Agent for ameliorating liver lesion caused by lesion in anterior pituitary gland and / or for preventing and / or treating diseases accompanied by said liver lesion, and screening method for agent for ameliorating liver lesion caused by lesion in anterior pituitary gland and / or for preventing and / or treating diseases accompanied by said liver lesion

Abstract

Provided are: an agent for ameliorating a liver lesion caused by a lesion in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by said liver lesion; and a screening method for an agent for ameliorating a liver lesion caused by a lesion in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by said liver lesion. This agent for ameliorating a liver lesion caused by a lesion in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by said liver lesion comprises one protein or polypeptide among the following (a)-(c): (a) a protein or polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 1 of the sequence listing; (b) a protein or polypeptide consisting of an amino acid sequence in which one or more amino acids are deleted, substituted, and / or added in the amino acid sequence set forth in SEQ ID NO: 1 of the sequence listing, and having an insulin-like action and a growth-promoting action; or (c) a protein or polypeptide consisting of an amino acid sequence that shares a sequence identity of at least 95% with the amino acid sequence set forth in SEQ ID NO: 1 of the sequence listing, and having an insulin-like action and a growth-promoting action.

Description

A method for screening an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases associated with the liver lesions, and an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases associated with the liver lesions

[0001] The present invention relates to an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, and a method for screening for an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions.

[0002] The pituitary gland controls the functions of various hormones, and the anterior pituitary gland in particular secretes growth hormone (GH), thyroid-stimulating hormone, adrenocorticotropic hormone, etc. Growth hormone (GH) promotes metabolism and bone and muscle growth in organs such as the liver and muscles, and is the most abundant of the pituitary hormones. Meanwhile, it is known that GH secretion disorders occur most quickly due to lesions (disease, dysfunction, hypofunction, hypoplasia, abnormalities, etc.) of the pituitary gland (particularly the anterior pituitary gland). Such impaired GH secretion can result in liver lesions (hepatosplenomegaly, hepatomegaly, liver dysfunction, etc.), which may eventually lead to diabetes, etc. However, no effective ameliorators for liver lesions caused by lesions in the anterior pituitary gland and / or effective preventive and / or therapeutic agents for diseases accompanied by such liver lesions have yet been discovered.

[0003] Niemann-Pick disease type C (hereinafter simply referred to as "NPC") is an autosomal recessive genetic disease (mutation of the NPC1 or NPC2 gene) that primarily develops in early childhood and presents with severe liver lesions (hepatosplenomegaly, hepatomegaly, liver dysfunction, etc.). It is designated as a lysosomal storage disease and a specific disease. Furthermore, no safe and effective preventive and / or therapeutic agents or treatments have been discovered, and many patients die in their teens. For example, the glucosylceramide synthase inhibitor miglustat is currently available as a therapeutic agent for NPC, but its therapeutic effect is ineffective against liver damage (Non-Patent Document 1). Therefore, there is an urgent need to develop novel preventive and / or therapeutic agents for NPC that are effective against liver damage.

[0004] Folia Pharmacol. Jpn. 141, 160-167 (2013)

[0005] The present invention has been made in consideration of the above-mentioned problems of the conventional art, and aims to provide an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, as well as a method for screening for an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions.

[0006] As a result of extensive research into the above-mentioned problems, the present inventors have found that lesions in the anterior pituitary gland cause impairment of the IGF-1 production and / or secretion system via the GH and insulin-like growth factor 1 (IGF-1) system, resulting in the formation of liver lesions. Furthermore, the present inventors have experimentally found that a significant decrease in serum IGF-1 concentration occurs as a result of lesions in the anterior pituitary gland. Furthermore, the present inventors have found that Npc1 mice, a model mouse for diseases accompanied by liver lesions due to lesions in the anterior pituitary gland, show a significant decrease in serum IGF-1 concentration. －／－ By subcutaneously administering mecasermin, a genetically engineered form of IGF-1 that is undersecreted, to mice (Npc1 gene-deficient mice), Npc1 －／－ It was demonstrated that the compound significantly inhibited liver lesions observed in mice. The present invention was completed based on the above findings. Specifically, the present invention is as follows.

[0007] <1> An agent for ameliorating liver lesions caused by lesions of the anterior pituitary gland and / or preventing and / or treating diseases accompanied by the liver lesions, comprising a protein or polypeptide selected from the following (a) to (c): (a) a protein or polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing; (b) a protein or polypeptide consisting of an amino acid sequence in which one or more amino acids are deleted, substituted, and / or added in the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, and having insulin-like and growth-promoting effects; or (c) a protein or polypeptide consisting of an amino acid sequence having 95% or more sequence identity with the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, and having insulin-like and growth-promoting effects. <2> The agent according to <1>, wherein the disease accompanied by liver lesions is at least one disease selected from the group consisting of diseases accompanied by central nervous system disorders, pediatric diseases, and hereditary diseases. <3> The agent according to <1>, wherein the disease accompanied by liver lesions is Niemann-Pick disease type C. <4> The agent according to <1>, intended for administration to a Niemann-Pick disease C patient having liver lesions caused by Niemann-Pick disease type C. <5> The agent according to any one of <1> to <4>, which is at least one selected from the group consisting of subcutaneous administration and continuous administration. <6> A method for screening for an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, the method comprising a step of screening using an improvement in IGF-1 production or activity by the liver as an index. <7> The method according to <6>, wherein the disease accompanied by liver lesions is at least one disease selected from the group consisting of diseases accompanied by central nervous system disorders, pediatric diseases, and genetic diseases. <8> The method according to <6>, wherein the disease accompanied by liver lesions is Niemann-Pick disease type C. <9> A method for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, comprising administering to a subject any of the proteins or polypeptides (a) to (c) above.

[0008] According to the present invention, it is possible to provide an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or a preventive and / or therapeutic agent for diseases accompanied by the liver lesions, and a method for screening an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or a preventive and / or therapeutic agent for diseases accompanied by the liver lesions. According to the present invention, it is possible to significantly improve liver lesions caused by lesions in the anterior pituitary gland and liver lesions in diseases accompanied by the liver lesions (preferably NPC).

[0009] 1 is a graph showing a significant decrease in pituitary weight and a significant decrease in the production and / or secretion of IGF-1 in pathological model mice of diseases (NPC, etc.) accompanied by liver lesions due to lesions in the anterior pituitary gland. 2 is a graph showing the results of a test in which mecasermin was administered subcutaneously daily to pathological model mice of diseases (NPC, etc.) accompanied by liver lesions due to lesions in the anterior pituitary gland (serum ALT and AST values). 3 is a graph showing the results of verification of the therapeutic effect of miglustat in pathological model mice. 4 is a graph showing a significant negative correlation between serum IGF-1 concentration and the liver damage marker ALT value in pathological model mice.

[0010] Hereinafter, embodiments of the present invention will be described in detail, but the present invention is not limited to the following embodiments and can be implemented with appropriate modifications within the scope of the object of the present invention.

[0011] <<Agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions>> A first aspect of the present invention is an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, comprising any one of the following proteins or polypeptides (a) to (c): (a) a protein or polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, (b) a protein or polypeptide consisting of an amino acid sequence in which one or more amino acids are deleted, substituted, and / or added in the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, and which has insulin-like action and growth-promoting action, or (c) a protein or polypeptide consisting of an amino acid sequence having 95% or more sequence identity with the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, and which has insulin-like action and growth-promoting action. The present invention also relates to the use of any one of the proteins or polypeptides (a) to (c) above for the manufacture of an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions.

[0012] In the present invention, anterior pituitary lesions include anterior pituitary dysfunction, hypofunction, hypoplasia, disease, abnormality, etc. Liver lesions include hepatosplenomegaly, hepatomegaly, liver dysfunction, etc.

[0013] In the present invention, "insulin-mimetic action" refers to a blood glucose lowering effect (preferably a blood glucose lowering effect mediated by the IGF-1 receptor). For example, an increase in glucose uptake in muscle-derived cells of any animal (e.g., mouse, rat, guinea pig, rabbit, dog, cat, cow, pig, monkey, or human) can be used as an indicator of "insulin-mimetic action." Furthermore, a decrease in blood glucose levels following administration of the agent or a test substance described below to any animal (e.g., mouse, rat, guinea pig, rabbit, dog, cat, cow, pig, or monkey) can be used as an indicator of "insulin-mimetic action" (e.g., Orphan Pacific, IF. 2015). More specific examples of glucose uptake activity include measurement of intracellular glucose concentration, measurement of the amount of intracellular uptake of a glucose-analogous tracer substance, and measurement of changes in glucose transporters. Methods for measuring glucose concentration include absorbance measurement methods such as enzymatic methods, methods for measuring the amount of intracellular uptake of glucose-related tracer substances include measuring the amount of [3H]-2'-deoxyglucose uptake, and methods for observing changes in glucose transporters include cell immunostaining and Western blotting, but those skilled in the art can also use other methods as appropriate. In addition, any primary cultured cells, established cell lines, or transformants of these cells may also be used.

[0014] Furthermore, the term "growth-promoting activity" refers to the cell proliferation-inducing activity in vitro, and to the activity of increasing muscle mass and / or body length in vivo. For example, the cell proliferation-inducing activity (e.g., EC growth) mediated by IGF-1 receptor in myoblasts of any animal (e.g., mouse, rat, guinea pig, rabbit, dog, cat, cow, pig, monkey, human) is not particularly limited. ５０The above-mentioned "cell proliferation-inducing activity" can be measured by measuring the cell number, the amount of DNA synthesis, the amount of change in metabolic enzyme activity, etc. More specifically, examples include cell number measurement, measurement of the amount of DNA synthesis, and measurement of the amount of change in metabolic enzyme activity. Methods for measuring cell number include a method using a hemocytometer or a method using a cell counting device such as a Coulter counter. Methods for measuring the amount of DNA synthesis include a measurement method based on the incorporation of [3H]-thymidine or 5-bromo-2'-deoxyuridine (BrdU), and methods for observing the amount of change in metabolic enzyme activity include the MTT method, the XTT method, and the WST method, but those skilled in the art can also use other methods as appropriate. Furthermore, any primary cultured cells, established cell lines, or transformants of these cells may also be used. In addition, in the above-mentioned in vivo measurement of the effect of increasing muscle mass and / or body length, measurements of the whole individual such as body weight, body length, limb circumference, etc., body composition measurement by impedance method, creatinine, height coefficient, etc. are used, and methods such as image analysis by CT or MRI, contrast methods using tracers such as isotopes or fluorescent substances are also used, and changes in muscle strength, etc. can also be used as an index. In addition, in animals other than humans, methods such as directly collecting muscles and measuring their weight and size are also possible.

[0015] The ameliorating, preventive and / or therapeutic agent according to the first aspect contains the above protein or polypeptide and is therefore capable of significantly ameliorating liver lesions caused by lesions in the anterior pituitary gland and liver lesions in diseases accompanied by the above liver lesions (preferably NPC) (preferably by a paracrine or autocrine action similar to that of IGF-1).

[0016] In the first aspect, the disease accompanied by the liver lesion is not particularly limited as long as the present invention can be applied, but from the viewpoint of targeting serious diseases, pediatric diseases (for example, diseases that occur in infancy) are preferred. Here, pediatric diseases refer to diseases that are not adult diseases, and include diseases that occur in infancy. Here, infancy in humans refers to 12 years or younger, preferably 11 years or younger, more preferably 10 years or younger, more preferably 9 years or younger, even more preferably 8 years or younger, particularly preferably 7 years or younger, and most preferably 6 years or younger. Furthermore, infancy refers to after the day of turning 1 year old, after turning 2 years old, after turning 3 years old, etc. Here, adult diseases refer to lifestyle-related diseases whose onset is deeply related to long-term lifestyle habits, and particularly include diseases that occur after middle age.

[0017] The disease accompanied by liver lesions is preferably a genetic disease, more preferably an autosomal recessive genetic disease, from the viewpoint of targeting serious diseases. The disease accompanied by liver lesions is preferably a disease accompanied by central nervous system disorders, from the viewpoint of targeting serious diseases. Preferred examples of the disease accompanied by liver lesions include Niemann-Pick disease type C, Niemann-Pick disease type A, and Niemann-Pick disease type B, with Niemann-Pick disease type C being preferred. The ameliorating, preventive, and / or therapeutic agent according to the first aspect is preferably an ameliorating, preventive, and / or therapeutic agent for administration to a Niemann-Pick disease type C patient having liver lesions caused by Niemann-Pick disease type C.

[0018] SEQ ID NO: 1 represents the amino acid sequence of IGF-1 (mecasermin). As used herein, the term "one to several" in the phrase "an amino acid sequence in which one or several amino acids have been deleted, substituted, and / or added" is not particularly limited, but preferably refers to approximately 1 to 10, more preferably 1 to 5, and even more preferably 1 to 3. As used herein, the term "an amino acid sequence having 95% or more sequence identity" refers to an amino acid sequence with 95% or more sequence identity, preferably 96% or more, more preferably 97% or more sequence identity. The side chains of amino acids that constitute proteins differ from one another in terms of hydrophobicity, charge, size, etc., but several highly conserved relationships that do not substantially affect the three-dimensional structure (also referred to as tertiary structure) of the entire protein are known empirically and through physicochemical measurements. For example, substitutions of amino acid residues include glycine (Gly) and proline (Pro), Gly and alanine (Ala) or valine (Val), leucine (Leu) and isoleucine (Ile), glutamic acid (Glu) and glutamine (Gln), aspartic acid (Asp) and asparagine (Asn), cysteine ​​(Cys) and threonine (Thr), Thr and serine (Ser) or Ala, and lysine (Lys) and arginine (Arg).

[0019] Therefore, even if a mutant protein or polypeptide is produced by substitution, insertion, deletion, or the like in the amino acid sequence of IGF-1 (mecasermin) set forth in SEQ ID NO: 1 in the Sequence Listing, as long as the mutation is highly conserved in the three-dimensional structure of IGF-1 (mecasermin) and the mutant protein is a physiologically active protein that, like IGF-1 (mecasermin), has specific insulin-like and growth-promoting effects, all of these fall within the scope of IGF-1 (mecasermin). There are no particular limitations on the method for obtaining the IGF-1 (mecasermin) protein or polypeptide, and the protein or polypeptide may be a protein or polypeptide synthesized by chemical synthesis, a naturally occurring protein or polypeptide isolated from a biological sample or cultured cells, or a recombinant protein or polypeptide produced by genetic engineering. Furthermore, IGF-1 (mecasermin) set forth in SEQ ID NO: 1 in the Sequence Listing may have three disulfide bonds within the molecule, as shown in the structure below.

[0020] The administration route of the improving, preventive, and / or therapeutic agent according to the first aspect is not particularly limited as long as it achieves the effects of the present invention, and may be systemic or local administration. Continuous administration or bolus administration may also be used, with continuous administration being preferred. The continuous administration form is not particularly limited as long as it achieves the effects of the present invention, and examples include an implantable pump (e.g., implanted in subcutaneous tissue) or an extracorporeal pump for administration in a dosage form such as an injection, as described below. An implantable pump is preferred from the viewpoints of reducing the risk of infection and not being limited by patient movement. The administration may be transdermal (subcutaneous, intramuscular, intravenous, or other injection), intracerebroventricular (injection), oral, or intraperitoneal (injection), but subcutaneous administration is preferred from the viewpoints of low invasiveness, not being limited by patient movement, etc. In the first aspect, at least one selected from the group consisting of subcutaneous administration and continuous administration is more preferably used, and continuous subcutaneous administration is particularly preferred from the viewpoints of low invasiveness, not being limited by patient movement, etc.

[0021] The dosage of the improving, preventing and / or treating agent according to the first aspect varies depending on the subject of administration, the administration route, the target disease, symptoms, etc., but a person skilled in the art can appropriately select an appropriate dosage. For example, when administered (preferably subcutaneously) to a mammal (e.g., human, mouse, rat, rabbit, dog, cat, cow, pig, monkey, etc.), preferably to a human (e.g., an adult human weighing 50 kg), the single dose of the protein or polypeptide as an active ingredient (e.g., a component having pharmacological activity in the body of an animal including a human, or a component that, when contacted with other substances such as microbial contaminants, causes a physical or chemical change in the other substances or in the active ingredient itself; furthermore, physical or chemical changes referred to here include binding, transfer, rearrangement, addition, elimination, degradation, cleavage, oxidation, reduction, labeling, color development, luminescence, etc.) is usually about 0.001 mg / kg or more, preferably 0.05 mg / kg or more, more preferably 0.1 mg / kg or more, even more preferably 0.3 mg / kg or more, particularly preferably 0.7 mg / kg or more, and most preferably 1.0 mg / kg or more). The upper limit of the single dose is usually about 10.0 mg / kg or less, preferably 5.0 mg / kg or less, more preferably 3.0 mg / kg or less, even more preferably 2.5 mg / kg or less, particularly preferably 2.0 mg / kg or less, particularly preferably 1.8 mg / kg or less, and most preferably 1.5 mg / kg or less. It is more preferable to administer the above dose once to three times a day (preferably once or twice a day).

[0022] The dosage of the improving, preventive, and / or therapeutic agent according to the first aspect may be determined by mixing the protein or polypeptide directly or with a pharmaceutically acceptable carrier to form a pharmaceutical composition. The pharmacologically acceptable carrier may be any of a variety of organic or inorganic carrier substances commonly used as formulation materials, and may be incorporated as excipients, lubricants, binders, or disintegrants in solid formulations, or as solvents, solubilizers, suspending agents, isotonicity agents, buffers, or soothing agents in liquid formulations. Furthermore, formulation additives such as preservatives, antioxidants, colorants, and sweeteners may also be used as needed. Suitable examples of excipients include lactose, sucrose, D-mannitol, D-sorbitol, starch, pregelatinized starch, dextrin, crystalline cellulose, low-substituted hydroxypropyl cellulose, sodium carboxymethylcellulose, gum arabic, dextrin, pullulan, light anhydrous silicic acid, synthetic aluminum silicate, and magnesium aluminometasilicate. Suitable examples of lubricants include magnesium stearate, calcium stearate, talc, colloidal silica, etc. Suitable examples of binders include pregelatinized starch, sucrose, gelatin, gum arabic, methylcellulose, carboxymethylcellulose, sodium carboxymethylcellulose, crystalline cellulose, sucrose, D-mannitol, trehalose, dextrin, pullulan, hydroxypropyl cellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, etc. Suitable examples of disintegrants include lactose, sucrose, starch, carboxymethylcellulose, calcium carboxymethylcellulose, croscarmellose sodium, sodium carboxymethyl starch, light anhydrous silicic acid, low-substituted hydroxypropylcellulose, etc. Suitable examples of solvents include water for injection, physiological saline, Ringer's solution, alcohol, propylene glycol, polyethylene glycol, sesame oil, corn oil, olive oil, cottonseed oil, etc.Suitable examples of solubilizing agents include polyethylene glycol, propylene glycol, D-mannitol, trehalose, benzyl benzoate, ethanol, trisaminomethane, cholesterol, triethanolamine, sodium carbonate, sodium citrate, sodium salicylate, and sodium acetate. Suitable examples of suspending agents include surfactants such as stearyltriethanolamine, sodium lauryl sulfate, laurylaminopropionic acid, lecithin, benzalkonium chloride, benzethonium chloride, and glycerin monostearate; hydrophilic polymers such as polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethylcellulose, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, and hydroxypropylcellulose; polysorbates, and polyoxyethylene hydrogenated castor oil. Suitable examples of isotonic agents include sodium chloride, glycerin, D-mannitol, D-sorbitol, and glucose. Suitable examples of buffering agents include buffer solutions such as phosphates, acetates, carbonates, and citrates. Suitable examples of soothing agents include benzyl alcohol.

[0023] Suitable examples of preservatives include parahydroxybenzoic acid esters, chlorobutanol, benzyl alcohol, phenethyl alcohol, dehydroacetic acid, sorbic acid, etc. Suitable examples of antioxidants include sulfites, ascorbic acid, etc. Suitable examples of coloring agents include water-soluble food tar dyes (e.g., food dyes such as Food Red No. 2 and No. 3, Food Yellow No. 4 and No. 5, Food Blue No. 1 and No. 2, water-insoluble lake dyes (e.g., aluminum salts of the above-mentioned water-soluble food tar dyes), natural dyes (e.g., β-carotene, chlorophyll, red iron oxide, etc.), etc. Suitable examples of sweeteners include saccharin sodium, dipotassium glycyrrhizinate, aspartame, stevia, etc.

[0024] Dosage forms of pharmaceutical compositions include oral preparations such as tablets, capsules (including soft capsules and microcapsules), granules, powders, syrups, emulsions, and suspensions; and parenteral preparations such as injections (e.g., subcutaneous injections, intravenous injections, intramuscular injections, intraperitoneal injections, and intravitreal injections), drip infusions, topical preparations (e.g., intranasal preparations, transdermal preparations, ointments), suppositories (e.g., rectal suppositories, vaginal suppositories), pellets, drip infusions, and sustained-release preparations, each of which can be safely administered orally or parenterally. Pharmaceutical compositions can be prepared by methods commonly used in the pharmaceutical technology field, such as those described in the Japanese Pharmacopoeia. Specific preparation methods for formulations are described in detail below.

[0025] For example, injections are produced by dissolving, suspending, or emulsifying the above-mentioned protein or polypeptide as an active ingredient in an aqueous solvent (e.g., distilled water, physiological saline, Ringer's solution, etc.) or an oily solvent (e.g., vegetable oils such as olive oil, sesame oil, cottonseed oil, and corn oil, propylene glycol, etc.) together with dispersants (e.g., polysorbate 80, polyoxyethylene hydrogenated castor oil 60, etc.), polyethylene glycol, carboxymethylcellulose, sodium alginate, etc.), preservatives (e.g., methylparaben, propylparaben, benzyl alcohol, chlorobutanol, phenol, etc.), isotonicity agents (e.g., sodium chloride, glycerin, D-mannitol, D-sorbitol, glucose, etc.). In this case, additives such as solubilizing agents (e.g., sodium salicylate, sodium acetate, etc.), stabilizers (e.g., human serum albumin, etc.), and soothing agents (e.g., benzyl alcohol, etc.) may be used, if desired.

[0026] Oral preparations are produced by adding, for example, excipients (e.g., lactose, sucrose, starch, D-mannitol, etc.), disintegrants (e.g., carboxymethylcellulose calcium, etc.), binders (e.g., pregelatinized starch, gum arabic, carboxymethylcellulose, hydroxypropylcellulose, polyvinylpyrrolidone, etc.), or lubricants (e.g., talc, magnesium stearate, polyethylene glycol 6000, etc.) to the above-mentioned protein or polypeptide as an active ingredient, followed by compression molding, and then coating with a coating base by a method known per se, as needed, for the purposes of taste masking, enteric coating, or sustained release. Examples of such coating bases include sugar coating bases, water-soluble film coating bases, enteric film coating bases, and sustained-release film coating bases. Sucrose is used as the sugar coating base, and one or more of talc, precipitated calcium carbonate, gelatin, gum arabic, pullulan, carnauba wax, etc. may also be used in combination. Examples of water-soluble film coating bases include cellulose polymers such as hydroxypropyl cellulose, hydroxypropylmethyl cellulose, hydroxyethyl cellulose, and methylhydroxyethyl cellulose; synthetic polymers such as polyvinyl acetal diethylaminoacetate, aminoalkyl methacrylate copolymer E (Eudragit E (trade name), Rohm Pharma Co., Ltd.), and polyvinylpyrrolidone; and polysaccharides such as pullulan.

[0027] Examples of enteric film coating bases include cellulose-based polymers such as hydroxypropyl methylcellulose phthalate, hydroxypropyl methylcellulose acetate succinate, carboxymethylethylcellulose, and cellulose acetate phthalate; acrylic acid-based polymers such as methacrylic acid copolymer L [Eudragit L (trade name), Rohm Pharma Co., Ltd.], methacrylic acid copolymer LD [Eudragit L-30D55 ​​(trade name), Rohm Pharma Co., Ltd.], and methacrylic acid copolymer S [Eudragit S (trade name), Rohm Pharma Co., Ltd.]; and natural products such as shellac. Examples of sustained-release film coating bases include cellulose-based polymers such as ethyl cellulose; and acrylic acid-based polymers such as aminoalkyl methacrylate copolymer RS ​​[Eudragit RS (trade name), Rohm Pharma Co., Ltd.] and ethyl acrylate-methyl methacrylate copolymer suspension [Eudragit NE (trade name), Rohm Pharma Co., Ltd.]. Two or more of the above-mentioned coating bases may be mixed in an appropriate ratio. In addition, a light-shielding agent such as titanium oxide or iron sesquioxide may be used during coating.

[0028] <Method for improving liver lesions caused by lesions in the anterior pituitary gland and / or preventing and / or treating diseases accompanied by the liver lesions> The present invention also relates to a method for improving liver lesions caused by lesions in the anterior pituitary gland and / or preventing and / or treating diseases accompanied by the liver lesions, comprising administering any one of the proteins or polypeptides (a) to (c) above to a recipient. Specific and preferred examples of any one of the proteins or polypeptides (a) to (c), the lesion in the anterior pituitary gland, the liver lesion, the recipient, the administration route, the dosage, the administration frequency, and the like are the same as the specific and preferred examples described above for the first aspect.

[0029] <Screening Method> A second aspect of the present invention is a screening method for an agent for ameliorating liver lesions caused by lesions of the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, the method comprising a screening step using an improvement in IGF-1 production or activity by the liver as an index. In the second aspect, screening means at least narrowing down the population of test substances, and preferably selects candidate agents for preventing and / or treating liver lesions caused by lesions of the anterior pituitary gland and / or for treating diseases accompanied by the liver lesions.

[0030] The reason why the second aspect can at least narrow down the population of candidate ameliorating, preventive and / or therapeutic agents that can significantly improve liver lesions caused by lesions in the anterior pituitary gland and diseases accompanied by the above-mentioned liver lesions is that if the test substance has the effect of "improving the production or activity of IGF-1 by the liver," it can significantly improve liver lesions caused by lesions in the anterior pituitary gland and liver lesions in diseases accompanied by the above-mentioned liver lesions, similar to IGF-1 (mecasermin) (preferably similar to the paracrine or autocrine action of IGF-1) (see Examples described below).

[0031] In the second aspect, specific and preferred examples of anterior pituitary lesions, liver lesions, etc. include the same specific and preferred examples as those described above for the first aspect. In the second aspect, the "step of screening using the improvement of IGF-1 production or activity by the liver as an index" preferably comprises the steps of: administering a test substance to an animal in vivo; determining a quantitative or qualitative state regarding IGF-1 production or activity by the liver (for example, determining serum transaminase levels, liver size (cm), γ-glutamyltransferase levels, etc.); and selecting, based on the results of the determination, a candidate agent for improving liver lesions caused by anterior pituitary lesions and / or for preventing and / or treating diseases accompanied by the liver lesions.

[0032] The route of administration of the test substance to the animals is not particularly limited and may be systemic or local administration. Continuous administration or bolus administration may also be used, with continuous administration being preferred. Administration may also be percutaneous (subcutaneous, intramuscular, intravenous, or other injection), intracerebroventricular (injection), oral, or intraperitoneal (injection), but subcutaneous administration is preferred from the viewpoints of minimal invasiveness, lack of limitations due to body movement, etc.

[0033] Examples of the serum transaminase level include ALT level, AST level, etc. Examples of the animal include any mammal other than humans (mouse, rat, pig, cow, monkey, etc.), and preferred are pathological model animals (mouse, rat, pig, cow, monkey, etc.) of liver lesions caused by lesions in the anterior pituitary gland (preferably, the diseases accompanied by liver lesions (NPC, etc.)). Examples of pathological model animals of liver lesions caused by lesions in the anterior pituitary gland (preferably, the diseases accompanied by liver lesions (NPC, etc.)) include, for example, Npc1 －／－ Animals (Npc1 gene-deficient animals (BALB / cNctr-Npc1 ｍ１Ｎ )) etc.

[0034] The measured value S1 in the presence of the test substance can be used as an index for identification, and the measured value S1 is preferably compared with the measured value S2 under control conditions. For example, when the "improvement in IGF-1 production or activity by the liver" is observed, the test substance is preferably identified as a candidate agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions.

[0035] Here, "control conditions" refers to conditions in the absence of a test substance, and means that the difference in measured values ​​between the presence and absence of the test substance (the above-mentioned "improvement in IGF-1 production or activity by the liver") is used as an index. The measured value S2 under the control conditions may be a measured value before the addition of the test substance, or may be a statistical value or range obtained by collecting data in advance.

[0036] For example, in the case of serum transaminase levels, liver size (cm), γ-glutamyltransferase levels, etc., if the measured value S1 is lower than the measured value S2 under the control conditions, it can be said that the "improvement in IGF-1 production or activity by the liver" has been confirmed.

[0037] The degree of the reduction is more preferably 3 / 4 or less, even more preferably 3 / 5 or less, particularly preferably 1 / 2 or less, especially preferably 1 / 3 or less, and most preferably 1 / 4 or less, compared to the control condition. There is no particular restriction on the lower limit of the reduction, but examples include 1 / 30 or more, 1 / 20 or more, and 1 / 10 or more. The degree of improvement is more preferably 1.2 times or more, even more preferably 1.5 times or more, especially preferably 2 times or more, especially preferably 3 times or more, and most preferably 4 times or more, compared to the control condition. There is no particular restriction on the upper limit of the improvement, but examples include 30 times or less, 20 times or less, and 10 times or less.

[0038] In the second aspect, the "screening step using the improvement of IGF-1 production or activity by the liver as an index" may include measuring the binding activity of a test substance to a target protein of IGF-1 (for example, at least one target protein selected from the group consisting of IGF-binding proteins (IGFBP)-1 to IGFBP-6 and IGF-1 receptor) in vitro, and using the measured intensity as an equivalent or alternative to "an index of the improvement of IGF-1 production or activity by the liver," to select candidate agents for improving liver lesions caused by lesions of the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by the liver lesions, based on the measurement results.

[0039] The method for measuring the binding activity of a test substance to a target protein of IGF-1 is not particularly limited. For example, the binding activity can be measured by a known method for measuring binding between substances. Examples of such methods include isothermal titration calorimetry (ITC), surface plasmon resonance (SPR), nuclear magnetic resonance (NMR), and fluorescence correlation spectroscopy (FCS).

[0040] Additionally, the strength of the response due to IGF-1 (mecasermin) may or may not be used as a benchmark "control condition."

[0041] (Test Substance) The test substance is not particularly limited. The test substance may consist of a single component (i.e., a pure substance) or a combination of two or more components (i.e., a mixture). When the test substance is a mixture, the number and composition ratio of the components constituting the mixture are not particularly limited. The test substance may be a known substance or a novel substance. The test substance may be a natural product or an artificial product. The type of test substance is not particularly limited, and may be a nucleic acid molecule (preferably an aptamer), an antibody, an individual low-molecular-weight synthetic compound (e.g., alcohols, ketones, aldehydes, ethers, esters, hydrocarbons, sugars, organic acids, nucleic acids, amino acids, peptides, lipids, and various other organic or inorganic components), a compound present in a natural product extract, or a synthetic peptide. Alternatively, the test substance may be a chemical library, a phage display library, or a combinatorial (compound) library created using combinatorial chemistry technology. Construction of a compound library is known to those skilled in the art, and commercially available compound libraries can also be used. The test substance is preferably a protein, (poly)peptide, low-molecular-weight compound (e.g., a compound library), nucleic acid molecule (preferably an aptamer), or antibody (monoclonal or polyclonal antibody), and from the viewpoint of high specificity for the target protein, a protein, (poly)peptide, nucleic acid molecule (preferably an aptamer), or monoclonal antibody is more preferred.

[0042] Here, an aptamer refers to a nucleic acid molecule composed of single-stranded RNA or DNA, which can bind to a target protein due to its three-dimensional structure and act as an agonist (enhancing activity) or antagonist (inhibiting activity) (Drug Delivery System 31-1, 2016, pp. 10-14). Aptamers have high binding affinity and specificity for target proteins, low immunogenicity, can be produced by chemical synthesis (preferably obtained by the SELEX method), and have high storage stability. The SELEX method refers to a method of screening for nucleic acid aptamers that selectively (preferably specifically) bind to a target protein from a mixture of nucleic acids called a nucleic acid library. The base length of the aptamer used as the test substance, which selectively (preferably specifically) binds to a target protein of IGF-1 (e.g., at least one target protein selected from the group consisting of IGF binding protein (IGFBP)-1 to IGFBP-6 and IGF-1 receptor), is not particularly limited as long as it selectively (preferably specifically) binds to a target protein of IGF-1 (e.g., at least one target protein selected from the group consisting of IGFBP-1 to IGFBP-6 and IGF-1 receptor), but is preferably 15 to 60 bases, more preferably 20 to 50 bases, even more preferably 25 to 47 bases, and particularly preferably 26 to 45 bases. Furthermore, polyclonal antibodies can be prepared by separating and purifying serum obtained from an animal immunized with an antigen (e.g., at least one target protein selected from the group consisting of IGFBP-1 to IGFBP-6 and IGF-1 receptor). Monoclonal antibodies can be prepared by fusing antibody-producing cells obtained from an animal immunized with an antigen (e.g., at least one target protein selected from the group consisting of IGFBP-1 to IGFBP-6 and IGF-1 receptor) with myeloma cells to produce hybridomas, culturing the hybridomas or administering them to an animal to cause ascites tumorigenesis in the animal, and isolating and purifying the culture medium or ascites fluid.

[0043] Examples of proteins and (poly)peptides as test substances include any or known proteins or (poly)peptides having insulin-like activity and growth-promoting activity, insulin-like growth factors (e.g., IGF-2), which are polypeptides highly similar in sequence to insulin, and splice variants of IGF-1 (e.g., mechano-growth factor: MGF). Furthermore, proteins and (poly)peptides as test substances are preferably selected from the proteins or polypeptides of (b) or (c) above.

[0044] Examples of low molecular weight compounds (e.g., compound libraries) to be tested include any or known compounds that have the effect of promoting growth hormone secretion, any or known compounds that have the effect of improving the production or activity of IGF-1 by the liver, and the like. For example, US Patent No. 3,239,345, US Patent No. 4,036,979, US Patent No. 4,411,890, US Patent No. 5,206,235, US Patent No. 5,284,841, US Patent No. 5,310,737, US Patent No. 5,317,017, European Patent Publication No. 0144,230, European Patent Publication No. 0513,974, PCT Patent Publication No. WO94 / 07486, PCT Patent Publication No. WO94 / 08583, PCT Patent Publication No. WO94 / 13696, Science, 260,1640-1643 (June 11, 1993) and the like compounds described in the compound library containing the compound group or the compound group may not be used as a test substance. Specific examples of low molecular weight compounds (e.g., compound libraries) as test substances include spiropiperidine compounds, benzolactam compounds, and the like, and more preferably, the compounds are selected from the spiropiperidine compounds described in PCT Patent Publication No. WO94 / 13696 and the benzolactam compounds described in U.S. Patent No. 5,206,235.

[0045] The spiropiperidine compound is preferably a compound represented by the following formula I or II, a pharmaceutically acceptable salt thereof, or each diastereomer.

[0046] [In the above formulas I and II, R １ is -C １ ~C１０ Alkyl, -aryl, -aryl-(C １ ~C ６ alkyl), -C ３ ~C ７ Cycloalkyl-(C １ ~C ６ alkyl), -C １ ~C ５ Alkyl-K-C １ ~C ５ Alkyl, -aryl (C ０ ~C ５ alkyl)-K-(C １ ~C ５ alkyl), -C ３ ~C ７ Cycloalkyl (C ０ ~C ５ alkyl)-K-(C １ ~C ５ K is selected from the group consisting of O, S(O) ｍ , N(R ２ )C(O), C(O)N(R ２ ), OC(O), C(O)O or -CR ２ =CR ２ - or -C≡C-, in which case the aryl group is as defined below, and R ２ and the alkyl group may further comprise 1 to 9 halogens, S(O) ｍ R ２ａ , 1 to 3 OR ２ａ or C(O)OR ２ａ The aryl group may be further substituted by phenyl, phenoxy, halophenyl, 1-3 C １ ~C ６ Alkyl, 1 to 3 halogens, 1 to 2 -OR ２ , methylenedioxy, —S(O) ｍ R ２ , 1 to 2 -CF ３ , -OCF ３ , nitro, -N(R ２ ) (R ２ ), -N(R ２ ) C(O)R ２ , -C(O)OR ２ , -C(O)N(R ２ ) (R２ ), -SO ２ N (R ２ ) (R ２ ), -N(R ２ ) S (O) ２ Aryl and —N(R ２ ) SO ２ (R ２ ) may be substituted by R ２ is hydrogen, C １ ~C ６ Alkyl, C ３ ~C ７ cycloalkyl, and two C １ ~C ６ When alkyl groups are present on one atom, the groups taken together may not be present on an oxygen, sulfur or NR ２ａ C may include ３ ~C ８ may form a ring, R ２ａ is hydrogen or C １ ~C ６ alkyl, R ３ａ and R ３ｂ is hydrogen, halogen, -C １ ~C ６ Alkyl, -OR ２ , cyano, -OCF ３ , methylenedioxy, nitro, -S(O) ｍ R, -CF ３ or -C(O)OR ２ and R ３ａ and R ３ｂ are in the ortho configuration, they may be combined with each other to form a C which may contain one or two heteroatoms selected from oxygen, sulfur or nitrogen. ５ ~C ８ may form an aliphatic ring or aromatic ring of the formula R ４ and R ５ is hydrogen, -C １ ~C ６ Alkyl, substituted C １ ~C ６ Alkyl (the substituents are 1 to 5 halogens, 1 to 3 hydroxyl groups, 1 to 3 C １ ~C １０Alkanoyloxy, 1-3 C １ ~C ６ Alkoxy, phenyl, phenoxy, 2-furyl, C １ ~C ６ Alkoxycarbonyl, —S(O) ｍ (C １ ~C ６ alkyl), or R ４ and R ５ Together - (CH ２ ) ｒ L ａ (CH ２ ) ｓ -, in which case L ａ HA-C(R ２ ) ２ -, O-, -S(O) ｍ - or -N(R ２ )-, r and s are independently integers from 1 to 3, R ２ is as defined above, and R ６ is hydrogen or C １ ~C ６ A is alkyl; or In the above formula, x and y are independently integers of 0 to 3, and Z is N-R ２ or O, R ７ and R ７ａ is hydrogen, -C １ ~C ６ Alkyl, -OR ２ , trifluoromethyl, phenyl, substituted C １ ~C ６ Alkyl (the substituents are imidazolyl, phenyl, indolyl, p-hydroxyphenyl, -OR ２ , 1 to 3 fluorines, —S(O) ｍ R ２ , -C(O)OR ２ , -C ３ ~C ７ cycloalkyl, —N(R ２ ) (R ２ ), -C(O)N(R ２ ) (R ２ ) or R７ and R ７ａ is independently R ４ and R ５ together with one or both of the groups ７ or R ７ａ and B, D, E, and F can form an alkylene bridge between the alkyl portions of the groups, the bridge having 1 to 5 carbon atoms, and ８ ) (R １０ )-, -O-, C=O, -S(O) ｍ -or-NR ９ -, and one or two of B, D, E or F may be absent to form a 5-, 6- or 7-membered ring, provided that B, D, E and F are not -C(R ８ ) (R １０ )- or C=O, provided that one of the remaining B, D, E and F groups is simultaneously -O-, -S(O) ｍ -or-NR ９ - or B and D or D and E together form -N=CR １０ -or-CR １０ =N-, or B and D or D and E together form -CR ８ =CR １０ - (However, the other of B and E or one of F is simultaneously -O-, -S(O) ｍ -or-NR ９ -) if and only if R ８ and R １０ is hydrogen, -R ２ , -OR ２ , -(CH ２ ) ｑ -aryl, -(CH ２ ) ｑ -C(O)OR ２ , -(CH ２ ) ｑ -C(O)O(CH ２ ) ｑ -aryl or -(CH ２ ) ｑ -(1H-tetrazol-5-yl), wherein aryl is independently selected from the group consisting of 1 to 3 halogens, 1 to 2 C１ ~C ８ alkyl, 1 to 3 -OR ２ or 1 to 2 -C(O)OR ２ and R ９ is -R ２ , -(CH ２ ) ｑ -aryl, -C(O)R ２ , —C(O)(CH ２ ) ｑ -aryl, -SO ２ R ２ , -SO ２ (CH ２ ) ｑ -aryl, -C(O)N(R ２ ) (R ２ ), -C(O)N(R ２ ) (CH ２ ) ｑ -aryl, -C(O)OR ２ , 1H-tetrazol-5-yl, —SO ３ H, -SO ２ NHC≡N, -SO ２ N (R ２ ) aryl, —SO ２ N (R ２ ) (R ２ ), wherein (CH ２ ) ｑ is 1-2 C １ ~C ４ may be substituted by alkyl, R ２ and aryl is 1 to 3 -OR ２ａ , -O(CH ２ ) ｑ aryl, 1 to 2 —C(O)OR ２ａ , 1 to 2 -C(O)O(CH ２ ) ｑ aryl, 1 to 2 —C(O)N(R ２ａ ) (R ２ａ ), 1 to 2 -C(O)N(R ２ａ ) (CH ２ ) ｑ aryl, 1 to 5 halogens, 1 to 3 C １ ~C ４Alkyl, 1,2,4-triazolyl, 1H-tetrazol-5-yl, —C(O)NHSO ２ R ２ａ , -S(O) ｍ R ２ａ , -C(O)NHSO ２ (CH ２ ) ｑ Aryl, —SO ２ NHC≡N, -SO ２ NHC(O)R ２ａ , -SO ２ NHC(O)(CH ２ ), aryl, —N(R ２ )C(O)N(R ２ａ ) (R ２ａ ), -N(R ２ａ )C(O)N(R ２ａ ) (CH ２ ) ｑ Aryl, —N(R ２ａ ) (R ２ａ ), -N(R ２ａ ) C(O)R ２ａ , -N(R ２ａ )C(O)(CH ２ ) ｑ Aryl, —OC(O)N(R ２ａ ) (R ２ａ ), -OC(O)N(R ２ａ ) (CH ２ ) ｑ Aryl, —SO ２ (CH ２ ) ｑ CONH-(CH ２ ) ｗ NHC(O)R １１ (where w is 2 to 6 and R １１ is biotin, aryl, or one or two OR ２, and aryl substituted with 1 to 2 halogens, azide, or nitro; m is 0, 1, or 2; n is 1 or 2; q can be 0, 1, 2, 3, or 4; G, H, I, and J are carbon, nitrogen, sulfur, or oxygen atoms, in which case at least one is a heteroatom, and one of G, H, I, or J may be absent to provide a 5- or 6-membered heteroaromatic ring.

[0047] The benzolactam compound is preferably a compound represented by the following formula:

[0048]

[0049] [In the above formula, L is n is 0 or 1, p is 0 to 3, q ​​is 0 to 4, w is 0 or 1, and x is m is 0 to 2, and R １ , R ２ , R １ａ , R ２ａ , R １ｂ and R ２ｂ are independently hydrogen, halogen, C １ -C ７ Alkyl, C １ -C ３ Perfluoroalkyl, C １ -C ３ Perfluoroalkoxy, —S(O) ｍ R ７ａ , cyano, nitro, R ７ｂ O (CH ２ ) Ｖ -, R ７ｂ COO (CH ２ ) ｖ -, R ７ｂ OCO (CH ２ ) ｖ -, phenyl or substituted phenyl (substituents are 1 to 3 halogens, C １ -C ６ Alkyl, C １ -C ６ alkoxy, or hydroxy) (v is 0 to 3); R ７ａ and R ７ｂare independently hydrogen, halogen, C １ -C ３ Perfluoroalkyl, C １ -C ６ Alkyl, substituted C １ -C ６ Alkyl (the substituents are phenyl or substituted phenyl, and the phenyl substituents are halogen, C １ -C ６ Alkyl, C １ -C ６ alkoxy, or hydroxy; ３ａ and R ３ｂ are independently hydrogen, R ９ , R ９ C substituted with １ -C ６ Alkyl, R ９ phenyl substituted with, or R ９ phenoxy substituted with R ９ teeth R ７ｂ O (CH ２ ) ｖ -, R ７ｂ COO (CH ２ ) ｖ -, R ７ｂ OCO (CH ２ ) ｖ -, R ７ｂ CO(CH ２ ) ｖ -, R ７ｂ O (CH ２ ) ｖ CO-, R ４ R ５ N (CH ２ ) ｖ -, R ７ｂ CON (R ４ ) (CH ２ ) ｖ -, R ４ R ５ NCO(CH ２ ) ｖ -, R ４ R ５ NCS (CH ２ ) ｖ -, R ４ R ５ NN (R ５ )CO(CH ２ )ｖ -, R ４ R ５ NN (R ５ ) CS (CH) ｖ -, R ７ｂ CON (R ４ ) N (R ５ )CO(CH ２ ) ｖ -, R ７ｂ CON (R ４ ) N (R ５ ) CS (CH ２ ) ｖ -, R ４ N (OR ７ｂ )CO(CH ２ ) ｖ - or R ７ａ CON (OR ７ｂ )CO(CH ２ ) ｖ - (wherein v is as defined above), and R ４ , R ４ａ , R ５ are independently hydrogen, phenyl, substituted phenyl, C １ -C １０ Alkyl, substituted C １ -C １０ Alkyl, C ３ -C １０ Alkenyl, substituted C ３ -C １０ Alkenyl, C ３ -C １０ Alkynyl, or substituted C ３ -C １０ Alkynyl (phenyl, alkyl, alkenyl, alkynyl substituents are 1 to 5 hydroxy, C １ -C ６ Alkoxy, C ３ -C ７ Cycloalkyl, phenyl C １ -C ３ Alkoxy, Fluoro, R １ Substitution or R １ and R ２ Phenyl C independently substituted with １ -C ３ Alkoxy, phenyl, R １ Substitution or R １ and R ２phenyl independently substituted with two of the following, wherein the substituents on the phenyl are as defined above; C １ -C ５ Alkanoyloxy, C １ -C ５ Alkoxycarbonyl, carboxy, formyl, or NR １０ R １１ (R １０ and R １１ are independently hydrogen, C １ -C ６ Alkyl, phenyl, phenyl C １ -C ６ Alkyl, C １ -C ５ Alkoxycarbonyl, or C １ -C ５ Alkanoyl C １ -C ６ alkyl), or R ４ and R ５ Let's go together-(CH ２ ) ｒ B (CH ２ ) ｓ - (B is CH ２ , O or S(O)m, or N—R １０ and r and s are independently 1 to 3; R １０ is as defined above), and R ６ is hydrogen, C １ -C １０ Alkyl, phenyl or phenyl C １ -C １０ A is alkyl; wherein x and y are independently 0-3; ８ and R ８ａ are independently hydrogen, C １ -C １０ Alkyl, trifluoromethyl, phenyl, substituted C １ -C １０ Alkyl (substituted with 1 to 3 of imidazolyl, indolyl, hydroxy, fluoro, S(O) ｍ R ７ａ , C １ -C ６ Alkoxy, C ３ -C ７ Cycloalkyl, phenyl C１ -C ３ Alkoxy, R １ Substitution or R １ and R ２ Phenyl C independently substituted with １ -C ３ Alkoxy, phenyl, R １ Substitution or R １ and R ２ phenyl independently substituted with １ -C ５ Alkanoyloxy, C １ -C ５ Alkoxycarbonyl, carboxy, formyl, or NR １０ R １１ (R １０ and R １１ is as defined above), or R ８ and R ８ａ Let's go together-(CH ｚ )t-(t is 2 to 6), or R ８ and R ８ａ are independently R ４ and R ５ may be taken together with either one or both of the following to form an alkyl bridge connecting the terminal nitrogen to the alkyl portion of the A group, the bridge containing 1 to 5 carbon atoms.

[0050] The present invention will be explained in more detail below by showing examples of the present invention, but the present invention is not limited to these examples and various applications are possible within the scope of the technical idea of ​​the present invention.

[0051] Reference Example 1 (Hypophysectomy of a Mouse Model of Disease Accompanied by Liver Lesions Caused by Lesions in the Anterior Pituitary Gland and Measurement of Mouse Serum IGF-1 Concentration) Npc1 is a mouse model of a disease accompanied by liver lesions (NPC, etc.) caused by lesions in the anterior pituitary gland. －／－ Mouse (Npc1 gene-deficient mouse (BALB / cNctr-Npc1 ｍ１Ｎ To evaluate pituitary hypoplasia in mice, Npc1 mice were used at 8 weeks of age. －／－The pituitary glands were removed from the mice and their weights were measured. In addition, the serum IGF-1 concentration, which is evaluated as a downstream signal of growth hormone (GH) secreted from the pituitary gland, was measured. －／－ Blood was collected from the inferior vena cava of rat and wild-type mice under isoflurane anesthesia. Serum IGF-1 concentrations were measured using a Mouse / Rat IGF1 ELISA Kit (Proteintech). The results are shown in Figures 1A and 1B. In Figure 1, values ​​are expressed as the mean ± standard error for n = 4 to 6 in each group, and the significance level relative to the control group (wild-type mice) is *: p < 0.05.

[0052] (Results) As is clear from the results shown in Figure 1A, pituitary weight was significantly lower in Npc1 gene-deficient mice compared to control (wild-type) mice. Furthermore, as is clear from the results shown in Figure 1B, serum IGF-1 concentrations were also significantly lower compared to controls. As described above, it was suggested that pituitary insufficiency occurs in liver lesions caused by lesions of the anterior pituitary gland (preferably, diseases accompanied by liver lesions (NPC, etc.) as described above), resulting in decreased production and / or secretion of IGF-1. This suggests the need for IGF-1 supplementation in liver lesions caused by lesions of the anterior pituitary gland (preferably, diseases accompanied by liver lesions (NPC, etc.) as described above).

[0053] Example 1 (Test of daily subcutaneous administration of mecasermin to pathological model mice with diseases (NPC, etc.) accompanied by liver lesions due to lesions in the anterior pituitary gland) Mecasermin (human IGF-1 preparation mecasermin (genetically recombinant)) was prepared once a week on a clean bench by dissolving lyophilized mecasermin powder in physiological saline to a concentration of 10 mg / mL and a pH of 1.8 to 3.8, and after preparation, it was stored at 4°C in the dark. －／－Mice were subcutaneously administered every 12 hours in the following order: neck, back, and rump. Mecasermin (human IGF-1 recombinant) was administered subcutaneously at 0.1-2 mg / kg / day twice daily. The dosage was determined weekly at the time of body weight measurement. Administration began at 6 weeks of age and continued until 9 weeks of age, when pathology due to anterior pituitary lesions became evident. Blood was collected from the inferior vena cava under isoflurane anesthesia. After incubation at room temperature (30 minutes), the serum was collected by centrifugation (4°C, 1000 g, 10 minutes). Serum ALT and AST levels were measured using a Fuji DryChem 7000 "Z" (Fujifilm Corporation). The results are shown in Figures 2A and 2B. In Figure 2, values ​​are expressed as mean ± standard error for n = 4-6 in each group. **: p < 0.01 for significance relative to the 0 mg / kg / day group (physiological saline administration).

[0054] (Results) As is clear from the results shown in Figures 2A and 2B, the mecasermin-administered group showed significantly lower serum transaminase levels (ALT and AST) than the increase observed in pathological model mice with diseases (such as NPC) accompanied by liver lesions due to lesions in the anterior pituitary gland. Compared to the increase in serum transaminase levels (ALT and AST) observed in pathological model mice with diseases (such as NPC) accompanied by liver lesions due to lesions in the anterior pituitary gland, it was found that doses of 1.0 (mg / kg / day) or more (daily) showed significantly lower levels. Furthermore, a comparison of serum transaminase levels at doses of 1.0 (mg / kg / day) and 2.0 (mg / kg / day) suggests that the improvement effect is saturated at doses of 1.0 (mg / kg / day) or more. As described above, it has been suggested that mecasermin has a remarkable effect on improving the pathological condition of liver lesions caused by lesions in the anterior pituitary gland (preferably the above-mentioned diseases accompanied by liver lesions (NPC, etc.)).

[0055] Comparative Example (Verification of the therapeutic effect of miglustat in the above-mentioned disease model mice) Bresaves (registered trademark) capsules (miglustat capsules containing miglustat; manufactured by Janssen Pharmaceuticals) were decapsulated and subjected to the same procedure as described in Zervas et al. Curr Biol 2001 Aug21;11(16):1283-7. doi:10.1016 / s0960-9822(01)00396-7. and Davidson et al. PLoS One 2009 Sep11;4(9):e6951. doi:10.1371 / journal.pone.0006951. According to the method of (1), the compound was mixed with powdered feed and administered daily to NPC mice, the above-mentioned pathological model mice, as follows, but many mice died from significant digestive disorders. Subsequently, the dosage was reduced with reference to Nietuppski et al. Mol Genet Metab. 2012 Apr; 105(4): 621-8. doi: 10.1016 / j. ymgme. 2012.01.020. Epub 2012 Feb 1. PMID: 22366055. 1200 mg / kg / day: 6 / 6 mice died. 600 mg / kg / day: 1 / 4 mice died (3 mice started administration at 5 weeks of age). 450 mg / kg / day: 3 / 5 mice died.

[0056] Figure 3 shows the results of verifying the therapeutic effect of miglustat in the above-mentioned pathological model mice. As is clear from the results shown in Figure 3, although the neuropathy score in surviving individuals tended to improve (not shown), the serum ALT levels were almost the same as those in NPC mice, indicating that miglustat does not have a therapeutic effect on liver damage.

[0057] Reference Example 2 (Correlation between serum IGF-1 concentration and liver damage marker ALT value in the above-mentioned pathological model mouse) －／－ The serum IGF-1 concentration and the liver damage marker ALT level of the mice were measured, and the Pearson correlation coefficient was calculated to evaluate the correlation between the serum IGF-1 concentration and the liver damage marker ALT level of the model mice. The results are shown in Figure 4 (n = 17).

[0058] 4 shows a significant negative correlation between the serum IGF-1 concentration and the liver damage marker ALT level in the pathological model mice. As is clear from the results shown in FIG. 4, the correlation coefficient r was −0.58, and the coefficient of determination r ２ = 0.336, p < 0.05, indicating a significant negative correlation between serum IGF-1 concentrations in the pathological model mice and the liver damage marker ALT levels. In other words, it can be said that there is a significant negative correlation between liver lesions caused by lesions in the anterior pituitary gland and serum IGF-1 concentrations.

Claims

1. An agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by said liver lesions, comprising any one of the following proteins or polypeptides (a) to (c): (a) a protein or polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, (b) a protein or polypeptide consisting of an amino acid sequence in which one or more amino acids have been deleted, substituted and / or added in the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, and which has insulin-like and growth-promoting effects, or (c) a protein or polypeptide consisting of an amino acid sequence having 95% or more sequence identity with the amino acid sequence set forth in SEQ ID NO: 1 in the Sequence Listing, and which has insulin-like and growth-promoting effects.

2. The agent according to claim 1, wherein the disease accompanied by liver lesions is at least one disease selected from the group consisting of diseases accompanied by central nervous system disorders, childhood diseases, and genetic diseases.

3. The agent according to claim 1, wherein the disease accompanied by liver lesions is Niemann-Pick disease type C.

4. The agent according to claim 1, for administration to a patient with Niemann-Pick disease type C who has liver lesions caused by Niemann-Pick disease type C.

5. The agent according to any one of claims 1 to 4, which is at least one selected from the group consisting of subcutaneous administration and continuous administration.

6. A method for screening for an agent for improving liver lesions caused by lesions in the anterior pituitary gland and / or for preventing and / or treating diseases accompanied by such liver lesions, the method comprising a step of screening using an improvement in the production or activity of IGF-1 by the liver as an indicator.

7. The method according to claim 6, wherein the disease accompanied by liver lesions is at least one disease selected from the group consisting of diseases accompanied by central nervous system disorders, childhood diseases, and genetic diseases.

8. The method according to claim 6, wherein the disease associated with liver lesions is Niemann-Pick disease type C.

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