Peptides having inhibitory activity against amyloid precursor protein and use thereof

Abstract

The present application relates to a peptide having inhibitory activity against amyloid precursor protein and uses thereof, and provides a peptide consisting of any one of the amino acid sequences set forth in SEQ ID NOs: 1 to 8 and SEQ ID NOs: 20 to 23, and a pharmaceutical composition for preventing or treating degenerative neurological diseases comprising the peptide as an active ingredient.

Description

[Technical Field]

[0001] This application relates to a peptide having inhibitory activity against amyloid precursor protein and its use. This patent application claims priority to Korean Patent Application No. 10-2021-0163629 filed with the Korean Intellectual Property Office on November 24, 2021, and Korean Patent Application No. 10-2021-0163630 filed with the Korean Intellectual Property Office on November 24, 2021, the disclosures of which are incorporated herein by reference. [Background technology]

[0002] Alzheimer's disease (AD) is a disease characterized by gradual memory loss and cognitive impairment, and its incidence has been increasing significantly as the average human lifespan has increased. One of the pathological characteristics of AD is the accumulation of senile plaques outside of neurons, and the causative agent is β-amyloid (Aβ).

[0003] β-amyloid (Aβ) is a protein fragment cleaved from the amyloid precursor protein (APP), and the most important enzyme involved in β-amyloid production is β-secretase, named β-site APP-cleaving enzyme (BACE1). APP cleaved by β-secretase (BACE1) is separated into an N-terminal domain called sAPPβ, which is approximately 90 kDa, and a cytoplasmic domain called CTFβ (C99).The sAPPβ thus produced is secreted outside the cell, and C99 is further cleaved by γ-secretase to produce 4 kDa β-amyloid.

[0004] Abnormal metabolism of amyloid precursor protein (APP) leads to the production of large amounts of β-amyloid (Aβ), which causes brain cell toxicity and leads to the development of Alzheimer's disease. Therefore, substances that inhibit the activity of Aβ will inhibit the production of β-amyloid and are therefore expected to be effective in preventing or treating Alzheimer's disease.

[0005] The majority of drugs currently on the market or under development are designed to improve the quality of life of Alzheimer's patients by improving their symptoms, including acetylcholinesterase inhibitors such as tacrine, donepezil, rivastigmine, and galantamine, which were developed based on the cholinergic nervous system hypothesis, and memantine, which has NMDA-glutamate receptor inhibitory function. However, most of these treatments only temporarily improve clinical symptoms in the early stages of the disease, and they also have side effects, making treatment difficult (Korean Patent Registration No. 10-2085358).

[0006] The present inventors have been conducting research to develop a therapeutic agent capable of inhibiting the production of and decomposing β-amyloid protein, with the aim of fundamentally treating Alzheimer's disease based on the cause of its onset. As a result, they have developed a substance that inhibits the expression and / or activity of amyloid precursor protein, and have completed the present invention based on this substance. Summary of the Invention [Problem to be solved by the invention]

[0007] One aspect is to provide a peptide consisting of any one of the amino acid sequences of SEQ ID NO:1 to SEQ ID NO:8 and SEQ ID NO:20 to SEQ ID NO:23.

[0008] Another aspect of the present invention is to provide a pharmaceutical composition for preventing or treating degenerative neurological diseases, which comprises the peptide as an active ingredient.

[0009] Yet another aspect is to provide a pharmaceutical use of said peptide for the prevention or treatment of a degenerative neurological disease, or a method for treating a degenerative neurological disease comprising administering to an individual a therapeutically effective amount of said peptide.

[0010] Other objects and advantages of the present application will become more apparent from the following detailed description taken in conjunction with the accompanying claims and drawings. Contents not described in this specification can be fully understood and inferred by those skilled in the art of the present application or a similar art, and therefore, the description thereof will be omitted. [Means for solving the problem]

[0011] Each description and embodiment disclosed in this application is applicable to each other description and embodiment. In other words, all combinations of the various elements disclosed in this application fall within the scope of this application. Furthermore, the scope of this application should not be limited by the specific description below.

[0012] One aspect provides a peptide consisting of any one of the amino acid sequences set forth in SEQ ID NOs: 1 to 8 and SEQ ID NOs: 20 to 23.

[0013] As used herein, the term "peptide" refers to a linear molecule formed by the binding of amino acid residues to each other via peptide bonds. The peptide can be prepared by chemical synthesis methods known in the art, particularly solid-phase synthesis or liquid-phase synthesis (U.S. Patent No. 5,516,891). The present inventors have made extensive efforts to develop peptides having biologically effective activities, and have identified peptides consisting of any one of the amino acid sequences set forth in SEQ ID NOs: 1 to 8. Here, the biologically effective activity may be any one or more of the following: (a) inhibition of amyloid precursor protein (APP) expression; (b) increased expression of ADAM10 (A Disintegrin And Metalloproteinase-containing protein 10) or ADAM17 (A Disintegrin And Metalloproteinase-containing protein 17) protein; (c) increased expression of IDE (insulin degrading enzyme) protein; (d) increased expression of anti-inflammatory cytokines; (e) promotion of neurite outgrowth; and (f) increased expression of brain-derived neurotrophic factor (BDNF).

[0014] Therefore, the peptide can be used to prevent or treat degenerative neurological diseases.

[0015] The peptide may have a protecting group attached to its N- or C-terminus to achieve chemical stability, enhanced pharmacological properties (e.g., half-life, absorption, potency, efficacy), altered specificity (e.g., a broader spectrum of biological activity), or reduced antigenicity. In one embodiment, the N-terminus of the peptide may be attached to any one protecting group selected from the group consisting of an acetyl group, a fluorenylmethoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, and polyethylene glycol (PEG); and / or the C-terminus of the peptide may be attached to any one protecting group selected from the group consisting of an amino group (-NH) and an azide (-NHNH). The peptides may also optionally further comprise targeting sequences, tags, labeled residues, or specifically engineered amino acid sequences to increase half-life or peptide stability.

[0016] The peptides are artificially synthesized or non-naturally occurring or engineered, where "non-naturally occurring or engineered" means that the peptides are produced by artificial modification rather than in the natural state. Here, the artificial modification may include artificially synthesizing an amino acid sequence by mimicking a plurality of amino acid structures, or engineering the peptides to obtain chemical stability, enhanced pharmacological properties, altered specificity, or reduced antigenicity, as described above.

[0017] As used herein, the term "stability" may refer not only to in vivo stability, which protects the peptide from attack by in vivo proteolytic enzymes, but also to storage stability (eg, storage stability at room temperature).

[0018] Another aspect provides a pharmaceutical composition for preventing or treating a degenerative neurological disease, comprising as an active ingredient a peptide consisting of any one of the amino acid sequences set forth in SEQ ID NOs: 1 to 8 and 20 to 23.

[0019] Among the terms and elements mentioned in the description of the peptide, those that are the same as those already mentioned are as described above.

[0020] As used herein, the term "prevention" refers to any action that inhibits or delays the onset of a disease by administering the composition.

[0021] As used herein, the term "treatment" refers to any form of care that provides a benefit to an individual suffering from or at risk of developing a disease, including improving the individual's condition (e.g., one or more symptoms), delaying disease progression, delaying the onset of symptoms, or slowing the progression of symptoms, etc. Thus, the terms "treatment" and "prevention" are not intended to mean a cure or complete elimination of symptoms.

[0022] The term "individual" refers to a subject in need of treatment for a disease, and more specifically refers to mammals such as human or non-human primates, mice, dogs, cats, horses, and cattle.

[0023] The "degenerative neurological disease", which is a target disease to be prevented or treated by the pharmaceutical composition, refers to a disease in which degenerative changes occur due to the gradual and steady death of nerve cells in the nervous system, causing various symptoms. Examples of the degenerative diseases include Alzheimer's disease, Parkinson's disease, Huntington's disease, multiple sclerosis, mild cognitive impairment, cerebral amyloid angiopathy, amyloid stroke, systemic amyloidosis, Dutch amyloidosis, Niemann-Pick disease, senile dementia, amyotrophic lateral sclerosis, spinocerebellar atrophy, Tourette's syndrome, Friedreich's ataxia, Machado-Joseph's disease, Lewy body dementia, dystonia, and progressive supranuclear palsy. The dementia may be selected from the group consisting of Alzheimer's disease, Alzheimer's disease, encephalopathy, frontotemporal dementia, peripheral neuropathy, and polyneuropathy, and is preferably, but not limited to, Alzheimer's disease.

[0024] According to one embodiment, the peptides (peptide-1 to peptide-8) can suppress the expression of APP, a major cause of degenerative neurological diseases such as Alzheimer's disease, while increasing the expression of ADAM10 and ADAM17, which cleave pre-formed APP on the cell membrane and release it in the form of sAPPa, and IDE, which degrades pre-formed senile plaques. Furthermore, the peptides can alleviate neuroinflammatory responses, one of the main causes of degenerative neurological diseases, and regenerate damaged nervous systems.

[0025] According to one embodiment, the peptides (peptide-9 to peptide-12) can suppress the expression of APP, a major cause of degenerative neurological diseases such as Alzheimer's disease, while increasing the expression of ADAM17, which cleaves pre-formed APP on the cell membrane surface and releases it in the form of sAPPa. Furthermore, the peptides can alleviate neuroinflammatory responses, one of the main causes of degenerative neurological diseases, and regenerate damaged nervous systems.

[0026] Therefore, the peptide can be used as an active ingredient in a pharmaceutical composition for preventing or treating degenerative neurological diseases.

[0027] The pharmaceutical composition may comprise, but is not limited to, a pharmaceutically effective amount of the peptide; and / or a pharmaceutically acceptable carrier.

[0028] The term "pharmaceutically effective amount" as used herein means an amount sufficient to achieve the efficacy of the pharmaceutical composition in preventing or treating a degenerative neurological disease.

[0029] The pharmaceutically acceptable carriers are those commonly used in formulations, and include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

[0030] The pharmaceutical composition may further contain, in addition to the above ingredients, lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives, etc., but is not limited thereto.

[0031] The pharmaceutical composition can be administered orally or parenterally, preferably parenterally. In the case of parenteral administration, it can be administered by intramuscular injection, intravenous injection, subcutaneous injection, intraperitoneal injection, topical administration, transdermal administration, etc., but is not limited thereto.

[0032] The dosage of the pharmaceutical composition may be, but is not limited to, 0.0001 to 1000 μg (micrograms), 0.001 to 1000 μg, 0.01 to 1000 μg, 0.1 to 1000 μg, or 1.0 to 1000 μg per day, and may be variously prescribed depending on factors such as formulation method, administration method, age, weight, sex, pathological condition, diet, administration time, administration route, excretion rate, and reaction sensitivity of the patient.

[0033] The pharmaceutical composition can be prepared in unit dose form or in a multi-dose container by formulating it with pharmaceutically acceptable carriers and / or excipients in a manner that can be easily carried out by a person skilled in the art to which the invention pertains.

[0034] The dosage form may be in the form of a solution, suspension or emulsion in an oily or aqueous medium, or in the form of an extract, powder, granules, tablet or capsule, and may further contain a dispersing agent and / or a stabilizing agent.

[0035] Yet another aspect provides a method for preventing or treating a degenerative neurological disease, comprising administering to an individual a pharmaceutical composition containing, as an active ingredient, a therapeutically effective amount of a peptide consisting of any one of the amino acid sequences of SEQ ID NO:1 to SEQ ID NO:8 and SEQ ID NO:20 to SEQ ID NO:23.

[0036] Among the terms or elements mentioned in the description of the peptide or pharmaceutical composition, the same as those already mentioned are as described above. [Effects of the Invention]

[0037] The peptide according to one embodiment can inhibit the expression of amyloid precursor protein and increase the degradation of the amyloid precursor protein and the expression of ADAM10, ADAM17 and IDE, which are related genes.

[0038] Furthermore, the peptide according to one embodiment can alleviate or ameliorate neuroinflammatory responses by increasing the production of anti-inflammatory cytokines, and can induce nerve cell regeneration such as neurite outgrowth.

[0039] Therefore, the peptide according to one embodiment can be used as an active ingredient in a pharmaceutical composition for preventing or treating degenerative neurological diseases. [Brief explanation of the drawings]

[0040] FIG. 1 shows the results of confirming the inhibition of amyloid precursor protein (APP) expression after adding a peptide according to one embodiment to a human neuronal cell line, where A in FIG. 1 shows the results of adding Peptide-1, B in FIG. 1 shows Peptide-4, C in FIG. 1 shows Peptide-5, D in FIG. 1 shows Peptide-7, E in FIG. 1 shows Peptide-8, and F in FIG. 1 shows the results of adding a comparison peptide.

[0041] Figure 2 shows the results of confirming increased ADAM10 expression after adding a peptide according to one embodiment to a human neuronal cell line, where A in Figure 2 shows the results after adding Peptide-3, B in Figure 2 shows the results after adding Peptide-4, C in Figure 2 shows the results after adding Peptide-5, D in Figure 2 shows the results after adding Peptide-7, and E in Figure 2 shows the results after adding Peptide-8.

[0042] Figure 3 shows the results of confirming increased ADAM17 expression after adding a peptide according to one embodiment to a human neuronal cell line, where Figure 3A shows the results after adding Peptide-1 and Figure 3B shows the results after adding Peptide-4.

[0043] Figure 4 shows the results of confirming increased expression of sAPPa and ADAM17 after adding a peptide according to one embodiment to a human neuronal cell line, where Figure 4A shows the results after adding Peptide-1, Figure 4B shows the results after adding Peptide-4, and Figure 4C shows the results after adding Peptide-7.

[0044] FIG. 5 shows the results of flow cytometric analysis of the distribution of APP on the membrane surface of nerve cells after addition of a peptide according to one embodiment.

[0045] Figure 6 shows the results of confirming increased IDE expression after adding a peptide according to one embodiment to a human neuronal cell line, where Figure 6A shows the results after adding Peptide-1, Figure 6B shows the results after adding Peptide-2, Figure 6C shows the results after adding Peptide-3, and Figure 6D shows the results after adding Peptide-4.

[0046] Figure 7 shows the results of confirming increased IDE expression after adding a peptide according to one embodiment to a human neuronal cell line, where Figure 7A shows the results after adding Peptide-5, Figure 7B shows the results after adding Peptide-6, Figure 7C shows the results after adding Peptide-7, and Figure 7D shows the results after adding Peptide-8.

[0047] FIG. 8 shows the results of confirming the increase in expression of anti-inflammatory cytokines by adding peptides according to one embodiment, where A in FIG. 8 shows the results when Peptide-1 was added, B in FIG. 8 shows the results when Peptide-3 was added, C in FIG. 8 shows the results when Peptide-4 was added, D in FIG. 8 shows the results when Peptide-5 was added, and E in FIG. 8 shows the results when Peptide-7 was added.

[0048] FIG. 9 shows the results of confirming the neurite outgrowth promoting effect after adding a peptide according to one embodiment to a human neuronal cell line, where FIG. 9A shows the results after adding Peptide-1, FIG. 9B shows the results after adding Peptide-2, and FIG. 9C shows the results after adding Peptide-6.

[0049] Figure 10 shows the results of confirming increased ADAM17 expression after adding a peptide according to one embodiment to a human neuronal cell line, where A in Figure 10 shows the results after adding Peptide-9, B in Figure 10 shows the results after adding Peptide-10, C in Figure 10 shows the results after adding Peptide-11, and D in Figure 10 shows the results after adding Peptide-12.

[0050] Figure 11 shows the results of confirming the inhibition of amyloid precursor protein (APP) expression after adding a peptide according to one embodiment to a human neuronal cell line, where Figure 11A shows the results of adding Peptide-11, Figure 11B shows the results of adding Peptide-12, and Figure 11C shows the results of adding a comparison peptide.

[0051] FIG. 12 shows the results of confirming the increase in expression of anti-inflammatory cytokines by adding a peptide according to one embodiment, where A in FIG. 12 shows the results when Peptide-9 was added, B in FIG. 12 shows the results when Peptide-11 was added, and C in FIG. 12 shows the results when Peptide-12 was added.

[0052] Figure 13 shows the results of confirming the increase in expression of brain-derived neurotrophic factor by adding a peptide according to one embodiment, where A in Figure 13 shows the results when Peptide-9 was added, B in Figure 13 shows the results when Peptide-10 was added, C in Figure 13 shows the results when Peptide-11 was added, and D in Figure 13 shows the results when Peptide-12 was added. DETAILED DESCRIPTION OF THE INVENTION

[0053] The present invention will be described in more detail below 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.

[0054] 1. Peptide-1 to Peptide-8

[0055] Example 1: Synthesis of peptides

[0056] Peptides having the amino acid sequences of SEQ ID NO:1 to SEQ ID NO:8 listed in Table 1 were synthesized using an automated peptide synthesizer (Milligen 9050, Millipore, USA). These synthesized peptides were purified and separated using C18 reverse-phase high-performance liquid chromatography (HPLC) (Waters Associates, USA). The column used was an ACQUITY UPLC BEH300 C18 (2.1 mm x 100 mm, 1.7 μm, Waters Co., USA).

[0057] [Table 1]

[0058] Example 2: Confirmation of the effect of inhibiting amyloid precursor protein expression

[0059] In this example, we aimed to confirm the inhibitory effect of peptide treatment on amyloid precursor protein (AP) expression. To this end, the prepared peptides were added at concentrations of 0.001, 0.01, 0.1, or 1 mM to the human neuronal cell line SH-SY5Y, isolated and expanded from the bone marrow of a human with neuroblastoma, and cultured for 24 hours. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the APP primers listed in Table 2 below to measure the expression level of APP. A control group was used without peptide treatment, and a comparative group was used with the addition of a peptide (NRP1: AFMVDNEAIYDICR (SEQ ID NO: 9)).

[0060] [Table 2]

[0061] As a result, as shown in Figure 1, the addition of a peptide according to one embodiment (Peptide-1, Peptide-4, Peptide-5, Peptide-7, or Peptide-8) inhibited the expression of APP, a protein that is a major cause of degenerative neurological diseases, including Alzheimer's disease. In particular, the inhibitory effect on APP expression was very significant, showing an 80-90% reduction in expression level compared to the control group.

[0062] Example 3: Confirmation of the effect of increasing the expression of genes related to the degradation of amyloid precursor protein

[0063] 3-1. Expression of ADAM10 (A Disintegrin And Metalloproteinase-containing protein 10) gene

[0064] ADAM10 is an α-secretase that cleaves APP into non-toxic fragments to inhibit the production of β-amyloid (Ab1-42, 1-40). Therefore, increased expression of ADAM10 can be used as an indicator of the therapeutic efficacy of degenerative neurological diseases. Specifically, the prepared peptide was added to the human neuronal cell line SH-SY5Y at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the ADAM10 primers listed in Table 3 below to measure the ADAM10 expression level. A control group was used without peptide treatment.

[0065] As a result, as shown in FIG. 2, the addition of a peptide according to one embodiment (Peptide-3, Peptide-4, Peptide-5, Peptide-7, or Peptide-8) increased the expression of ADAM10.

[0066] [Table 3]

[0067] 3-2. Expression of ADAM17 (A Disintegrin And Metalloproteinase-containing protein 17) gene

[0068] ADAM17 is an enzyme that cleaves APP into the non-toxic fragment sAPPa, removing it from the cell membrane. Therefore, increased expression of ADAM17 can be used as an indicator of the therapeutic efficacy of degenerative neurological diseases. Specifically, the prepared peptide was added to the SH-SY5Y neuronal cell line at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the ADAM17 primers listed in Table 4 below to measure the ADAM10 expression level. The prepared peptide was also added to the SH-SY5Y human neuronal cell line at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. The expression of ADAM17 protein and sAPPa, which is released by cleavage from APP by ADAM17 and ADAM10, was then confirmed by Western blotting using antibodies against each protein. Flow cytometric analysis was also used to analyze the distribution of APP on the neuronal membrane. A control group was used without the addition of peptide.

[0069] [Table 4]

[0070] As a result, as shown in Figures 3 and 4, the addition of peptides according to one embodiment (Peptide-1, Peptide-4, and Peptide-7) increased the expression of sAPPa and ADAM17, and as shown in Figure 5, it was found that this change in expression contributed to a decrease in the level of APP present on the surface of the neuronal membrane.

[0071] 3-3. Expression of IDE (Insulin degrading enzyme) gene

[0072] IDE is an enzyme that degrades senile plaques formed and aggregated by β-amyloid (Ab1-42, 1-40). Therefore, increased expression of IDE can be used as an indicator of the therapeutic efficacy of degenerative neurological diseases. Specifically, the prepared peptide was added to the human neuronal cell line SH-SY5Y at concentrations of 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the IDE primers listed in Table 5 below to measure the IDE expression level. A control group was used without peptide treatment.

[0073] [Table 5]

[0074] As a result, as shown in Figures 6 and 7, the addition of peptides according to one embodiment (Peptide-1, Peptide-2, Peptide-3, Peptide-4, Peptide-5, Peptide-6, Peptide-7, and Peptide-8) increased the expression of IDE.

[0075] Taking the above experimental results into consideration, the peptide according to one embodiment can prevent the onset of degenerative neurological diseases or improve associated symptoms by increasing the expression of ADAM10 and ADAM17, which cleave APP on the cell membrane surface and release it in the form of the non-toxic fragment sAPPa, and by reducing the expression of IDE, which degrades senile plaques in brain tissue caused by pathological factors.

[0076] Example 4: Confirmation of the effect of increasing the expression of anti-inflammatory cytokines

[0077] In this example, we aimed to confirm the inhibitory effect of peptide treatment on neuroinflammatory responses, one of the main causes of degenerative neurological diseases, by examining the effect of increasing the expression of IL-10, an anti-inflammatory cytokine. Specifically, the prepared peptide was added to the human neuronal cell line SH-SY5Y at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was then extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the IL-10 primers listed in Table 6 below to measure IL-10 expression levels. A control group was used without peptide treatment.

[0078] [Table 6]

[0079] As a result, as shown in FIG. 8, the addition of a peptide according to one embodiment (Peptide-1, Peptide-3, Peptide-4, Peptide-5, or Peptide-7) increased the expression of IL-10.

[0080] Example 5: Confirmation of neurite outgrowth promoting effect

[0081] In this example, we investigated the growth of neurites to confirm the regeneration of neurons by peptide treatment. Specifically, the prepared peptides were added to the human neuronal cell line SH-SY5Y at concentrations of 0.01, 0.1, or 1 mM and cultured for 24 hours. The grown neurites were then observed under a microscope and their lengths were measured and evaluated.

[0082] As a result, as shown in FIG. 9, the addition of a peptide according to one embodiment (Peptide-1, Peptide-2, or Peptide-6) increased the length of neurites.

[0083] From the above results, it was found that the peptide according to one embodiment can contribute to inhibiting the occurrence and progression of degenerative neurological diseases, including Alzheimer's disease, Parkinson's disease, Huntington's disease, multiple sclerosis, etc.

[0084] 2. Peptide-9 to Peptide-12

[0085] Example 1: Synthesis of peptides

[0086] Peptides having the amino acid sequences of SEQ ID NOs: 20 to 23 listed in Table 7 were synthesized using an automated peptide synthesizer (Milligen 9050, Millipore, USA). These synthesized peptides were purified and separated using C18 reverse-phase high-performance liquid chromatography (HPLC) (Waters Associates, USA). The column used was an ACQUITY UPLC BEH300 C18 (2.1 mm x 100 mm, 1.7 μm, Waters Co., USA).

[0087] [Table 7]

[0088] Example 2: Confirmation of the effect of increasing the expression of genes related to the degradation of amyloid precursor protein

[0089] The prepared peptides were added to the SH-SY5Y neuronal cell line at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the ADAM17 primers listed in Table 4 to measure the ADAM17 expression level. A control group was used without peptide addition.

[0090] As a result, as shown in FIG. 10, the addition of a peptide according to one embodiment (Peptide-9, Peptide-10, Peptide-11, or Peptide-12) increased the expression of ADAM17, and this tendency showed a concentration-dependent tendency.

[0091] Taking the above experimental results into consideration, the peptide according to one embodiment can prevent the onset of degenerative neurological diseases or ameliorate associated symptoms by increasing the expression of ADAM17, which cleaves APP on the cell membrane surface and releases it in the form of the non-toxic fragment sAPPa.

[0092] Example 3: Confirmation of the effect of inhibiting amyloid precursor protein expression

[0093] In this example, we aimed to confirm the inhibitory effect of peptide treatment on amyloid precursor protein (AP) expression. To this end, the prepared peptides were added at concentrations of 0.001, 0.01, 0.1, or 1 mM to the human neuronal cell line SH-SY5Y, isolated and expanded from the bone marrow of a human with neuroblastoma, and cultured for 24 hours. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the APP primers listed in Table 2 to measure the expression level of APP. A control group was used without peptide administration, and a comparative group was used with the administration of a peptide (NRP1: AFMVDNEAIYDICR (SEQ ID NO: 9)).

[0094] As a result, as shown in Figure 11, the addition of a peptide according to one embodiment (Peptide-11 or Peptide-12) inhibited the expression of APP, a protein that is a major cause of degenerative neurological diseases including Alzheimer's disease. In particular, the inhibitory effect on APP expression was very significant, showing an 80-90% lower expression level compared to the control group.

[0095] Example 4: Confirmation of the effect of increasing the expression of anti-inflammatory cytokines

[0096] In this example, we aimed to confirm the inhibitory effect of peptide treatment on neuroinflammatory responses, one of the main causes of degenerative neurological diseases, by examining the effect of increasing the expression of IL-10, an anti-inflammatory cytokine. Specifically, the peptides prepared above were added to the human neuronal cell line SH-SY5Y at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was then extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the IL-10 primers listed in Table 6 to measure IL-10 expression levels. A control group was used without peptide treatment.

[0097] As a result, as shown in Figure 12, the addition of a peptide according to one embodiment (Peptide-9, Peptide-11, or Peptide-12) increased IL-10 expression. In particular, this anti-inflammatory effect was more pronounced in the group to which Peptide-9 was added.

[0098] Considering the above experimental results, the peptide according to one embodiment can inhibit the degradation of amyloid precursor protein and at the same time induce an anti-inflammatory response at the lesion site.

[0099] Example 5. Confirmation of the effect of increasing the expression of brain-derived neurotrophic factor

[0100] In this example, we aimed to confirm the therapeutic or ameliorative effects of peptide treatment on degenerative neurological disorders through nervous system regeneration by examining the upregulation of brain-derived neurotrophic factor (BDNF) expression. Specifically, the prepared peptides were added to the human neuronal cell line SH-SY5Y at concentrations of 0.001, 0.01, 0.1, or 1 mM and cultured for 24 hours at 37°C and 5% CO2. Total RNA was extracted from the culture, cDNA was synthesized, and real-time qPCR was performed using the BDNF primers listed in Table 8 below to measure BDNF expression levels. A control group was used without peptide treatment.

[0101] [Table 8]

[0102] As a result, as shown in Figure 13, the addition of a peptide according to one embodiment (Peptide-9, Peptide-10, Peptide-11, or Peptide-12) increased BDNF expression. In particular, this increased expression effect was more pronounced in the group to which Peptide-11 or Peptide-12 was added.

[0103] From the above results, it was found that the peptide according to one embodiment can contribute to inhibiting the occurrence and progression of degenerative neurological diseases, including Alzheimer's disease, Parkinson's disease, Huntington's disease, multiple sclerosis, etc.

[0104] The above description of the present invention is for illustrative purposes only, and those skilled in the art will understand that the present invention can be easily modified into other specific forms without changing the technical spirit or essential features of the present invention. Therefore, the above-described embodiments should be understood to be illustrative in all respects and not limiting.

Claims

1. A peptide consisting of any one of the amino acid sequences of SEQ ID NO:1 to SEQ ID NO:6, SEQ ID NO:8, and SEQ ID NO:20 to SEQ ID NO:

23.

2. 2. The peptide according to claim 1, wherein the N-terminus of the peptide is bound to any one protecting group selected from the group consisting of an acetyl group, a fluorenylmethoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, and polyethylene glycol (PEG).

3. The C-terminus of the peptide is an amino group (-NH ２ ), and azide (-NHNH ２ 2. The peptide of claim 1, wherein the peptide is bonded to any one of the protecting groups selected from the group consisting of:

4. The peptide of claim 1, wherein the peptide exhibits one or more of the following properties: (a) inhibition of amyloid precursor protein (APP) expression; (b) increased expression of ADAM10 (A Disintegrin And Metalloproteinase-containing protein 10) or ADAM17 (A Disintegrin And Metalloproteinase-containing protein 17) protein; (c) increased expression of IDE (insulin degrading enzyme) protein; (d) increased expression of anti-inflammatory cytokines; (e) promoting neurite outgrowth; and (f) Increased expression of brain-derived neurotrophic factor (BDNF).

5. A pharmaceutical composition for preventing or treating a degenerative neurological disease, comprising the peptide according to any one of claims 1 to 4 as an active ingredient, The pharmaceutical composition, wherein the degenerative neurological disease is selected from the group consisting of Alzheimer's disease, Parkinson's disease, and Lewy Body Dementia.

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