CN113336837B

CN113336837B - Polypeptide and application thereof in preparing anti-neurodegenerative disease medicine

Info

Publication number: CN113336837B
Application number: CN202110827432.XA
Authority: CN
Inventors: 高向东; 陈松; 杨倩华; 周岳茜; 姚文兵
Original assignee: China Pharmaceutical University
Current assignee: China Pharmaceutical University
Priority date: 2021-07-21
Filing date: 2021-07-21
Publication date: 2022-12-23
Anticipated expiration: 2041-07-21
Also published as: CN115925811A; CN116444643A; CN113336837A

Abstract

The present invention relates to polypeptides and uses thereof. The polypeptide is a series of polypeptides, and the sequence is one of SEQ ID NO 1 to SEQ ID NO 14. The polypeptide is used for preparing a medicament for treating or preventing neurodegenerative diseases. The series of polypeptides of the invention can treat or prevent neurodegenerative diseases and have good application prospect.

Description

Polypeptide and application thereof in preparing anti-neurodegenerative disease medicine

Technical Field

The invention relates to polypeptides and application thereof in preparing medicines for resisting neurodegenerative diseases, wherein the polypeptides are specific polypeptide fragments of DVL-2, and belong to the technical field of biological medicines.

Background

Neurodegenerative diseases (such as Alzheimer's disease, parkinson's disease, etc.) are characterized mainly by specific neuronal loss and are a progressive group of diseases. Neurodegenerative diseases including Alzheimer's disease, parkinson's disease and the like mostly have complex mechanisms, specific causes are not yet determined, and effective curing means is lacked. There is a great need for research and development of methods and drugs effective in preventing and treating neurodegenerative diseases.

Parkinson's disease is one of the most common neurodegenerative diseases, and is most frequent in the elderly population. The most important pathological changes of the Parkinson disease comprise abnormal degeneration of mesencephalic dopaminergic neurons, obvious reduction of striatal dopamine and formation of lewy bodies caused by the abnormal degeneration, and clinically manifested by dyskinesia such as resting tremor, bradykinesia, muscular rigidity, abnormal gait in postures and the like, and even non-motor symptoms such as cognitive dysfunction and the like. The Parkinson disease patients in China account for about half of the world high-incidence Parkinson disease countries, and the social burden caused by the Parkinson disease is increasingly increased along with the continuous acceleration of the aging of the population.

Alzheimer's disease is the most common neurodegenerative disease, and is clinically manifested as cognitive function and learning disorders, memory decline, and the like. With the increasing aging of population, the number of patients suffering from alzheimer disease is increasing sharply in recent years, and the social burden caused by the aging is increasing, especially in china, the number of patients suffering from alzheimer disease is the first to live in the world, and the market demand of the medicine for preventing and treating alzheimer disease is huge.

Dishevelled (DVL) is an important regulator in the canonical Wnt signaling pathway. DVL in mammals consists primarily of DVL-1, DVL-2 and DVL-3, all of which contain three basic domains, the highly conserved DIX, PDZ and DEP domains. The inventors of the present invention have conducted extensive research, experiments and research efforts, and have now achieved research results relating to DVL-2.

Disclosure of Invention

The main purposes of the invention are: according to the recent research results of the inventor, aiming at the problems in the prior art, a polypeptide is provided, and can be applied to the preparation of the anti-neurodegenerative disease medicine. Also provides the application of the polypeptide.

The technical scheme for solving the technical problems of the invention is as follows:

a polypeptide, characterized in that the amino acid sequence of the polypeptide is one of SEQ ID NO 1 to SEQ ID NO 14.

Preferably, the amino acid sequence of the polypeptide is one of SEQ ID NO. 2, SEQ ID NO. 5, SEQ ID NO. 7 and SEQ ID NO. 8.

The series of polypeptides are specific polypeptide fragments of DVL-2 protein, and can be applied to preparation of medicines for resisting neurodegenerative diseases.

A polypeptide comprising a polypeptide as described above linked to a cell penetrating peptide, wherein the cell penetrating peptide is located at the N-terminus of the polypeptide; the amino acid sequence of the cell penetrating peptide is one of SEQ ID NO. 15 to SEQ ID NO. 47.

A polypeptide, which is characterized in that the polypeptide is formed by connecting the polypeptide with a cell penetrating peptide through a connecting peptide, wherein the connecting peptide and the cell penetrating peptide are positioned at the N end of the polypeptide; the amino acid sequence of the cell penetrating peptide is one of SEQ ID NO. 15 to SEQ ID NO. 47, and the amino acid sequence of the connecting peptide is one of SEQ ID NO. 48 to SEQ ID NO. 72.

Use of a polypeptide as hereinbefore described for the manufacture of a medicament for the treatment or prevention of a neurodegenerative disease.

Wherein the neurodegenerative disease comprises Alzheimer's disease or Parkinson's disease.

The invention also proposes:

a nucleic acid encoding a polypeptide as hereinbefore described.

A pharmaceutical composition comprising a polypeptide as hereinbefore described or a nucleic acid as hereinbefore described.

Use of a pharmaceutical composition as described hereinbefore for the preparation of a pharmaceutical formulation for the treatment or prevention of a neurodegenerative disease.

Through a large amount of exploration, experiments and research work, the inventor of the invention carries out structure-activity relationship analysis based on DVL-2, designs a plurality of polypeptide sequences and carries out activity detection, and finally obtains a series of polypeptides with biological activity for preventing and treating neurodegenerative diseases. The series of polypeptides related by the invention can treat or prevent neurodegenerative diseases, and has good application prospects.

Drawings

FIG. 1 is a graph showing the results of example 1 of the present invention, wherein the ordinate of each of graphs A and B shows the relative cell viability. In the graph A, the Control group is a Control group, the Rotenone group is a model group, and the Rotenone + peptide (2, 5,7, 8) group is a polypeptide group to be detected (the polypeptide to be detected is polypeptide SEQ ID NO:2, polypeptide SEQ ID NO:5, polypeptide SEQ ID NO:7, polypeptide SEQ ID NO:8, respectively). In the diagram B, the Control group is a Control group, the Rotenone group is a model group, the other experimental groups are polypeptide groups to be detected, and the polypeptides to be detected in the Rotenone + Tat group are cell penetrating peptides SEQ ID NO. 15, and the polypeptides to be detected in the Rotenone + Tat-peptide group (2, 5,7, 8) are second group polypeptides containing polypeptides SEQ ID NO. 2, SEQ ID NO. 5, SEQ ID NO. 7 or SEQ ID NO. 8, respectively.

FIG. 2 is a graph showing the results of example 2 of the present invention, in which Hoechst marks the nucleus and appears blue; PI marks apoptotic or necrotic cells, appearing red. In the figure, the Control group is a Control group, the Rotenone group is a model group, and the Rotenone + peptide2 group is a polypeptide group to be detected (the polypeptide to be detected is the polypeptide SEQ ID NO: 2).

FIG. 3 is a graph showing the results of example 3 of the present invention. In the figure, the Rotenone (-) Peptide2 (-) group was a control group, the Rotenone (+) Peptide2 (-) group was a model group, and the Rotenone (+) Peptide2 (+) group was a test polypeptide group (the test polypeptide was the polypeptide SEQ ID NO: 2).

FIG. 4 is a graph showing the results of example 4 of the present invention, in which the ordinate represents the relative cell survival rate, control group represents the Control group, and A β represents _25-35 Group as model group, abeta _25-35 The + peptide (2, 5,7, 8) group is a polypeptide group to be detected (the polypeptide to be detected is polypeptide SEQ ID NO:2, polypeptide SEQ ID NO:5, polypeptide SEQ ID NO:7, polypeptide SEQ ID NO:8, respectively).

FIG. 5 is a schematic diagram showing the results of example 5 of the present invention, in which Hoechst marks the nucleus and appears blue; PI marks apoptotic or necrotic cells, appearing red. In the figure, control group is a Control group, abeta _25-35 Group as model group, abeta _25-35 The + peptide2 group is a polypeptide group to be detected (the polypeptide to be detected is the polypeptide SEQ ID NO: 2).

FIG. 6 is a graph showing the results of example 6 of the present invention. In the figure, the Control group is a Control group, the MPTP group is a model group, the other experimental groups are polypeptide groups to be detected, the polypeptide to be detected in the MPTP + Tat group is a cell penetrating peptide SEQ ID NO. 15, and the polypeptide to be detected in the MPTP + Tat-peptide (2, 5,7, 8) group is 4 polypeptides containing a polypeptide SEQ ID NO. 2, a polypeptide SEQ ID NO. 5, a polypeptide SEQ ID NO. 7 or a polypeptide SEQ ID NO. 8 respectively.

FIG. 7 is a graph showing the results of example 7 of the present invention. Wherein, the A picture is the detection of TH expression level in substantia nigra of each group of experimental mice, and the B picture is the detection of TH expression level in striatum of each group of experimental mice. In the figure, con group is a control group, MPTP-Non group is a model group, the other experimental groups are to-be-detected polypeptide groups, the to-be-detected polypeptides in MPTP + Tat group are cell penetrating peptides SEQ ID NO:15, and the to-be-detected polypeptides in MPTP + Tat-peptide (2, 5,7, 8) group are 4 polypeptides containing polypeptide SEQ ID NO:2, polypeptide SEQ ID NO:5, polypeptide SEQ ID NO:7 or polypeptide SEQ ID NO:8 respectively.

Detailed Description

The present invention will be described in further detail with reference to the drawings in conjunction with embodiments. The invention is not limited to the examples given.

Example 1

This example is to examine the effect of each polypeptide on neuronal cell death in a parkinsonian-like model.

The test polypeptides of this example are as follows:

(1) A first group of polypeptides: polypeptide SEQ ID NO. 2, polypeptide SEQ ID NO. 5, polypeptide SEQ ID NO. 7, polypeptide SEQ ID NO. 8.

(2) A second group of polypeptides: (1) polypeptide SEQ ID NO:2+ linker peptide SEQ ID NO:48+ cell penetrating peptide SEQ ID NO:15; (2) polypeptide SEQ ID NO 5+ linker peptide SEQ ID NO 48+ cell penetrating peptide SEQ ID NO 15; (3) polypeptide SEQ ID NO 7+ linker peptide SEQ ID NO 57+ cell penetrating peptide SEQ ID NO 15; (4) polypeptide SEQ ID NO 8+ linker peptide SEQ ID NO 48+ cell penetrating peptide SEQ ID NO 15. Note: cell penetrating peptide SEQ ID NO 15 is cell penetrating peptide Tat.

The specific experimental procedure of this example is as follows:

test polypeptide groups: after 6 days of 10. Mu.M retinoic acid-induced differentiation of SH-SY5Y cells, post-differentiation SH-SY5Y cells in logarithmic growth phase were selected at 3X 10 ⁴ The cells were seeded in 96-well plates (100. Mu.L per well) at a density of one/mL, and the CO was 5% at 37 ℃ ₂ Culturing in a cell culture box. Rotenone (0.1. Mu.M) was added after 24h of culture (note: rotenone, rotenone), and the polypeptide to be tested (40. Mu.M) was added, and 6 wells were set. After 24h of culture, 5mg/mL MTT solution with 10% of culture medium volume is added into each hole, the incubation is carried out for 4h in a constant-temperature cell culture box at 37 ℃, the supernatant is discarded, 150 microliter DMSO is added into each hole, and the shaking is carried out for 10min at 500r/min on an enzyme label plate oscillator, so that the blue-violet crystal formazan is completely dissolved. After the oscillation is finished, 570nm is taken as a detection wavelength and 630nm is taken as a reference wavelength in an enzyme labeling instrument, the OD value of each hole is measured, and the survival condition of the cells is calculated.

Compared with the polypeptide group to be detected, the comparison group does not contain Rotenone and the polypeptide to be detected, and the model group does not contain the polypeptide to be detected.

As shown in fig. 1, the first and second groups of polypeptides both alleviated parkinsonism-like model neuronal cell death compared to the model group, whereas Tat, the cell-penetrating peptide alone, showed no significant cytoprotective effect.

Example 2

This example is to examine the effect of polypeptides on apoptosis in parkinsonism-like model nerve cells.

The polypeptide to be tested in this example is the polypeptide SEQ ID NO 2.

The specific experimental procedure of this example is as follows:

the polypeptide groups to be tested: after SH-SY5Y cells are induced to differentiate for 6 days by 10 mu M retinoic acid, differentiated SH-SY5Y cells in the logarithmic growth phase are selected to be 5 multiplied by 10 ⁴ cell/mL density was inoculated in a 6-well plate (1000. Mu.L per well), and CO was 5% ₂ Culturing in a cell culture box. Rotenone (0.1. Mu.M) was added after 24h of culture, and the polypeptide to be tested (40. Mu.M) was added, 3 replicate wells were set. After culturing for 24h, removing the culture medium in the 6-well plate, and washing the cells for 3 times with PBS buffer solution, 3min each time; add 1000. Mu.L of medium containing Hoechst 33342 (7 mM) per well and incubate for 25min in an incubator at 37 ℃. Cells were washed 2 times with PBS buffer, 1000. Mu.L of PI (2 mM) containing medium was added to each well and incubated for 15min at room temperature in the absence of light. After the staining was completed, the cells were washed 3 times with PBS buffer, and after the washing was completed, photographs were taken by observation under an inverted fluorescence microscope.

As shown in FIG. 2, hoechst marks the nucleus of the cell and appears blue; PI marks apoptotic or necrotic cells, appearing red. Compared with a model group, the polypeptide SEQ ID NO 2 can effectively reduce the apoptosis of Parkinson disease-like model nerve cells.

Example 3

This example is to examine the effect of polypeptides on pathological abnormalities of neural cells of the Parkinson's disease-like model.

The polypeptide to be detected in this example is the polypeptide SEQ ID NO 2.

The specific experimental procedure of this example is as follows:

the polypeptide groups to be tested: after 6 days of 10. Mu.M retinoic acid-induced differentiation of SH-SY5Y cells, post-differentiation SH-SY5Y cells in logarithmic growth phase were selected at 5X 10 ⁴ Per mL density was inoculated into 6-well plates (1000. Mu.L per well), and the content of CO was 5% ₂ Culturing in a cell culture box. Rotenone (0.1. Mu.M) was added after 24h of culture, and the polypeptide to be tested (40. Mu.M) was added, 3 duplicate wells were set. After 24h of culture, extracting cell protein and preparing samples, wherein the specific process is as follows: the medium in the 6-well plate was discarded, the cells were washed 3 times with PBS buffer, collected in an EP tube, and 300 μ L of RIPA lysate (a mixture of protease inhibitor and phosphatase inhibitor was added at a ratio of 1. Centrifuging at 12000r/min at 4 deg.C for 20min, collecting supernatant, determining protein concentration by BCA protein concentration determination kit and adjusting protein concentration to the same level by PBS buffer solution, adding 5 xSDS loading buffer solution with 1/4 total volume into protein solution, mixing well, heating in boiling water for 10min, and storing in-20 deg.C refrigerator. Protein samples were analyzed by western blotting, which was performed as follows: preparing SDS-PAGE gel, placing the SDS-PAGE gel in an electrophoresis tank, adding electrophoresis buffer solution, adding a protein sample into the sample loading hole, and performing constant-pressure electrophoresis. And stopping electrophoresis when the bromophenol blue band is transferred to the bottom of the gel plate, transferring the target protein band to a PVDF membrane by using a semi-dry transfer membrane instrument, sealing for 2h by using 3% skimmed milk powder, incubating overnight at 4 ℃ by using an anti-TH antibody, incubating by using a corresponding horseradish peroxidase-coupled secondary antibody, and finally exposing the band by using a chemiluminescent reagent.

Compared with the polypeptide group to be detected, the reference group does not contain Rotenone and the polypeptide to be detected, and the model group does not contain the polypeptide to be detected.

The results are shown in FIG. 3, compared with the model group, the polypeptide SEQ ID NO. 2 can increase TH expression level of parkinsonism-like model nerve cells and relieve pathological abnormality of the parkinsonism-like model nerve cells.

Example 4

This example is to examine the effect of each polypeptide on neuronal cell death in an alzheimer-like model.

The polypeptides to be tested in this example were 4 polypeptides: polypeptide SEQ ID NO. 2, polypeptide SEQ ID NO. 5, polypeptide SEQ ID NO. 7, and polypeptide SEQ ID NO. 8.

The specific experimental procedure of this example is as follows:

the polypeptide groups to be tested: SH-SY5Y cells in logarithmic growth phase were selected at 3X 10 ⁴ cell/mL density was inoculated in a 96-well plate (100. Mu.L per well), and CO was 5% ₂ Culturing in a cell culture box. After 24h of culture, abeta is added _25-35 (0.05. Mu.M) (Note: A.beta. _25-35 I.e., the beta-amyloid fragment), and the polypeptide to be tested (40 μ M) was added, and 6 multiple wells were set. After 24h of culture, 5mg/mL MTT solution with 10% of culture medium volume is added into each hole, incubation is carried out for 4h in a constant-temperature cell culture box at 37 ℃, supernatant is discarded, 150 muL DMSO is added into each hole, and then the mixture is shaken for 10min at a speed of 500r/min on an enzyme label plate oscillator, so that the purple crystal formazan is completely dissolved. After the oscillation is finished, 570nm is taken as a detection wavelength and 630nm is taken as a reference wavelength in an enzyme labeling instrument, the OD value of each hole is measured, and the survival condition of the cells is calculated.

Compared with the polypeptide group to be detected, the control group does not add Abeta _25-35 And the polypeptide to be detected is not added in the model group.

The results are shown in fig. 4, and compared with the model group, 4 polypeptides can relieve the death of nerve cells in the Alzheimer disease-like model.

Example 5

This example is to examine the effect of polypeptides on apoptosis in neural cells in an Alzheimer's disease-like model.

The polypeptide to be tested in this example is the polypeptide SEQ ID NO 2.

The specific experimental procedure of this example is as follows:

the polypeptide groups to be tested: SH-SY5Y cells in logarithmic growth phase were selected at 5X 10 ⁴ The cells were seeded at a density of one/mL in 6-well plates (1000. Mu.L per well) and the CO was 5% at 37 ℃ ₂ Culturing in a cell culture box. After 24h of culture, abeta is added _25-35 (0.05. Mu.M), and the polypeptide to be tested (40. Mu.M) is added, and 3 multiple wells are set. After culturing for 24h, removing the culture medium in the 6-well plate, and washing the cells for 3 times with PBS buffer solution, 3min each time; add 1000. Mu.L of medium containing Hoechst 33342 (7 mM) per well and incubate for 25min in an incubator at 37 ℃. Cells were washed 2 times with PBS buffer, 1000. Mu.L of PI (2 mM) containing medium was added to each well and incubated for 15min at room temperature in the absence of light. After staining was complete, cells were washed 3 times with PBS buffer and after washing was complete, visualized by inverted fluorescenceAnd observing and photographing under a micro mirror.

As shown in FIG. 5, hoechst marks the nucleus and appears blue; PI marks apoptotic or necrotic cells, appearing red. Compared with a model group, the polypeptide SEQ ID NO. 2 can effectively reduce the apoptosis of nerve cells of an Alzheimer disease-like model.

Example 6

This example is to examine the effect of each polypeptide on parkinsonism-like behavioral abnormalities in model mice.

The polypeptides to be tested in this example were 4 polypeptides: (1) polypeptide SEQ ID NO 2+ linker peptide SEQ ID NO 48+ cell penetrating peptide SEQ ID NO 15; (2) polypeptide SEQ ID NO 5+ linker peptide SEQ ID NO 48+ cell penetrating peptide SEQ ID NO 15; (3) polypeptide SEQ ID NO 7+ linker peptide SEQ ID NO 57+ cell penetrating peptide SEQ ID NO 15; (4) polypeptide SEQ ID NO 8+ linker peptide SEQ ID NO 48+ cell penetrating peptide SEQ ID NO 15.

The specific experimental procedure of this example is as follows:

the male C57BL/6 mice with the age of 8 weeks are taken and raised in a raising room with the temperature of 22-24 ℃, the humidity of 50 percent and the daily illumination time of 12 hours, and the mice can freely obtain food and water. Experimental mice were randomly divided into 7 groups: a Control group, an MPTP + Tat-peptide 2 group, an MPTP + Tat-peptide5 group, an MPTP + Tat-peptide 7 group and an MPTP + Tat-peptide 8 group. The polypeptide solution to be tested (1. Mu.g/g, 1 time per day) was intraperitoneally injected on days 1-6, MPTP (20 mg/kg; 2 hours apart, 4 times in total) was intraperitoneally injected on day 2, and the same amount of physiological saline was injected to the control group. Open field experiments were performed on day 8, and after the completion of the behavioural experiments, mice were sacrificed for brain tissue collection.

The open field experiment device consists of an open field reaction box and an automatic data collecting and processing system, wherein the bottom side length of the open field reaction box of a mouse is 50cm, the height of the reaction box is 35cm, the inner wall of the reaction box is coated with light tone, the reaction box is divided into four congruent squares by partition plates, a camera is erected right above the reaction box by 1-2 meters, and the visual field of the camera can cover the inside of the whole open field. The experiment was performed in a quiet environment. Open field experiment supporting software at first, accomplish parameter setting according to the experiment requirement after, put the mouse gently in the bottom surface center of reaction tank, make a video recording and the collection of data simultaneously. The mouse freely explores the environment in the reaction box for 5min, and the activity of the mouse in the open field within 5min is observed and recorded.

The result is shown in figure 6, compared with a model group mouse, the 4 polypeptides can improve the autonomous activity and space exploration capacity of a Parkinson disease model mouse, reduce the Parkinson disease-like behavior abnormality of the model mouse, and the cell penetrating peptide Tat alone does not show a significant improvement effect.

Example 7

The embodiment is to detect the influence of each polypeptide on pathological abnormalities of substantia nigra and striatum tissues of a mouse model of the Parkinson's disease.

The specific experimental procedure of this example is as follows:

mice were housed and treated in the same manner as in example 6. After sacrifice, the mouse brain substantia nigra and striatum were separated on ice and placed in a weighed EP tube and RIPA tissue lysate was added at 10 μ L/mg (protease inhibitor, phosphatase inhibitor cocktail at 1. Homogenizing on ice with tissue homogenizer, placing on vortex instrument, shaking thoroughly for cracking once every 5min for 5 times, centrifuging at 4 deg.C at 12000r/min with high speed refrigerated centrifuge for 10min, and collecting supernatant. Measuring protein concentration with BCA protein concentration determination kit, adjusting protein concentration to the same level with PBS buffer, adding 1/4 total volume of 5 xSDS loading buffer into protein solution, mixing, heating in boiling water for 10min, and storing in-20 deg.C refrigerator. Protein samples were analyzed by western blotting in the same manner as in example 3.

The result is shown in figure 7, compared with the model group mice, 4 polypeptides can improve the expression quantity of TH in substantia nigra and striatum of the Parkinson disease model mice, relieve Parkinson disease-like pathological abnormality of the model mice, and the cell penetrating peptide Tat alone does not show a remarkable improvement effect.

In the present invention, the polypeptide may also be a sequence other than SEQ ID NO 2, SEQ ID NO 5, SEQ ID NO 7, SEQ ID NO 8 in the following table.

The cell penetrating peptide may also be a sequence other than SEQ ID NO:15 in the following table.

SEQ ID NO	Sequence of
		15	YGRKKRRQRRR
16	GRKKRRQRRR
		17	GRKKRRQRRRPPQ
18	CYGRKKRRQRRR
		19	RKKRRQRRR
20	RRRQRRKKRGY
		21	RRRR
22	RRRRRR
		23	RRRRRRR
24	RRRRRRRR
		25	RRRRRRRRR
26	RRRRRRRRRR
		27	RRRRRRRRRRRR
28	RRRRRRRRRRRRRRR
		29	RQIKIWFQNRRMKWKK
30	YARAAARQARA
		31	LSTAADMQGVVTDGMASGLDKDYLKPDD
32	KETWWETWWTEWSQPKKRKV
		33	CVKRGLKLRHVRPRVTRMDV
34	DAATATRGRSAASRPTERPRAPARSASRPRRPVD
		35	RGGRLSYSRRRFSTSTGR
36	RRIRPRPPRLPRPRPRPLPFPRPG
		37	PIEVCMYREP
38	VPTLK
		39	CHHHHHRRRRRRRRRHHHHHC
40	MVTVLFRRLRIRRACGPPRVRV
		41	Ac-GLWWRLWWRLRSWFRLWFRA-Cya
42	LLIILRRRIRKQAHAHSK
		43	CRWRWKCCKK
44	KWRRKLKKLRPKKKRKV
		45	MASIWVGHRG
46	RRRKIKR
		47	YGRKKRRQRRRLLRAALRKAAL

The linker peptide may also be a sequence other than SEQ ID NO 48, SEQ ID NO 57.

In addition to the above embodiments, the present invention may have other embodiments. All technical solutions formed by adopting equivalent substitutions or equivalent transformations fall within the protection scope of the claims of the present invention.

<110> university of Chinese pharmacy

<120> polypeptide and application thereof in preparation of anti-neurodegenerative disease drugs

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Claims

1. The polypeptide is characterized in that the amino acid sequence of the polypeptide is SEQ ID NO. 2.

2. A polypeptide comprising the polypeptide of claim 1 linked to a cell penetrating peptide, wherein the cell penetrating peptide is located at the N-terminus of the polypeptide; the amino acid sequence of the cell penetrating peptide is one of SEQ ID NO. 15 to SEQ ID NO. 47.

3. A polypeptide, which is formed by connecting the polypeptide of claim 1 with a cell-penetrating peptide through a connecting peptide, wherein the connecting peptide and the cell-penetrating peptide are both positioned at the N-terminal of the polypeptide; the amino acid sequence of the cell penetrating peptide is one of SEQ ID NO. 15 to SEQ ID NO. 47, and the amino acid sequence of the connecting peptide is one of SEQ ID NO. 48 to SEQ ID NO. 72.

4. Use of a polypeptide according to any one of claims 1 to 3 for the preparation of a medicament for the treatment or prevention of a neurodegenerative disease.

5. Use according to claim 4, characterized in that the neurodegenerative disease is Alzheimer's disease.

6. Use according to claim 4, characterized in that the neurodegenerative disease is Parkinson's disease.

7. A nucleic acid encoding the polypeptide of any one of claims 1 to 3.

8. A pharmaceutical composition comprising a polypeptide according to any one of claims 1 to 3 or a nucleic acid according to claim 7.

9. Use of the pharmaceutical composition of claim 8 for the preparation of a pharmaceutical formulation for the treatment or prevention of a neurodegenerative disease.