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Sclerostin and application thereof in preparation of related products for treating or preventing Alzheimer's disease

A technology for Alzheimer's disease and sclerostin, applied in the field of biomedicine, can solve problems such as degrading protease dysfunction, aggravating cognitive function, losing binding and stabilizing microtubules, etc., to inhibit the production of Aβ protein and accelerate The production of Aβ protein and the effect of alleviating Alzheimer's disease

Pending Publication Date: 2021-09-07
蒋青
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, current studies believe that in the preclinical stage of Alzheimer's disease, the clearance efficiency of AβO in the cerebrospinal fluid may decrease and the concentration of AβO in the cerebrospinal fluid may increase, which may be related to the dysfunction of these degradation proteases
In addition, not all AβO can be cleared, and some oligomers dissociated from Aβ fibrils may not be cleared and have some toxicity to adjacent synapses, promoting the hyperphosphorylation and aggregation of microtubule-associated Tau protein , so that it loses the role of binding and stabilizing microtubules, aggregates to form pairs of helical filaments, and aggravates the degeneration of cognitive function

Method used

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  • Sclerostin and application thereof in preparation of related products for treating or preventing Alzheimer's disease
  • Sclerostin and application thereof in preparation of related products for treating or preventing Alzheimer's disease
  • Sclerostin and application thereof in preparation of related products for treating or preventing Alzheimer's disease

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1 Sclerostin preparation

[0030] The amino acid sequence of mouse sclerostin (sclerostin) with chemical synthesis purity greater than 98% is shown in SEQNO.1:

[0031] MQPSLAPCLICLLVHAAFCAVEGQGWQAFRNDATEVIPGLGEYPEPPPENNQTMNRAENGGRPPHHPYDAKDVSEYSCRELHYTRFLTDGPCRSAKPVTELVCSGQCGPARLLPNAIGRVKWWRPNGPDFRCIPDRYRAQRVQLLCPGGAAPRSRKVRLVASCKCKRLLTFHNQSELKDFGPETARPQKGRANKPRAPGARGAK

[0032]The synthesized sclerostin powder was dissolved in double-distilled water to prepare a solution with a concentration of 1 μg / μl, and stored at -80°C for future use after aliquoting. When in use, it is diluted with double distilled water to a corresponding concentration and stored for later use. In this embodiment, it is diluted to 200 ng / ml.

Embodiment 2

[0033] Example 2 Construction of sclerostin neutralizing antibody based on sclerostin recombinant protein

[0034] According to the amino acid sequence of mouse sclerostin, a neutralizing antibody that can penetrate into the mouse body was constructed. The specific construction method is as follows:

[0035] Material preparation stage: Immunogen verification

[0036] Immunogen preparation: the mouse sclerostin constructed according to Example 1 was used as an immunogen, and the protein antigen was detected by SDS-PAGE. Responsible for coupling it with Immunoplus as an immunogen to break the immune tolerance of mice.

[0037] Quality control (QC): polyacrylamide gel electrophoresis (SDS-PAGE) method, purity>75%.

[0038] Phase 1: Animals immunized with SC2033-PF

[0039] A conventional immunization scheme was used to immunize 3 BALB / C mice + 3 C57 mice with the target protein-Immunoplus;

[0040] The routine immunization schedule is shown in the table below.

[0041]

...

Embodiment 3

[0063] According to the amino acid sequence of sclerostin, the sost overexpression plasmid was constructed and combined with the relatively mature bone-targeting virus AAV9-DSS-Nlter (standard reference: Yang, Y.S., et al. (2019)."Bone -targeting AAV-mediated silencing of Schnurri-3 prevents bone loss inosteoporosis." Nat Commun 10(1):2958.), construct a bone-targeting overexpression sost virus, which is used to inject into mice for related experimental verification.

[0064] Obtain the target gene by PCR or enzyme digestion, construct the target vector by ligation or homologous recombination / assembly, and transform E.coli DH5α competent cells. Transformants were screened by colony PCR, and positive clones were sent for sequencing. The verified clones were subjected to plasmid extraction. In this embodiment, PCR is used for construction.

[0065] Query the target gene and the upstream and downstream sequences from GenBank, and use VectorNTI software to design primers. The se...

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Abstract

The invention belongs to the field of research of related medicines for Alzheimer's disease, and particularly discloses sclerostin and application thereof in preparation of related products for treating or preventing Alzheimer's disease. The sclerostin is a protein secreted by bone cells and used for regulating and controlling bone mass, and the amino acid sequence of the sclerostin is shown as SEQ NO.1. Through synthesis of sclerostin recombinant protein and a corresponding neutralizing antibody, it is verified throughcell and zoology experiments that the sclerostin recombinant protein can accelerate generation of A beta protein outside brain tissue and accelerate cognitive function decline; after the neutralizing antibody is added, the BACE1 protein expression is obviously inhibited, so that the production of A beta protein is inhibited, and the effect of relieving Alzheimer's disease is achieved. The neutralizing antibody prepared on the basis of the sclerostin has a treatment and / or prevention effect, and can be used for preparing a medicine or a health care product for treating and / or preventing the Alzheimer's disease, so that a brand new choice and thought are provided for treating and / or preventing the Alzheimer's disease, and the selection field for treating the Alzheimer's disease is widened.

Description

technical field [0001] The invention relates to a new application in the field of biomedicine, belongs to the field of drug research related to Alzheimer's disease, and specifically relates to sclerostin and its application in the preparation of related products for treating or preventing Alzheimer's disease. Background technique [0002] Alzheimer's disease (AD) is a gradually aggravating neurodegenerative disease that widely affects the cerebral cortex and hippocampus of patients, leading to progressive memory loss and seriously affecting the cognitive function of normal elderly people. It brings serious living burden to patients, and also brings heavy economic pressure to society. By 2018, about 44 million people worldwide will suffer from Alzheimer's disease, and this number will reach 150 million in 2050. However, the specific mechanism is not very clear, and there is no significantly improved drug treatment. [0003] At present, the pathogenesis of Alzheimer's disease...

Claims

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Application Information

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IPC IPC(8): C07K14/47C07K16/18C12N15/12A61K38/17A61K39/395A61P25/28A23L33/18
CPCC07K14/4703A61P25/28A23L33/18A61K38/00A61K2039/505A23V2002/00A23V2200/322A23V2250/542
Inventor 蒋青郭保生石天舒
Owner 蒋青
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