Application of a humanized DKK-1 monoclonal antibody in the preparation of a drug for treating Alzheimer's disease

Abstract

The present invention belongs to the field of biomedicine. Specifically, the present invention provides an application of a humanized DKK-1 monoclonal antibody in the preparation of a drug for treating Alzheimer's disease. The light and heavy chain sequences of the humanized DKK-1 monoclonal antibody are shown in SEQ ID NO: 1-2, respectively. Animal experiments have confirmed that the humanized DKK-1 monoclonal antibody can prolong the survival of Alzheimer's mice, improve the cognitive level of Alzheimer's mice, improve the memory ability of Alzheimer's mice, and reduce blood sugar, indicating that the humanized DKK-1 monoclonal antibody has the effect of treating Alzheimer's disease and can alleviate the symptoms of senile dementia.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and specifically relates to the use of a humanized DKK-1 monoclonal antibody in the preparation of a drug for treating Alzheimer's disease. Background Art

[0002] At present, the reported DKK-1 antibodies mainly include: DKN-01, BHQ-880, DKK1-A2, RH2-18, and Hetero-DS.

[0003] Among them, DKN-01 is a humanized anti-DKK-1 IgG4 monoclonal antibody developed by LeapTherapeutics, which is used to treat a variety of tumors, such as bile duct cancer, endometrial cancer, gastric cancer, esophageal cancer, liver cancer, ovarian cancer, and prostate cancer, and is in clinical phase II. BHQ-880 can be used to treat multiple myeloma and osteosarcoma. DKK1-A2 recognizes the DKK1 P20 peptide and is used to treat blood and solid cancer cells. RH2-18 can treat postmenopausal osteoporosis. Hetero-DS can treat postmenopausal osteoporosis and fractures.

[0004] As can be seen from the above, the current effects of DKK-1 antibodies are mainly focused on treating fractures, osteoporosis and cancer, while there are few reports on related research on treating Alzheimer's disease, lowering blood sugar, and prolonging life. Summary of the invention

[0005] In order to solve the above problems, the present invention provides a use of a humanized DKK-1 monoclonal antibody in the preparation of a drug for treating Alzheimer's disease, and specifically provides the following technical solutions:

[0006] The first aspect of the present invention provides a use of a humanized DKK-1 monoclonal antibody in the preparation of a drug for treating Alzheimer's disease, wherein the light and heavy chains of the humanized DKK-1 monoclonal antibody are as shown in SEQ ID NO.1-2.

[0007] In one embodiment, the application includes:

[0008] (1) Prolong the survival of patients with Alzheimer's disease; and / or

[0009] (2) Improving cognitive function in patients with Alzheimer's disease; and / or

[0010] (3) Improving memory in patients with Alzheimer's disease; and / or

[0011] (4) Lower blood sugar levels in patients with Alzheimer’s disease.

[0012] In one embodiment, the medicament is a vaccine.

[0013] The second aspect of the present invention provides a drug for treating Alzheimer's disease, wherein the active ingredient of the drug is the aforementioned humanized DKK-1 monoclonal antibody.

[0014] In one embodiment, the drug further comprises other pharmaceutical excipients.

[0015] In a preferred embodiment, the drug is a vaccine. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The survival curve of mice after DKK-1 monoclonal antibody treatment is shown;

[0017] Figure 2 shows the experimental data of new object recognition. Figure 2 A is the time it takes for mice to explore new things. Figure 2 B is the frequency of mice exploring new things;

[0018] Figure 3 shows the data of the Y-maze experiment. Figure 3 A is the time the mouse spends exploring the new arm, Figure 3 B is the frequency of mice exploring the new arm;

[0019] Figure 4 shows the random blood glucose values ​​of mice;

[0020] Figure 5 shows the expression levels of Alzheimer's disease marker molecules, Figure 5 A is the ELISA test to detect the level of β-amyloid 42 (Aβ42) in serum; Figure 5 B is the immunohistochemical experiment to detect the expression level of β-amyloid 40 (Aβ40) in brain tissue. DETAILED DESCRIPTION

[0021] Example 1 Preparation of DKK-1 monoclonal antibody

[0022] The preparation of humanized DKK-1 monoclonal antibody and the verification of biological activity are described in the prior patent 202410979827.5 (application number) and are incorporated herein. Specifically, the amino acid sequences of the light and heavy chains of the DKK-1 monoclonal antibody used in this example are as follows:

[0023] Light chain sequence (SEQ ID NO.1):

[0024] DIQMTQSPSSSLSASVGDRVTITCRASQGISNYLNWYQQKPGKAPKLLISYTSSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCLQYSKLPLTFGQGTKVEIK RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0025] Heavy chain sequence (SEQ ID NO.2):

[0026] EVQLVESGGGLVQPGGSLRLSCAASGFTFSNYAMSWVRQAPGKGLEWVSSISTGGTTYYPDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCVSEIYWPMDFWGQGT LVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPP CPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT ISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG

[0027] Example 2 Verification of the effect of DKK-1 monoclonal antibody in the treatment of Alzheimer's disease

[0028] Alzheimer's disease model animals: app-pSl-tau gene mice (Beijing Huafukang Biotechnology Co., Ltd.), male and female, 16, 4 months old. The mice were divided into two groups: control group: IgG antibody (Control, purchased from Dakoway Company, catalog number 400197), experimental group: DKK-1 monoclonal antibody (anti-DKK-1), 8 mice in each group.

[0029] Antibody regimen: intraperitoneal injection, 25 mg / kg / time, twice a week, for 3 months.

[0030] Experimental steps:

[0031] According to the above injection scheme, the mice were observed every day and the death of mice was recorded.

[0032] Following the above injection protocol, behavioral testing began 2 months after antibody intervention:

[0033] 2.1 Novel Object Recognition Test: Rodents will spend more time (A) and frequency (B) exploring new objects than objects they are already familiar with. In this task, the test animals are exposed to an object during an initial familiarization phase. In the subsequent test phase, the animals will be exposed to the same familiar object and a new object at the same time. It is expected that the animals will remember the initial object and spend more time exploring the new object, assessing the animals' memory and cognitive abilities, especially in terms of object recognition and memory cognitive functions.

[0034] 2.2 Y-maze experiment: The Y-maze is a commonly used tool for animal behavior research and can be effectively used to study the spatial learning and memory ability of animals. The Y-maze consists of three arms of equal length, and the arrangement of these three arms is similar to the letter "Y", hence the name Y-maze. The design of the Y-maze allows animals to explore new arms for a certain period of time and frequency, which can also reflect their spatial learning and memory abilities.

[0035] 2.3 Blood sugar: Blood was collected from the rat tail to test random blood sugar (after collecting blood from the rat tail, drop the blood onto the test strip of a blood sugar tester (Roche) and record the blood sugar value).

[0036] 2.4 Changes in the expression of Aβ40 and Aβ42 amyloid proteins: The expression levels of Aβ40 and Aβ42 are closely related to Alzheimer's disease and are often used as marker molecules for Alzheimer's disease. Abnormal accumulation of Aβ40 and Aβ42 may form plaques in the brain, leading to neuronal damage and cognitive decline. We perfused mice, fixed and precipitated brain tissue, and then sectioned and immunohistochemically stained it.

[0037] Experimental results:

[0038] The survival curve shows that DKK-1 monoclonal antibody treatment can effectively improve the survival rate of Alzheimer's mice ( Figure 1 In the new object recognition experiment, mice in the DKK-1 monoclonal antibody group explored the new objects more fully, indicating that DKK-1 monoclonal antibody can improve the cognitive and memory abilities of Alzheimer's mice ( Figure 2 In the Y-maze experiment, the learning and memory abilities of mice in the DKK-1 monoclonal antibody group were improved, indicating that DKK-1 monoclonal antibody can improve the cognitive and memory abilities of Alzheimer's disease mice ( Figure 3In the blood sugar experiment, the blood sugar of mice in the DKK-1 monoclonal antibody group was low, indicating that DKK-1 has a blood sugar-lowering effect ( Figure 4 In the study of changes in the expression of Aβ40 and Aβ42 amyloid proteins, ELISA experiments detected a decrease in the level of Aβ42 in the serum of mice in the DKK-1 monoclonal antibody group; immunohistochemical staining experiments showed that the expression of Aβ40 in mice in the DKK-1 monoclonal antibody group was downregulated ( Figure 5 ), the above results show that DKK-1 monoclonal antibody can alleviate Alzheimer's disease, lower blood sugar and significantly prolong the survival of Alzheimer's patients.

[0039] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. Use of a humanized DKK-1 monoclonal antibody in the preparation of a drug for treating Alzheimer's disease, characterized in that: The light and heavy chains of the humanized DKK-1 monoclonal antibody are shown in SEQ ID NO.1-2, respectively.

2. The use according to claim 1, characterized in that The applications include: (1) Prolong the survival of patients with Alzheimer's disease; and / or (2) Improving cognitive function in patients with Alzheimer's disease; and / or (3) Improving memory in patients with Alzheimer's disease; and / or (4) Lower blood sugar levels in patients with Alzheimer’s disease.

3. A drug for treating Alzheimer's disease, characterized in that: The active ingredient of the drug is a humanized DKK-1 monoclonal antibody, and the light and heavy chains of the antibody are shown in SEQ ID NO.1-2 respectively.

4. The drug according to claim 3, characterized in that The medicine also includes other pharmaceutical excipients.

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