time cell analyzer used was an xCELLigence real-time cell analyzer (Agilent), and the cells were incubated at 37°C and 5% CO ２ The cells were placed in an incubator under controlled conditions.

[0082] First, 50 μl / well of HuMedia-KG2 medium was added to an E-Plate VIEW 96 PET (Agilent) plate. The plate was then placed in the real-time cell analyzer, and the background electrical resistance value was measured. The plate was then removed, and the human primary keratinocytes were seeded onto the plate at 100 μl / well to give a density of 25,000 cells / well, and the plate was then placed back into the real-time cell analyzer. Simultaneously with the placement, electrical resistance measurements were initiated and recorded at 15-minute intervals (Day 0).

[0083] On Day 1, the plate was removed. After 50 μl / well of culture supernatant was removed from the wells of the plate, 100 μl / well of calcium solution was added so that the final calcium concentration was 1.2 mmol / L. The plate was then placed back into the real-time cell analyzer, and measurement of electrical resistance was resumed. Next, on Day 2, the plate was removed. After 100 μl / well of culture supernatant was removed from the wells of the plate, 50 μl / well of Compound A was added. The plate was then placed back into the real-time cell analyzer, and measurement of electrical resistance was resumed. Then, 3 hours after the addition of Compound A, the plate was removed, and 50 μl / well of bisphenol solution was added to the plate. The plate was then placed back into the real-time cell analyzer, and measurement of electrical resistance was resumed.

[0084] Three groups were set: a bisphenol-free group, a bisphenol-added group (compound A not added), and an example group (bisphenol-added / compound A-added group). At the time of adding the bisphenol, the final concentration of compound A in the wells was 0 μmol / L (compound A not added), 0.01 μmol / L, 0.1 μmol / L, or 1 μmol / L, and the final concentration of bisphenol was 0 μmol / L (bisphenol not added) or 4 μmol / L.

[0085] The normalized electrical resistance value (normalized cell index; nCi) 60 hours after the addition of the bisphenol was used as an index of inhibition of intercellular adhesion disorders. nCi was normalized so that the electrical resistance value (cell index) immediately before the addition of the bisphenol was 1.

[0086] <Statistical Analysis> Statistical analysis was performed using SAS (Statistical Analysis System). The Student's t-test was used to compare the bisphenol-free group with the bisphenol-added group. The pharmacological action of Compound A was assessed by comparing the bisphenol-added group with the bisphenol-added / Compound A-added group, and the Williams multiple comparison test was used as the test method.

[0087] Compound A was evaluated in an in vitro model of epidermal cell-cell adhesion disorder induced by bisphenol. The test was performed four times, and the mean values ​​and standard errors of nCi at 60 hours after bisphenol addition were calculated and statistically analyzed. The results are shown in Figure 2.

[0088] 2 is a graph showing the relationship between administration of MPBS monohydrate and intercellular adhesion in an in vitro model of epidermal cell-cell adhesion disorder, with the vertical axis representing normalized electrical resistance. As shown in FIG. 2, the bisphenol-added group, in which bisphenol was added to cultured human primary keratinocytes under conditions of a final calcium concentration of 1.2 mmol / L, showed a statistically significant decrease in nCi compared to the bisphenol-free group ( ＃＃ p<0.01). Furthermore, it was found that, compared to the bisphenol-added group, the bisphenol-added / compound A-added group could recover the nCi that had decreased due to the addition of bisphenol by further pretreating with compound A. In particular, by pretreating with a compound A concentration of 0.1 to 1 μmol / L, the nCi that had decreased due to the addition of bisphenol was statistically significantly recovered ( ＊＊ p<0.005). These results suggest that Compound A has an effect of improving bisphenol-induced epidermal cell adhesion disorders.

[0089] Example 3 Transcriptome analysis in an in vitro epidermal cell-cell adhesion disorder model The transcriptome, i.e., the expression changes of gene transcription products, were comprehensively evaluated in the epidermal cell-cell adhesion disorder model for Compound A. The gene expression changes were evaluated based on the changes in RNA count values ​​or the changes in RNA expression levels normalized to the RNA count values ​​using a NextSeq500 / 550 sequencer.

[0090] <Test Design and Data Analysis Method> The epidermal cell-cell adhesion disorder model was prepared by culturing under the same conditions as in Example 2, except as described below.

[0091] The experimental groups (1) to (3) of this example are shown below. (1) Group 1 (Control Group): On Day 1, calcium was added to the wells, and the cells were cultured for 75 hours in the presence of a final calcium concentration of 1.2 mmol / L. This experimental group was not treated with bisphenol, simulating normal keratinocytes in the presence of calcium. (2) Group 2 (Bisphenol-Added Group): On Day 1, calcium was added to the wells, and the cells were cultured for 24 hours in the presence of a final calcium concentration of 1.2 mmol / L, followed by the addition of DMSO solution. After 3 hours of culture following the addition of DMSO, bisphenol (final concentration 4 μmol / L) was added and the cells were cultured for 48 hours. This experimental group was exposed to bisphenol, simulating the conditions that cause HHD. (3) Group 3 (Compound A-Added Group): On Day 1, calcium was added to the wells, and the cells were cultured for 24 hours in the presence of a final calcium concentration of 1.2 mmol / L, followed by the addition of Compound A (final concentration 1 μmol / L). After 3 hours of incubation from the addition of Compound A, bisphenol (final concentration: 4 μmol / L) was further added and the cells were incubated for 48 hours. This experimental group mimicked the conditions in which Compound A was administered to subjects with HHD.

[0092] After incubation, cells were collected from each experimental group (1) to (3), and total RNA was extracted using an RNeasy® Mini QIAcube column (Qiagen) according to the manufacturer's protocol. All extracted RNA samples were then treated with DNase. Using a NanoDrop (Thermo Fisher Scientific) and a Bioanalyzer RNA6000 Nanochip (Agilent), all extracted RNA samples were confirmed to be of high quality, with an RIN value of 9.4 or higher, indicating RNA quality, and a total RNA content of 1.3 μg or higher.

[0093] Using the extracted total RNA (10 ng) as a template, a cDNA library was prepared using NEBNext® Ultra II Directional RNA Library prep for Illumina (NEB), NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB), and NEBNext® Multiplex Oligos for Illumina (96 Unique Dual Index Primer Pairs, NEB) according to the product protocols.

[0094] Next, the sequence of the cDNA library was determined using a NextSeq500 / 550 system (Illumina) according to the product protocol. Quality control was performed using FASTQC (version 0.12.1) and Trimmomatic-0.39, and the data was converted into a FASTQ file.

[0095] Next, using StrandNGS software (version 4.0, Strand Life Science) and Star (version 2.7.10b), the gene types were identified from the 150-base sequence information contained in the FASTQ file, referring to the human genome assembly and transcript annotation (release 101) obtained from the Ensembl database. The expression levels of these transcripts were then calculated. The read count values ​​and normalized TPM values ​​were used to determine the expression levels of the transcripts.

[0096] Expression variation analysis was performed using DESeq2 (a two-group comparison package) in the statistical analysis software R (version 4.3.2), and the log2 ratio of RNA expression levels and significance tests were calculated. Significance tests were performed based on the p-value calculated by a Wald test with multiple testing correction based on the Benjamini-Hochberg method (BH method). In this analysis, genes with an absolute log2 ratio of 0.585 or greater (expression variation ratio of 1.5-fold or greater) or 0.263 or greater (expression variation ratio of 1.2-fold or greater), and a p-value of less than 0.1 by the Wald test between any two groups, were identified as "genes with altered expression levels" (expression variation genes; DEGs). DEGs include both upregulated and downregulated genes.

[0097] Gene ontology enrichment analysis was performed by comparing the ontology terms defined as Biological Processes in the Gene Ontology database with the genetic information associated with the ontology terms and the DEGs. Ontology terms with a p-value of less than 0.05 in the Fisher exact test with multiple testing correction based on the BH method were considered significant, and annotation of the DEGs was performed. This identified ontology terms of biological functions associated with the DEGs.

[0098] The expression levels of the transcripts in each experimental group were compared. As a result, in a comparison between the first and second groups, 4,827 genes were identified as DEGs (log2 ratio standard of 0.585 or more, 2,025 genes increased in the second group, which is the pathological group, and 2,802 genes decreased). In addition, in a comparison between the second and third groups, 1,515 genes were identified as DEGs (log2 ratio standard of 0.263 or more, 538 genes increased in the third group, which is the drug-treated group, and 977 genes decreased).

[0099] The results of the enrichment analysis are shown in Tables 2 and 3. Table 2 shows the biological function ontology terms that increased with bisphenol and decreased with compound A, and Table 3 shows the biological function ontology terms that decreased with bisphenol and increased with compound A. Of the biological function ontology terms associated with the obtained DEGs, six ontology terms were identified that were associated with DEGs that increased in the bisphenol-added group (group 2; pathological group) but decreased in the compound A-added group (group 3) (Table 2). Furthermore, 10 ontology terms were identified that were associated with DEGs that increased in the compound A-added group (group 3) but decreased in the bisphenol-added group (group 2; pathological group) (Table 3). These results suggest that bisphenol addition activates endoplasmic reticulum (ER) stress and alters Golgi apparatus function, and that compound A has an inhibitory effect on these alterations. Furthermore, it was suggested that the addition of bisphenols inhibited epidermal differentiation and weakened intercellular adhesion, and that Compound A normalized these changes. These findings suggest that Compound A has an ameliorative effect on bisphenol-induced epidermal intercellular adhesion disorders.

[0100]

[0101]

[0102] Although the present invention has been described above with reference to the embodiments and examples, the present invention is not limited to the above-described embodiments. Various modifications that can be understood by those skilled in the art can be made to the configuration and details of the present invention within the scope of the present invention.

[0103] This application claims priority based on Japanese Patent Application No. 2024-104617, filed June 28, 2024, the disclosure of which is incorporated herein in its entirety by reference.

[0104] According to the pharmaceutical composition of the present invention, by containing 5-methyl-2-(1-piperazinyl)benzenesulfonic acid, it is possible to treat or prevent Hailey-Hailey disease.

Claims

1. A pharmaceutical composition for treating Hailey-Hailey disease, comprising 5-methyl-2-(1-piperazinyl)benzenesulfonic acid.

2. The pharmaceutical composition according to claim 1, which inhibits acantholysis in Hailey-Hailey disease.

3. The pharmaceutical composition according to claim 1 or 2, wherein the dosage of 5-methyl-2-(1-piperazinyl)benzenesulfonic acid is 1 to 1000 mg / day.

4. A pharmaceutical composition for acantholysis comprising 5-methyl-2-(1-piperazinyl)benzenesulfonic acid.

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