KR20180099342A - Composition for preventing or treating inherited peripheral neuropathy comprising aminosalicylic acid and use thereof - Google Patents

Abstract

Provided are: a pharmaceutical composition for preventing or treating hereditary peripheral neuropathy; a composition for increasing cell survival rate of Schwann cells or inhibiting apoptosis; a method for preventing or treating the hereditary peripheral neuropathy of an individual; and a method for increasing cell survival rate of Schwann cells or inhibiting apoptosis in an individual.

Description

Composition for preventing or treating hereditary peripheral neuropathy comprising aminosalicylic acid,

A pharmaceutical composition for preventing or treating hereditary peripheral neuropathy, a composition for increasing the cell survival rate of Schwann cells or inhibiting apoptosis, a method for preventing or treating hereditary peripheral neuropathy in an individual, To increase cell viability or to inhibit cell death.

Hereditary peripheral neuropathy (IPN) is a congenital disorder that has family history and is characterized by dysfunctions of the motor and sensory nervous system due to gene mutations. It is divided into hereditary motor and sensory neuropathy (HMSN), hereditary motor neuropathy (HMN), and hereditary sensory neuropathy (HSN). Of these, HMSN accounts for the majority, which is known as Charcot-Marie-Tooth disease (CMT).

Although accurate genetic diagnosis of hereditary peripheral neuropathy has made it possible to prevent the onset and tailor the treatment to the genetic cause, the development of efficient hereditary peripheral neuropathy has not been developed yet. Currently, ascorbic acid is effective in animal models of peripheral neuropathy, but it has been found to be ineffective in clinical trials, and a fundamental therapeutic agent is required. Accordingly, the inventors of the present invention have completed the present invention by confirming the causative agent of hereditary peripheral neuropathy and the customized therapies suitable for the hereditary peripheral neuropathy.

One aspect provides a pharmaceutical composition for preventing or treating hereditary peripheral neuropathy.

Another aspect provides a composition for increasing the cell viability of Schwann cells or inhibiting apoptosis.

Another aspect provides a method of preventing or treating a hereditary peripheral neuropathy of an individual.

Another aspect provides a method of increasing the cell survival rate of Schwann cells or inhibiting apoptosis in an individual.

One aspect provides a pharmaceutical composition for preventing or treating hereditary peripheral neuropathy, comprising an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof.

The amino salicylic acid may be an amino derivative of salicylic acid having the structure of the following formula 1a, 1b, or 1c. The aminosalicylic acid may be 3-aminosalicylic acid (3-ASA), 4-aminosalicylic acid (4-ASA), 5-aminosalicylic acid (5-ASA) Or a combination thereof.

(식 1a)

(Formula 1a)

(식 1b)

(Formula 1b)

(식 1c)

(Equation 1c)

The aminosalicylic acid may be in the form of a pharmaceutically acceptable salt thereof. Such salts include those derived from inorganic acids such as the customary acid addition salts used in the pharmaceutical field, for example in the field of hereditary peripheral neurological diseases, such as hydrochloric acid, bromic acid, sulfuric acid, sulfamic acid, phosphoric acid or nitric acid, Or an organic acid such as acetic acid, succinic acid, glycolic acid, stearic acid, citric acid, malonic acid, methanesulfonic acid, tartaric acid, malic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, 2- acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, , ≪ / RTI > The salt may be a base addition salt such as ammonium, dimethylamine, monomethylamine, monoethylamine or diethylamine. In addition, the salt may be in the form of a conventional metal salt, for example, a salt derived from a metal such as sodium, potassium, lithium, magnesium, or calcium. The acid addition salt, the base addition salt or the metal salt may be produced by a conventional method. For example, a base addition salt or a metal salt may be obtained by reacting the ionic form of the amino salicylic acid, i.e., aminosalicylate, with an appropriate base or metal ion.

The pharmaceutically acceptable salt of the aminosalicylic acid may be one having the structure of Formula 2a, 2b or 2c.

(식 2a)

(Formula 2a)

(식 2b)

(Formula 2b)

(식 2c)

(Equation 2c)

In formula 2a, 2b or 2c, M represents sodium, potassium, lithium, magnesium, or calcium.

The pharmaceutically acceptable salt of the aminosalicylic acid may be sodium 4-aminosalicylic acid (s4-ASA) or potassium 4-aminosalicylic acid (p4-ASA).

The aminosalicylic acid may also be in the form of its solvate. "Solvate" means a complex or aggregate formed by one or more solute molecules, i.e., an amino salicylic acid or a pharmaceutically acceptable salt thereof, and one or more solvent molecules. The solvate may be, for example, a complex or aggregate formed with water, methanol, ethanol, isopropanol or acetic acid.

The amino salicylic acid may also be in the form of its stereoisomer. The stereoisomers include all stereoisomers such as enantiomers and diastereomers. The compound may be a stereoisomerically pure form or a mixture of one or more stereoisomers, for example, a racemic mixture. The separation of certain stereoisomers can be carried out by any of the conventional methods known in the art.

The aminosalicylic acid may be chemically synthesized or commercially available.

The composition may be a pharmaceutical composition. The composition may further comprise a pharmaceutically acceptable carrier. In such compositions, the term "pharmaceutically acceptable carrier" refers to a substance, generally an inert substance, used in combination with the active ingredient to aid in the application of the active ingredient. Such carriers include conventional pharmaceutically acceptable excipients, additives or diluents. The carrier may be, for example, a filler, a binder, a disintegrant, a buffer, a preservative, an antioxidant, a lubricant, a flavoring agent, a thickener, a coloring agent, A stabilizer, and an isotonic agent.

The composition may comprise a "therapeutically effective amount" of the aminosalicylic acid or a pharmaceutically acceptable salt, solvate, or combination thereof. As used herein, "therapeutically effective amount" means an amount sufficient to produce a therapeutic effect when administered to a subject in need of treatment. The term "treatment" means treating a disease or medical condition, such as a hereditary neurological disease, in a mammal, including a human, including an individual, including: (a) Prevention, i.e. prophylactic treatment of the patient; (b) relieving the disease or medical condition, i. e., eliminating or ameliorating the disease or medical condition in the patient; (c) inhibiting the disease or medical condition, i.e. slowing or stopping the progression of the disease or medical condition in the individual; Or (d) relieving the disease or medical condition in the subject. The "effective amount" may be 0.01 mg to 10,000 mg, 0.1 mg to 1000 mg, 1 mg to 100 mg, 0.01 mg to 1000 mg, 0.01 mg to 100 mg, 0.01 mg to 10 mg, or 0.01 mg to 1 mg. The subject may be a mammal, for example a human. The subject may have a hereditary peripheral neuropathy or Schwann cell and a disease associated with apoptosis. The composition may be one comprising as an active ingredient an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof or a combination thereof. "Active ingredient" is intended to carry out the function referred to in the composition and excludes those that do not fulfill the function as they are included in minor amounts as impurities.

The composition may be administered orally, or parenterally, including intravenously, intraperitoneally, subcutaneously, rectally, and topically. Thus, the composition may be formulated into various forms such as tablets, capsules, aqueous solutions or suspensions. In the case of tablets for oral use, excipients such as lactose and corn starch, and lubricants such as magnesium stearate may be usually added. In the case of capsules for oral administration, lactose and / or dried corn starch may be used as a diluent. If an oral aqueous suspension is required, the active ingredient may be combined with an emulsifying agent and / or a suspending agent. If desired, certain sweetening and / or flavoring agents may be added. For intranasal, intramuscular, intraperitoneal, subcutaneous and intravenous administration, sterile solutions of the active ingredient are usually prepared and the pH of the solution should be suitably adjusted and buffered. For intravenous administration, the total concentration of solutes should be adjusted to give the formulation isotonicity. The composition may be in the form of an aqueous solution comprising a pharmaceutically acceptable carrier, such as saline, at a pH of about 7.4. The solution can be introduced into a patient's intramuscular or nerve blood flow by local bolus injection.

The composition may be combined with one or more other therapeutic agents for treating hereditary peripheral neuropathy. In addition, the composition may be free of other active ingredients for treating hereditary peripheral neuropathy, and may include an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof.

The composition may be for treating hereditary peripheral neuropathy. Hereditary peripheral neuropathy (IPN) is a congenital disorder that has family history and is characterized by mutations in genes that cause dysfunctions of the motor and sensory nervous system. It is divided into hereditary motor and sensory neuropathy (HMSN), hereditary motor neuropathy (HMN), and hereditary sensory neuropathy (HSN). Hereditary peripheral neuropathy is, for example, a hereditary sensory and autonomic neuropathy (HSAN), distant hereditary motor neuropathy (dHMN), hereditary neuropathy with liability to pressure pancreatic cancer, palsies: HNPP), Charcot-Marie-Tooth disease (CMT), Dejerine-Sottas disease (Dejerine-Sottas syndrome: DSS) hypomyelination neuropathy: CH). The disease may be a disease, a disease, or a disease.

The hereditary peripheral neuropathy may be caused by reduced survival rate or increased death of Schwann cells. Schwann cells are basic glial cells of the peripheral nervous system. Schwann cells form a few seconds in the axon with a few seconds, and Schwann cells that do not form a few seconds function to maintain the axons. When Schwann cells are killed by Schwann cell death, they can not form a few seconds and can reduce neuronal signal transduction. In turn, this reduction in delivery may cause a decrease in sensation and / or motor function. The hereditary peripheral neuropathy may be Charcot-Marie-Tooth disease (CMT) or Dejerine-Sottas neuropathy (DSN). CMT is a disease characterized by the loss of sensory function and may be classified as CMT1, CMT2, CMT4, IntCMT or CMTX depending on the phenotype or genotype. CMT1 refers to a dehydrative neurological disorder with autosomal dominant inheritance and slow neurotransmission rate during CMT disease, and may be CMT1A, CMT1B, CMT1C, CMT1D, CMT1E, CMT1F or CMT1X. DSN means a disease that develops at a young age, exhibits symptoms of muscle atrophy, sensory changes, eye shaking and spinal curvature, and autosomal dominant or recessive inheritance.

The hereditary peripheral neuropathy may be associated with cell viability or cell death of schwann cells. The apoptosis may be due to apoptosis or necrosis. The Schwann cells may have a reduced survival rate or increased death, and the reduced survival rate or increased death may be caused by a mutation in the amino acid sequence of the myelin protein zero (MPZ) protein. MPZ is encoded by the MPZ gene, and the locus of the MPZ gene is chromosome 1q22. MPZ is a major constituent of about 50% or more of total plants, and is a major component of compact myelin. MPZ stabilizes a few seconds of assembly through the ability to cover and protect the nerves, the ability to promote nerve conduction, and the ability to control planting (encapsulation). MPZ has been reported to have more than 200 variants (http://hihg.med.miami.edu/code/http/cmt/public_html/index.html#/). Various mutations of MPZ can cause the hereditary peripheral neuropathy. Various variations of MPZ can cause CMT or DSN. The mutation of the amino acid sequence of the MPZ protein means that the amino acid sequence of MPZ is changed. The variation of the amino acid sequence of MPZ protein or the amino acid sequence of MPZ is also referred to as mutant MPZ. Changes in the amino acid sequence include missense mutation, silent mutation, nonsense mutation, neutral mutation, mutations in introns, and frame shift mutation , Frame shift: fs), and the like. A change in the amino acid sequence of MPZ may be a change of amino acid residues corresponding to V169, R98 positions or a combination thereof in a polypeptide consisting of the amino acid sequence of SEQ ID NO: 1. The modified amino acid sequence of MPZ may be V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: 1. The change in the amino acid sequence of MPZ may be due to a change in the nucleotide corresponding to the 292nd, 506th, or the combination thereof in the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2. The change in the amino acid sequence of MPZ may be due to a combination of c.292C> T, c.506delT, or a combination thereof in a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2. The Schwann cells may be present in vivo or in vitro.

Another aspect provides a composition for increasing the cell survival rate or inhibiting apoptosis of Schwann cells, comprising an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof.

The composition may include an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, as an "amount effective to increase the cell viability of a Schwann cell in a subject or to inhibit apoptosis " . The "effective amount" can be appropriately selected by a person skilled in the art. The Schwann cells may have a reduced survival rate or increased death, and the reduced survival rate or increased mortality may be due to mutations in the amino acid sequence of the MPZ protein. The mutation of the amino acid sequence of the MPZ protein may be V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: The mutation of the amino acid sequence of the MPZ protein may be by a combination of c.292C> T, c.506delT, or a combination thereof in a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2.

The amino acid sequence of the amino acid, the amino acid, the amino acid, the aminosalicylic acid, the pharmaceutically acceptable salt thereof, the solvate thereof or the combination thereof, the Schwann cell, the cell survival rate, the cell death, the mutation of the amino acid sequence of MPZ and MPZ protein.

Another aspect provides a composition for inhibiting the occurrence of ER stress in Schwann cells, or for reducing ER stress, comprising an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof.

Said composition comprising an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof as "an amount effective to inhibit the occurrence of ER stress in a subject or to reduce ER stress " . The "effective amount" can be appropriately selected by a person skilled in the art. Endoplasmic (ER) stress means that an abnormal protein is accumulated in the endoplasmic reticulum, for example, a protein having an incorrect fold. Such misfolded proteins can produce oxidative stress in the endoplasmic reticulum, for example, reactive oxygen species (ROS). Schwann cells may be caused or increased in ER stress, and the ER stress may be caused or increased by mutation of the amino acid sequence of the MPZ protein. Mutations in the amino acid sequence of the MPZ protein can form MPZ proteins with false folds. Accumulation of MPZ proteins with misfolding in the endoplasmic reticulum can produce oxidative stress. The mutation of the amino acid sequence of the MPZ protein may be V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: The mutation of the amino acid sequence of the MPZ protein may be by a combination of c.292C> T, c.506delT, or a combination thereof in a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2.

Aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, mutation of the amino acid sequence of Schwann cells, MPZ, MPZ protein, and the like are as described above.

Another aspect provides the use of an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof or a combination thereof as defined above in the treatment of hereditary peripheral neuropathy.

Another aspect provides the use of an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, as defined above, in the manufacture of a medicament for the treatment of hereditary peripheral nerve diseases.

Another aspect provides a method for preventing or treating hereditary peripheral neuropathy by administering to an individual an amount of an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof effective to prevent or treat hereditary peripheral neuropathy A method for preventing or treating a hereditary peripheral neuropathy of an individual.

"An amount effective to prevent or treat a hereditary peripheral neuropathy" means an amount sufficient to prevent or treat a hereditary peripheral neuropathy when administered to a subject in need of prevention or treatment of the hereditary peripheral neuropathy.

The hereditary peripheral neuropathy may be caused by reduced survival rate or increased death of Schwann cells. The Schwann cells may have a reduced survival rate or increased death, and the reduced survival rate or increased mortality may be due to mutations in the amino acid sequence of the MPZ protein. The hereditary peripheral neuropathy may be due to a mutation in the amino acid sequence of the MPZ protein. The mutation of the amino acid sequence of the MPZ protein may be V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: The mutation of the amino acid sequence of the MPZ protein may be by a combination of c.292C> T, c.506delT, or a combination thereof in a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2.

The hereditary peripheral neuropathy may be CMT or DSN. The CMT may be CMT1B. Mutations in the amino acid sequence of the MPZ protein can increase ER stress, induce apoptosis of Schwann cells, and induce CMT. Mutations in the amino acid sequence of the MPZ protein can increase ER stress mediated by unfolded protein response (UPR) and induce CMT1B by inducing apoptosis in Schwann cells.

In the above method, the ordinarily skilled artisan can appropriately select the route of administration upon administration according to the condition of the patient. The administration may be oral, parenteral, or local administration. The local administration may be administration to the nervous system, for example, the brain.

In this method, the dosage varies depending on various factors such as the condition of the patient, the route of administration, the judgment of the attending physician, etc., as described above. Effective doses can be estimated from dose-response curves obtained from in vitro experiments or animal model studies. The proportions and concentrations of the aminosalicylic acid, its pharmaceutically acceptable salts, solvates thereof, or combinations thereof present in the composition to be administered can be determined according to chemical characteristics, route of administration, therapeutic dose, and the like. The dosage can be administered to an individual in an effective amount of from about 1 μg / kg to about 1 g / kg per day, or from about 0.1 mg / kg to about 500 mg / kg per day. The dose may vary depending on the age, weight, susceptibility, or symptom of the individual.

The method may further comprise collecting information relating to hereditary peripheral nerve disease in the subject prior to administration. Information related to hereditary peripheral neuropathy may be a result of judging the clinical symptoms, electrophysiologic examination (nerve conduction test and electromyography test), and neurobiology. The information associated with the hereditary peripheral neuropathy may include highly arched foot, contracture, sensorineural hearing loss, vocal cord palsy, hoarseness, Myotonic dystrophy, Functional Disability Scale, and the like. The diagnosed result may be a result of a diagnosis of a subject suspected of having or possibly having a hereditary peripheral neuropathy. The method may further include determining whether the subject has a hereditary peripheral nerve disease based on the diagnosis result, in comparison with the control group. The control group may be a result of the diagnosis of a healthy normal person. The method may further comprise administering to the subject an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, for an individual determined to have a hereditary peripheral nerve disorder in the determining step, as compared to the control .

Aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, hereditary peripheral neuropathy, prevention and treatment, and the like are as described above.

Another aspect is to provide a method of increasing the cell survival rate of Schwann cells in a subject by administering to the subject an amount of an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof effective to inhibit Schwann cell apoptosis in the subject Or inhibiting apoptosis, comprising the step of administering to a subject an effective amount of a compound of the present invention.

"An amount effective to inhibit apoptosis of Schwann cells in an individual" increases the cell survival rate of Schwann cells or inhibits apoptosis when administered to a subject in need of inhibiting apoptosis Quot; means an amount sufficient to exhibit the effect. The method of increasing the cell survival rate of Schwann cells or inhibiting apoptosis in the individual can effectively increase the cell survival rate or apoptosis of Schwann cells of the individual. The Schwann cells may have a reduced survival rate or increased death, and the reduced survival rate or increased mortality may be due to mutations in the amino acid sequence of the MPZ protein. The mutation of the amino acid sequence of the MPZ protein may be V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: The mutation of the amino acid sequence of the MPZ protein may be by a combination of c.292C> T, c.506delT, or a combination thereof in a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2.

The method may further comprise collecting information relating to apoptosis of Schwann cells in the subject prior to the administration. Information related to Schwann cell death may be cell death rate, cell survival rate, apoptosis rate, shape of Schwann cells, and the like. Information related to the cell death of Schwann cells can be measured by visualizing Schwann cells by optical or fluorescence microscopy, or by using Schwann cell-specific markers. Thus, the method may further comprise measuring the reduced survival rate or increased death of Schwann cells. The Schwann cells may be of an individual suspected of having or suspected of having a disease associated with reduced survival rate or increased death of schwann cells. The method further includes determining whether cell death is progressing or progressing as compared to a control group based on the information collected by collecting information related to the reduced survival rate or increased death of the Schwann cells can do. The control group may be about the cell survival rate of normal Schwann cells derived from an individual showing no apoptosis of the Schwann cells or the average cell survival rate of schwann cells. In addition, the method may further comprise administering to the subject determined to be progressing or progressing in Schwann cell apoptosis relative to the control group in the determining step, an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, To the patient.

The method may further comprise collecting information relating to the mutation of the amino acid sequence of the MPZ protein in the subject prior to the administration. Information related to the mutation of the amino acid sequence of the MPZ protein may be the amino acid sequence of the MPZ, the structure of the MPZ protein, the folding of the MPZ protein, the localization of the MPZ protein, the nucleotide sequence of the MPZ gene, and the like. The information related to the mutation of the amino acid sequence of the MPZ protein may be obtained by sequencing the amino acid sequence of the MPZ of the individual, sequencing the nucleotide sequence of the MPZ, visualizing the sequence with a laser scanning confocal microscope, observing the position of the MPZ in the cell, It can be measured using an MPZ-specific marker. Thus, the method may further comprise detecting the mutation of the amino acid sequence of the MPZ protein. The MPZ may be derived from an individual suspected of having or having a disease associated with mutant MPZ. The method may further comprise determining whether the amino acid sequence of the MPZ is altered relative to the control, based on the information collected by detecting the mutation of the amino acid sequence of the MPZ protein. The control group may be a normal MPZ amino acid sequence derived from an individual having no mutation in the amino acid sequence of the MPZ protein, an MPZ amino acid sequence derived from an average individual, or an MPZ amino acid sequence derived from a reference genome. In addition, the method may further comprise administering to the subject an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, to an individual determined to have a mutation in the amino acid sequence of the MPZ protein relative to the control in the determining step .

The amino acid sequence, amino acid sequence, dosage, dose, etc. of the amino salicylic acid, its pharmaceutically acceptable salt, its solvate or combination thereof, Schwann cells, MPZ and MPZ protein are as described above.

Another aspect relates to a method of inhibiting Schwann cell apoptosis comprising contacting an Schisan cell of an individual with an amount of an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, And suppressing apoptosis of the Schwann cells in the individual, wherein the suppression of apoptosis is inhibited. The Schwann cells may be isolated from an individual, or present in vitro. The contacting may be carried out in a liquid medium, for example a culture medium of the Schwann cells, or a buffer.

Aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof, Schwann cells and the like are as described above.

According to one aspect of the present invention, there is provided a pharmaceutical composition for preventing or treating hereditary peripheral neuropathy, which has an effect of preventing or treating hereditary peripheral neuropathy.

According to another aspect of the present invention, there is provided a composition for inhibiting apoptosis of Schwann cells, which has an effect of increasing the cell survival rate of Schwann cells or inhibiting apoptosis.

The method for preventing or treating a hereditary peripheral neuropathy of an individual according to another aspect has an effect of effectively preventing or treating a hereditary peripheral neuropathy of an individual.

According to the method for inhibiting Schwann cell apoptosis according to another aspect, it has an effect of increasing the cell survival rate of Schwann cells or inhibiting apoptosis in the individual.

FIG. 1A shows an image obtained by culturing RT4 cells transformed with the wild-type MPZ or the mutant MPZ gene, and microscopically observing the degree of attachment of the cells to the plate. Figure 1b shows the results of counting the number of cells after culturing RT4 cells transfected with wild-type MPZ or mutant MPZ genes (n = 3; scale bar = 200 μm; *, p <0.05; ***, p < 0.001). 1C to 1F show the viability of wild-type MPZ or RT4 cells transformed with the mutant MPZ gene after FACS analysis (n = 3; ***, p <0.001) ). Figures 1g and 1h show the expression levels of caspase-3, truncated caspase-3 and MPZ by Western blotting after culturing RT4 cells transfected with wild-type MPZ or mutant MPZ genes (n = 3; *, p &lt;0.05; ***, p < 0.001).
FIGS. 2A to 2C show the results of culturing RT4 cells transfected with wild-type MPZ or mutant MPZ genes and then confirming expression levels of ER stress markers BiP and CHOP by Western blotting (n = 3; *, p < 0.05). FIG. 2d is an image obtained by culturing RT4 cells transfected with wild-type MPZ or mutant MPZ gene, and showing the expression level and position of MPZ using Myc and the ER region using a laser scanning confocal microscope using PDI n = 3; scale bar = 10 m). FIG. 2E shows the results of culturing RT4 cells transfected with wild-type MPZ or mutant MPZ gene, and then measuring the intracellular ROS level by using DCF-DA and measuring the oxidation of DCF by fluorescence (n = 3; *, p &lt;0.05; ***, p < 0.001).
FIG. 3A shows the results obtained by adding 4-amino salicylic acid, sodium 4-amino salicylic acid, and 5-amino salicylic acid to RT4 cells transformed with wild-type MPZ or mutant MPZ genes, Represents the observed image. FIG. 3b shows the results of counting the number of cells after culturing with addition of 4-amino salicylic acid, sodium 4-amino salicylic acid, and 5-amino salicylic acid to RT4 cells transfected with wild-type MPZ or mutant MPZ genes. * Represents statistical significance for the MPZ V169fs control group. # Indicates statistical significance for the MPZ R98C control group. 3c, 3d, 3g, and 3h show the results of the mutation type (n = 3; scale bar = 200 m *, p &lt;0.05; **, p & 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid were added to RT4 cells transfected with the MPZ gene, and the cell viability was confirmed using FACS analysis. Figures 3e, 3f, 3i and 3j show that RT4 cells transduced with the mutant MPZ gene were cultured after addition of 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid, respectively, (N = 3; *, p <0.01; ***, p <0.001), confirming the number of apoptotic cells or necrosis cells.
FIGS. 4A to 4F show that the RT4 cells transduced with the mutant MPZ gene were supplemented with 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid, respectively, and then Western blotting revealed expression levels of BiP, CHOP and MPZ (N = 3; *, p <0.05; **, p <0.01; ***, p <0.001). FIG. 4g shows the result of culturing RT4 cells transfected with GFP-tagged mutant MPZ gene by adding 100 μM of 4-aminosalicylic acid, sodium 4-aminosalicylic acid, and 5-aminosalicylic acid, (N = 3; scale bar = 20 μm) using the PDI and the expression and localization of MPZ. 4h, 4i and 4j, RT-4 cells transduced with the mutant MPZ gene were cultured by adding 1, 10, and 100 μM of 4-aminosalicylic acid, and the expression levels of total JNK and P-JNK were confirmed by Western blotting (N = 3; *, p < 0.05).
FIGS. 5A and 5B show that RT4 cells transduced with the mutant MPZ gene were cultured by adding 1, 10, and 100 μM of 4-aminosalicylic acid, and then DCF-DA assay was used and oxidation of DCF was measured by fluorescence. (N = 3). FIG. 5c shows the results of measuring the ROS levels in the cells by measuring the oxidation of DCF using DCF-DA assay and the fluorescence of DCF-DA assay after adding 600 μM ascorbic acid as a positive control to RT4 cells transduced with the mutant MPZ gene (N = 3; *, p < 0.05).

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

Example  1: Aminosalicylic acid is a hereditary peripheral neuropathy on  Effect

1. Experimental Method

(1) wild type by Schwann cell culture and transduction MPZ  And variants MPZ  Gene expression

RT4 cells (RT4-D6P2T, CRL-2768, ATCC; Manassas, Va.), A rat-derived schwann cell, were cultured in RPMI 1640 supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin were cultured in high glucose Dulbecco's modified eagle medium (Biowest, Nuaille, France) containing penicillin-streptomycin at 37 ° C and 5% CO ₂ .

RT4 cells were plated on a 6-well plate at a cell number of 2 x 10 ⁵ per well. MPZ and mutant MPZ, namely pCMV-Myc-MPZ WT and pCMV-MPZ WT, containing the MPZ gene according to the manufacturer's protocol using the Lipofectamine 3000 reagent (Invitrogen, Paisley, UK) Myc-MPZ V169fs, pCMV-Myc-MPZR98C and pEGFP (C1) -MPZ WT, pEGFP (C1) -MPZV169fs vectors. The transduced cells were cultured at 37 DEG C for 48 hours. Or transfected cells were cultured at 37 ° C for 24 hours and the cells were treated with 4-aminosalicylic acid (4-ASA), sodium 4-aminosalicylic acid (s4-ASA ) And 5-aminosalicylic acid (5-ASA) (all Sigma-Aldrich, St Louis, MO, USA) were added and cultured for 24 hours.

(2) Classification of fluorescent marker cells  (Fluorescence-activated cell sorting: FACS ) analysis

The degree of cell death was measured by fluorescence-activated cell sorting (FACS). RT4 cells were plated on a 6-well plate at a cell number of 2 x 10 ⁵ per well. The wild-type MPZ and mutant MPZ genes, pCMV-Myc-MPZ WT, pCMV-Myc-MPZ V169fs and pCMV-Myc-MPZR98C vectors were transiently transfected into RT4 cells for 24 hours. Subsequently, aminosalicylic acid was added and cultured for 24 hours, and then cells to which aminosalicylic acid was added were obtained. The control group was obtained by transfecting RT4 cells transiently with wild-type MPZ and mutant MPZ genes and culturing for 48 hours without addition of aminosalicylic acid.

The cells to which the aminosalicylic acid was added were incubated in the dark state and at room temperature with saturated concentrations of Annexin V fluorescein isothiocyanate (FITC) and propidium iodide (PI) (BD (10 mM HEPES, 140 mM NaCl, and 2.5 mM CaCl ₂ , pH 7.4) containing 10 mM sodium phosphate buffered saline (Biosciences Pharmingen, San Jose, Calif., USA) PI staining was performed. The cultured cells were washed with cold phosphate buffered saline (PBS), pelleted, and analyzed with a FACS VERSE analyzer (Becton Dickinson, Francklin Lakes, NJ, USA).

(3) DCFDA  analysis

Reactive oxygen species (ROS) were measured by H ₂ DCFDA analysis.

The wild-type MPZ and mutant MPZ genes, pCMV-Myc-MPZ WT, pCMV-Myc-MPZ V169fs and pCMV-Myc-MPZR98C vectors were transfected into RT4 cells and cultured for 48 hours. As a control, RT4 cells were cultured for 48 hours with addition of 5 [mu] M of Brefeldin A (Brefeldin A). H ₂ DCFDA analysis was performed according to the manufacturer's protocol (Invitrogen). RT4 cells were obtained and 10 [mu] M H ₂ DCFDA was added followed by incubation at 37 ° C for 40 minutes. After cooling on ice, the cells were pelleted and analyzed with a FACS VERSE analyzer. The fluorescence intensity of H ₂ DCFDA produced by the reaction between H ₂ DCFDA and intracellular ROS of more than 10,000 living cells in each sample was analyzed by excitation at 488 nm and emission at 525 nm. The experiment was repeated three or more times, and the most representative data was selected using the histogram data.

(4) Western Blot  (western blotting) analysis

Expression of MPZ protein and ER stress markers was confirmed by western blotting.

Cells were harvested and lysed with RIPA buffer (Thermo scientific, Rockford, IL, USA). The cell lysate was centrifuged at 4 ° C for 15 minutes, and the supernatant was quantified. 20 μg of the protein was separated and transferred onto a polyvinylidene fluoride (PVDF) membrane. Then, the PVDF membrane was stained with a 0.1% Ponceau S staining solution (Sigma-Aldrich) to confirm that the same amount of protein was analyzed. Then, the PVDF membrane was reacted with blocking buffer (Casein blocking buffer 10x, Sigma-Aldrich) Lt; / RTI &gt; The PVDF membrane was reacted with the primary antibody overnight at 4 ° C. The primary antibodies were labeled with anti-myc-tag antibody (1: 2000; Abcam, Cambridge, UK), binding immunoglobulin protein (BiP) Secreted caspase-3 (Asp175) antibody, phospho-AKT (ser473) antibody, phospho-SAPK / JNK (Thr183 / Tyr185) (Sigma-Aldrich) and anti-beta actin antibody (1: 1000; Cell Signaling Technology, Danvers, MA, USA) The cells were then reacted with anti-mouse or anti-rabbit IgG, secondary antibody to which HRP was coupled (Cell Signaling Technology) and the protein was detected using Western Blot luminol reagent (Santa Cruz Biotechnology, Dallas, TX, USA) Respectively. The absorbance of the protein band in which the immune response occurred was measured using Image J software (https://imagej.nih.gov).

(5) Laser scanning Confocal  Microscopy (laser scanning confocal microscopy) image analysis

For immunofluorescence staining, RT4 cells were plated on a cover slip in a 24-well culture dish at 2 x 10 ⁴ cells per well. Wild-type MPZ and mutant MPZ (pCMV-Myc-MPZ WT, pCMV-Myc-MPZ-V169fs, pCMV-Myc-MPZ-R98C and pEGFP-MPZ WT and pEGFP-MPZ V169fs) were transfected into the RT4 cells. The transduced cells were cultured for 24 hours. The cultured cells were supplemented with 100 [mu] M of 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid and cultured for 24 hours. The cultured cells were fixed in 4% paraformaldehyde for 20 minutes, washed three times for 5 minutes with phosphate buffered saline, washed with 5% normal goat containing 0.1% Triton X-100, Serum was reacted at room temperature for 1 hour to block non-specific binding. Fixed cells were reacted with primary antibody overnight at 4 ° C. Each of the primary antibodies used was an anti-myc-tag antibody (1: 500; Abcam) and an anti-protein disulfide isomerase (PDI) antibody (1: 100; Cell Signaling Technology). Each cell was washed three times for 5 minutes with PBS and the cells were stained with Alexa Fluor 488 goat anti-rabbit IgG (1: 1000) and Alexa Fluor 568 goat anti-mouse IgG (1: 1000) (Molecular Probes, Eugene, OR , USA). &Lt; / RTI &gt; Each cell was washed and mounted on a VECTASHIELD HardSet Antifade Mounting Medium (Vector Laboratories, Burlingame, Calif.) Containing 4 ', 6-diamidino-2-phenylindole (DAPI) , CA, USA). The stained fragment was observed and imaged using a laser scanning confocal microscope (Carl Zeiss CLSM700, Oberkochen, Germany).

(6) Statistical analysis

All experiments (western blot, FACS, immunochemistry, quantitation) were performed three or more times independently. Comparisons between groups were made with Student's t -test. p <0.05 was considered statistically significant.

2. Mutation type MPZ  Confirmation of increased cell death of Schwann cells by expression

It was confirmed whether apoptosis was increased in Schwann cells expressing mutant MPZ.

FIG. 1A shows an image obtained by culturing RT4 cells transformed with the wild-type MPZ or the mutant MPZ gene, and microscopically observing the degree of attachment of the cells to the plate. 1A, it was confirmed that RT4 cells transfected with the wild-type MPZ or the mutant MPZ gene did not adhere to the bottom of the dish.

FIG. 1B shows the results of counting the number of cells after culturing RT4 cells transfected with wild type MPZ or mutant MPZ gene. When living cells were counted using trypan blue staining, the number of living cells was significantly reduced when the mutant MPZ gene was expressed. When the mutation of V169fs was expressed, it caused 65% apoptosis compared to the wild type. When the mutation of R98C was expressed, it caused 32% apoptosis compared to the wild type.

FIGS. 1C to 1F show the results of culturing RT4 cells transfected with wild-type MPZ or mutant MPZ genes, and then confirming cell viability using FACS analysis. Referring to FIG. 1d, when the mutant MPZ gene was expressed, the number of living cells was remarkably decreased. When the mutation of V169fs was expressed, the number of living cells was reduced to 38% as compared with the wild type. Referring to FIG. 1E, it can be seen that such apoptosis is due to apoptotic cell clusters.

Figures 1g and 1h show the results of culturing RT4 cells transfected with wild-type MPZ or mutant MPZ gene, followed by Western blotting to confirm expression levels of caspase-3, truncated caspase-3 and MPZ. When the mutant MPZ gene was expressed, the level of truncated caspase 3 expression was increased compared to the wild type. When the mutation of V169fs was expressed, the level of truncated caspase 3 expression was increased about twice as compared with the wild type. Mutations of mutant MPZ, V169fs and R98C, are known to induce Schwann cell death via apoptosis pathway.

3. Mutation type MPZ  Expression of ER stress in schwann cells and Active oxygen species  reactive oxygen species ROS ) Is increasing  Check whether

We confirmed whether MPZ localization, ER stress and ROS were increased in Schwann cells expressing mutant MPZ.

When ER stress increases, unfolded protein response (UPR) is activated. Unfolded protein response is a collection of signaling pathways to resolve false protein folding and restore the protein folding environment, such as increasing the production of chaperone to perform protein folding. Proteins acting downstream of the unfolded protein response may be responsible for pro-apoptosis. Thus, if the unfolded protein reaction does not occur smoothly, the unfolded protein response induces apoptosis.

Specifically, as the amount of unfolded protein increases, ER membrane bound proteins may be activated at an early stage when the unfolded protein response is activated. The ER membrane protein is expressed by ER kinase-like ER kinase (PERK), inositol-requiring enzyme 1 alpha (IRE1 alpha) or activating transcription factor 6 (ATF6) . Thus, translation by PERK is reduced and the cell cycle is stopped.

Activated PERK, IRE1a and ATF6 are expressed in pre-cells containing C / EBP homologous protein (CHOP), c-Jun N-terminal kinase (JNK) It can trigger apoptotic cascades mediated by apoptotic proteins.

FIGS. 2A to 2C show the results of culturing RT4 cells transfected with wild-type MPZ or mutant MPZ gene, and then Western blotting revealed expression levels of ER stress markers BiP and CHOP. When the mutation of V169fs was expressed, the expression level of BiP increased about 1.3 times. When the mutations of V169fs and R98C were expressed, the expression level of CHOP was significantly increased compared to the wild type.

FIG. 2D is an image obtained by culturing RT4 cells transfected with wild-type MPZ or mutant MPZ gene, expressing and locating MPZ using Myc, and confirming the ER region with a laser scanning confocal microscope using PDI. When wild-type and R98C mutations were expressed, the mutant MPZ protein was expressed in the cytosol whereas when the mutant of V169fs was expressed, the mutant MPZ protein was expressed in the same co-localization as PDI . That is, when the mutation of V169fs was expressed, the mutant MPZ protein was located in a relatively larger amount in the ER region than in the wild-type and R98C mutant expressions. When the R98C mutation was expressed, the RT4 cell morphology was narrow and bipolar.

FIG. 2E shows the results of confirming intracellular ROS levels by using DCF-DA and measuring the oxidation of DCF by fluorescence after culturing RT4 cells transformed with wild-type MPZ or mutant MPZ gene. Locating the unfolded protein in the ER may increase the level of oxidative stress in the ER. When the mutations of V169fs and R98C were expressed, the ROS levels were increased by about 2-fold and about 1.2-fold, respectively.

ER stress levels and localization in Schwann cells may vary depending on the variation of the amino acid sequence of the MPZ protein. Mutations of V169fs and R98C can induce Schwann cell apoptosis through apoptosis pathway, and this apoptosis can be induced by UPR and ROS increase in the endoplasmic reticulum.

4. Mutation type MPZ  The expressing Schwann cells were incubated with aminosalicylic acid ( aminosalicylic  acid: ASA) to determine if cell death is reduced

After Schwann cells expressing mutant MPZ were reacted with aminosalicylic acid, it was confirmed that the apoptosis of Schwann cells was reduced.

Aminosalicylic acid used 4-amino salicylic acid, sodium 4-amino salicylic acid and 5-amino salicylic acid. Each of these concentrations was selected in the range of about 0.1 to 1 nM, about 0.5 to 500 μM, about 0.75 to 250 μM, about 1 to 250 μM, about 5 to 250 μM, about 7.5 to 250 μM, or about 10 to 100 μM. 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid, respectively, were prepared at concentrations of 1, 10 and 100 μM, respectively. The aminosalicylic acid was added to RT4 cells transformed with the mutant MPZ gene and then cultured.

FIG. 3A shows the results obtained by adding 4-amino salicylic acid, sodium 4-amino salicylic acid, and 5-amino salicylic acid to RT4 cells transformed with wild-type MPZ or mutant MPZ genes, Represents the observed image. 3A, RT4 cells transfected with wild-type MPZ or mutant MPZ gene were found to be more attached to the bottom of the dish than when they were not reacted with aminosalicylic acid after reacting with aminosalicylic acid.

FIG. 3b shows the results of counting the number of cells after culturing with addition of 4-amino salicylic acid, sodium 4-amino salicylic acid, and 5-amino salicylic acid to RT4 cells transfected with wild-type MPZ or mutant MPZ genes. As a result of counting living cells using trypan blue staining, the number of living cells was significantly decreased when the mutant MPZ gene was expressed, but it was significantly increased when cultured with the addition of aminosalicylic acid. When the mutation of V169fs was expressed and cultured with addition of 4-amino salicylic acid and 4-amino salicylic acid, respectively, the number of living cells increased depending on the concentration of 4-amino salicylic acid and sodium 4-amino salicylic acid.

3c, 3d, 3g and 3h were obtained by adding 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid to RT4 cells transformed with the mutant MPZ gene, As shown in Fig. When the mutant MPZ gene was expressed, the number of living cells was remarkably decreased, but it was significantly increased when cultured with the addition of aminosalicylic acid.

FIGS. 3e, 3f, 3i and 3j show the results obtained by adding 4-aminosalicylic acid to RT4 cells transduced with the mutant MPZ gene and culturing the cells using apoptosis cells and necrosis cells using FACS analysis The results are shown in Fig. When the mutant MPZ gene was expressed and cultured with 4-aminosalicylic acid, live cells were significantly increased, and apoptosis and necrosis cells were significantly decreased. The addition of both sodium 4-aminosalicylic acid and 5-aminosalicylic acid resulted in the same results, showing a significant increase in living cells and a significant decrease in apoptosis and necrosis cells.

Aminosalicylic acid can reduce apoptosis induced by mutant MPZ. Specifically, 4-aminosalicylic acid, sodium 4-aminosalicylic acid and 5-aminosalicylic acid can reduce apoptosis and necrosis of schwann cells and increase the cell viability of Schwann cells. Thus, it can be seen that aminosalicylic acid can be used as a substance for preventing or treating hereditary peripheral neuropathy.

5. Mutation type MPZ  Determine whether ER stress is reduced by reacting expressing schwann cells with amino salicylic acid

After Schwann cells expressing mutant MPZ were reacted with aminosalicylic acid, the ER stress of Schwann cells was decreased.

FIGS. 4A to 4F show that 4-aminosalicylic acid, sodium 4-aminosalicylic acid, and 5-aminosalicylic acid were added to RT4 cells transduced with the mutant MPZ gene and cultured for 24 hours. Western blotting revealed that BiP, CHOP and MPZ Expression level is confirmed. When the mutant MPZ gene was expressed and cultured with addition of 4-amino salicylic acid, sodium 4-amino salicylic acid and 5-amino salicylic acid, ER stress marker expression was decreased. In the case of expressing the mutation of V169fs and culturing with addition of 4-aminosalicylic acid, the expression levels of BiP and CHOP decreased in a concentration dependent manner on the concentration of 4-aminosalicylic acid.

FIG. 4g shows the expression and localization of MPZ using GFP after addition of 100 μM of 4-aminosalicylic acid, sodium 4-amino salicylic acid, and 5-amino salicylic acid to RT4 cells transduced with the mutant MPZ gene, And an image of the ER region confirmed by a laser scanning confocal microscope using PDI. When the mutant of V169fs was expressed and cultured with addition of 4-aminosalicylic acid, the cell phenotype of Schwann cells was similar to that of RT4 expressing wild type MPZ. MPZ expression was not limited to the cell body or the endoplasmic reticulum All over Schwann cells. Expression of V169fs mutation and incubation with addition of sodium 4-aminosalicylic acid and 5-aminosalicylic acid, respectively, also revealed MPZ expression throughout Schwann cells.

4h, 4i and 4j, RT-4 cells transduced with the mutant MPZ gene were cultured by adding 1, 10, and 100 μM of 4-aminosalicylic acid, and the expression levels of total JNK and P-JNK were confirmed by Western blotting . When the R98C mutation was expressed and cultured with addition of 4-aminosalicylic acid, the expression level of P-JNK was significantly decreased.

Aminosalicylic acid can reduce ER stress induced by mutant MPZ. Aminosalicylic acid can significantly reduce ER stress and UPR-mediated pro-apoptotic markers, CHOP and P-JNK.

6. Mutation type MPZ  Expressing Schwann cells reacted with amino salicylic acid ROS  Check if it is decreasing

After Schwann cells expressing mutant MPZ were reacted with aminosalicylic acid, it was confirmed that the ROS of Schwann cells was decreased.

FIGS. 5A and 5B show that RT4 cells transduced with the mutant MPZ gene were cultured by adding 1, 10, and 100 μM of 4-aminosalicylic acid, and then DCF-DA assay was used and oxidation of DCF was measured by fluorescence. It shows the result of checking the ROS level. FIG. 5c shows the results of measuring the ROS levels in the cells by measuring the oxidation of DCF using DCF-DA assay and the fluorescence of DCF-DA assay after adding 600 μM ascorbic acid as a positive control to RT4 cells transduced with the mutant MPZ gene Results are shown. When V169fs and R98C mutants were expressed and cultured with addition of 4-aminosalicylic acid, there was no significant change in intracellular ROS levels. On the other hand, the ROS level in the positive control group was significantly decreased.

4-aminosalicylic acid does not affect the increased ROS levels due to ER stress mediated by mutant MPZ expression.

<110> Samsung Life Public Welfare Foundation <120> Composition for prevention or treating inherited peripheral          neuropathy comprising aminosalicylic acid and use thereof <130> PN116883 <160> 2 <170> Kopatentin 2.0 <210> 1 <211> 248 <212> PRT <213> Homo sapiens <400> 1 Met Ala Pro Gly Ala Pro Ser Ser Ser Pro Ser Ile Leu Ala Val   1 5 10 15 Leu Leu Phe Ser Ser Leu Val Leu Ser Pro Ala Gln Ala Ile Val Val              20 25 30 Tyr Thr Asp Arg Glu Val His Gly Ala Val Gly Ser Arg Val Thr Leu          35 40 45 His Cys Ser Phe Trp Ser Ser Glu Trp Val Ser Asp Asp Ile Ser Phe      50 55 60 Thr Trp Arg Tyr Gln Pro Glu Gly Gly Arg Asp Ala Ile Ser Ile Phe  65 70 75 80 His Tyr Ala Lys Gly Gln Pro Tyr Ile Asp Glu Val Gly Thr Phe Lys                  85 90 95 Glu Arg Ile Gln Trp Val Gly Asp Pro Arg Trp Lys Asp Gly Ser Ile             100 105 110 Val Ile His Asn Leu Asp Tyr Ser Asp Asn Gly Thr Phe Thr Cys Asp         115 120 125 Val Lys Asn Pro Pro Asp Ile Val Gly Lys Thr Ser Gln Val Thr Leu     130 135 140 Tyr Val Phe Glu Lys Val Pro Thr Arg Tyr Gly Val Val Leu Gly Ala 145 150 155 160 Val Ile Gly Gly Val Leu Gly Val Val Leu Leu Leu                 165 170 175 Tyr Val Val Arg Tyr Cys Trp Leu Arg Arg Gln Ala Ala Leu Gln Arg             180 185 190 Arg Leu Ser Ala Met Glu Lys Gly Lys Leu His Lys Pro Gly Lys Asp         195 200 205 Ala Ser Lys Arg Gly Arg Gln Thr Pro Val Leu Tyr Ala Met Leu Asp     210 215 220 His Ser Arg Ser Thr Lys Ala Val Ser Glu Lys Lys Ala Lys Gly Leu 225 230 235 240 Gly Glu Ser Arg Lys Asp Lys Lys                 245 <210> 2 <211> 747 <212> DNA <213> Homo sapiens <400> 2 atggctcctg gggctccctc atccagcccc agccctatcc tggctgtgct gctcttctct 60 tctttggtgc tgtccccggc ccaggccatc gtggtttaca ccgacaggga ggtccatggt 120 gctgtgggct cccgggtgac cctgcactgc tccttctggt ccagtgagtg ggtctcagat 180 gacatctcct tcacctggcg ctaccagccc gaagggggca gagatgccat ttcgatcttc 240 cactatgcca agggacaacc ctacattgac gaggtgggga ccttcaaaga gcgcatccag 300 tgggtagggg accctcgctg gaaggatggc tccattgtca tacacaacct agactacagt 360 gacaatggca cgttcacttg tgacgtcaaa aaccctccag acatagtggg caagacctct 420 caggtcacgc tgtatgtctt tgaaaaagtg ccaactaggt acggggtcgt tctgggagct 480 gtgatcgggg gtgtcctcgg ggtggtgctg ttgctgctgc tgcttttcta cgtggttcgg 540 tactgctggc tacgcaggca ggcggccctg cagaggaggc tcagtgctat ggagaagggg 600 aaattgcaca agccaggaaa ggacgcgtcg aagcgcgggc ggcagacgcc agtgctgtat 660 gcaatgctgg accaccacag aagcaccaaa gctgtcagtg agaagaaggc caaggggctg 720 ggggagtctc gcaaggataa gaaatag 747

Claims (20)

A pharmaceutical composition for preventing or treating a hereditary peripheral neuropathy, comprising an amino salicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof. The composition according to claim 1, wherein the disease is a disease caused by decreased survival rate or increased death of schwann cells. 3. The composition of claim 2, wherein the Schwann cell has a reduced survival rate or increased death, and wherein the reduced survival rate or increased death is due to a mutation in the amino acid sequence of the MPZ protein. 4. The composition according to claim 3, wherein the mutation of the amino acid sequence of the MPZ protein is V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: 1. The composition of claim 1, wherein the disease is Charcot Marie Tooth disease or Dejerine-Sottas disease. 6. The composition of claim 5, wherein the CMT is CMT1B. The pharmaceutical composition according to claim 1, wherein the aminosalicylic acid is 3-aminosalicylic acid, 4-aminosalicylic acid, 5-aminosalicylic acid, or a combination thereof, and the pharmaceutically acceptable salt of the aminosalicylic acid is sodium 4-aminosalicylic acid, -Aminosalicylic acid, or a combination thereof. A composition for increasing the cell viability or inhibiting apoptosis of Schwann cells, comprising an amino salicylic acid, a pharmaceutically acceptable salt thereof or a solvate thereof. 9. The composition of claim 8, wherein the Schwann cell has a reduced survival rate or increased death, and wherein the reduced survival rate or increased death is due to a mutation in the amino acid sequence of the MPZ protein. The composition according to claim 9, wherein the mutation of the amino acid sequence of the MPZ protein is V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: 1. Administering to an individual an amount of an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof effective to prevent or treat a hereditary peripheral nerve disorder, thereby preventing or treating hereditary peripheral neuropathy A method for preventing or treating a hereditary peripheral neuropathy of an individual. 12. The method of claim 11, wherein the disease is a disease caused by reduced survival or increased death of Schwann cells. 13. The method of claim 12, wherein the Schwann cell has a reduced survival rate or increased death, and the reduced survival rate or increased death is due to a mutation in the amino acid sequence of the MPZ protein. 14. The method according to claim 13, wherein the mutation of the amino acid sequence of the MPZ protein is V169fs, R98C, or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: 1. 12. The method according to claim 11, wherein the disease is Charcot Marie Tooth disease (CMT) or Dejerine-Sottas disease. 16. The method of claim 15, wherein the CMT is CMT1B. 12. The method of claim 11, wherein the aminosalicylic acid is 3-aminosalicylic acid, 4-aminosalicylic acid, 5-aminosalicylic acid, or a combination thereof, wherein the pharmaceutically acceptable salt of the aminosalicylic acid is sodium 4-aminosalicylic acid, -Aminosalicylic acid, or a combination thereof. An effective amount of an aminosalicylic acid, a pharmaceutically acceptable salt thereof, a solvate thereof, or a combination thereof in an amount effective to inhibit apoptosis of a Schwann cell in an individual is administered to an individual to increase the cell survival rate of the Schwann cell in the individual, And inhibiting apoptosis of the Schwann cells in the subject. 19. The method of claim 18, wherein the Schwann cells have reduced survival rate or increased death, and the reduced survival rate or increased mortality is due to mutations in the amino acid sequence of the MPZ protein. The method according to claim 19, wherein the mutation of the amino acid sequence of the MPZ protein is V169fs, R98C or a combination thereof in the polypeptide consisting of the amino acid sequence of SEQ ID NO: 1.

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