Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Aptamer of sialic acid-binding immunoglobulin-like lectin-15 protein and application thereof

A nucleic acid aptamer and protein technology, applied in biochemical equipment and methods, instruments, analytical materials, etc., can solve the problems of antibody instability, slow antibody production cycle, easy inactivation cost, etc.

Pending Publication Date: 2021-07-16
XIAMEN UNIV
View PDF5 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, many immune checkpoint antibodies including Siglec-15 are still not on the market, because the production cycle of antibodies is slow, coupled with the disadvantages of instability, easy inactivation and high cost of antibodies, so there is an urgent need to develop other more stable, More effective and lower cost immunotherapy drugs

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Aptamer of sialic acid-binding immunoglobulin-like lectin-15 protein and application thereof
  • Aptamer of sialic acid-binding immunoglobulin-like lectin-15 protein and application thereof
  • Aptamer of sialic acid-binding immunoglobulin-like lectin-15 protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The screening of embodiment 1 Siglec-15 nucleic acid aptamer

[0022] Chemically synthesize an 85-base DNA initial library. The two ends of the DNA initial library are fixed sequences, including a random sequence of 45 bases in the middle, which is 5'-AAGGAGCAGCGTGGAGGATA-45N-TTAGGGTGTGTCGTCGTGGT-3', library capacity for 10 15 above. Take 5nmol DNA initial library dissolved in binding buffer (12mmol / L PBS, pH 7.4, 150mmol / L NaCl, 5mmol / L KCl, 0.55mmol / L MgCl 2 ) for denaturation treatment: 10 minutes at 95°C, 10 minutes on ice, and then 10 minutes at 25°C.

[0023] 2) The recombinant Siglec-15 protein (purchased from Sino Biological) expressed by polyhistidine tag was used as the screening target of the aptamer. Siglec-15 protein is modified on Ni agarose microspheres, and Siglec-15-Ni agarose microspheres are formed by interacting with Ni through the epitope of His-tag.

[0024] 3) The obtained pretreated initial library solution was incubated with Siglec-15-Ni aga...

Embodiment 2

[0027] Example 2 Investigating the Binding Ability and Affinity of Nucleic Aptamers and Siglec-15 by Flow Cytometry

[0028]1) Use His-tag Siglec-15 modified Ni agarose microspheres to monitor the process of enrichment. The results of flow cytometry showed that the fluorescence intensity combined with Siglec-15-Ni-beads increased significantly with the increase of the number of rounds of screening ( figure 1 A), while the fluorescence intensity of Ni-beads did not increase ( figure 1 B), showing that after 11 rounds of screening, the DNA library was successfully enriched with the recognition sequence of Siglec-15 protein.

[0029] 2) Some sequences were obtained by TA clone sequencing of the enriched library, namely WXY1, WXY2, WXY3, WXY4, WXY5, WXY6 and WXY7 shown in SEQ ID NO: 1 to SEQ ID NO: 7 (Table 1). figure 2 A is the initial library, the enrichment library and the combination of 7 nucleic acid aptamers screened out and Siglec-15-Ni agarose microspheres. Although di...

Embodiment 3

[0032] Example 3 Nucleic acid aptamer in Siglec-15 / CD8 + Application of T cell immunotherapy

[0033] 1) CFSE staining CD8 + T detection aptamer pair CD8 + effect on T cell proliferation. Add activated CD8 to Siglec-15 protein + T cells and different nucleic acid aptamers: random sequence, SEQ ID NO: 3 at a final concentration of 2 μM, supplemented every 12 hours. After 72 hours of incubation, the results of flow cytometry showed that compared with only CD8 + T system, add CD8 + After T cells, the activity of T cell proliferation decreased by about 50%, and the nucleic acid aptamer SEQ ID NO: 3 restored the proliferation ability of T cells to a large extent, for only CD8 + 93% of the system of T ( Figure 5 A).

[0034] 2) Use CCK-8 to detect nucleic acid aptamer pair CD8 + The effect of T cells on the killing ability of cancer cells. Add activated CD8 to U87 cells + T cells and different nucleic acid aptamers: random sequence, SEQ ID NO: 3 at a final concentration ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses an aptamer of sialic acid-binding immunoglobulin-like lectin-15 (Siglec-15) protein. The aptamer comprises nucleotide sequences as shown in SEQ IDNO: 1- SEQ ID NO: 7. The aptamer provided by the invention can provide rapid and accurate detection on Siglec-15 protein. Meanwhile, The aptamer can block the interaction with T cell surface receptors, and reactivate proliferation of T cells to a certain extent, thereby normalizing immunity in tumor microenvironment without causing serious autoimmune response. The aptamer for Siglec-15 can be used as a medicine for cancers with high Siglec-15 expression in tumor microenvironment, and becomes a potential nucleic acid medicine for tumor immunotherapy. The aptamer of the present invention is possibly effective for patients who have no response to anti-PD-L1 treatment, and further supplements existing tumor diagnosis and immunotherapy systems.

Description

technical field [0001] The invention relates to a nucleic acid aptamer of a sialic acid-binding immunoglobulin-like lectin-15 (Siglec-15) protein and an application thereof. Background technique [0002] Cancer is a global public health problem and a major threat to human health. With the advancement of science and the development of cancer-related disciplines, tumor immunology research and immunotherapy have developed rapidly. Among them, the immunosuppressant represented by PD-1 / L1 is the most eye-catching in tumor immunotherapy and has brought new hope to the majority of tumor patients. However, this immunotherapy is not effective for all tumor patients. Under normal circumstances, the overall effective rate of PD-1 / L1 antibodies against solid tumors is still only about 20%. [0003] Sialic acid-binding immunoglobulin-like lectin-15 (Sialic acid-binding immunoglobulin-like lectin-15, Siglec-15) can mediate the interaction between cells and cells or pathogens by recogniz...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/115G01N33/574
CPCC12N15/115G01N33/57484C12N2310/16
Inventor 杨朝勇宋彦龄魏心语陈福德黄梦娇吴秋月周雷激
Owner XIAMEN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com