Anti-human PTH1R extracellular region blocking type single-domain antibody and application thereof

A single-domain antibody, anti-human technology, applied in the field of biomedicine

Active Publication Date: 2022-08-09
NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Current treatments for SHPT include active vitamin D compounds (such as calcitriol and some vitamin D analogs), phosphate binders, and more recently calcimimetics (such as cinacalcet), but these agents have limitations sex [6]

Method used

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  • Anti-human PTH1R extracellular region blocking type single-domain antibody and application thereof
  • Anti-human PTH1R extracellular region blocking type single-domain antibody and application thereof
  • Anti-human PTH1R extracellular region blocking type single-domain antibody and application thereof

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Experimental program
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Effect test

Embodiment 1

[0032] 1. Antigen Preparation

[0033] 1.1 Construction and purification of wild-type and mutant human PTH1R ECD-hFc plasmids

[0034] According to the structure of human PTH1R ECD, the key site of its binding to PTH was mutated and used as a negative antigen for subsequent identification of antibody characteristics. Construction of wild-type and mutant human PTH1R ECD-hFc plasmids using pFUSE as a vector ( figure 1 ), the human Fc tag is beneficial to the purification of the antibody by the Protein A adsorption column. Using the purified wild-type and mutant human PTH1R ECD-hFc protein, it is expected that the antibody that can compete with PTH for binding to the key site of PTH1R can be obtained. Blocks the interaction of PTH with PTH1R.

[0035] 1.2 Identification of wild-type and mutant human PTH1R ECD-hFc proteins

[0036] The purified wild-type and mutant human PTH1R ECD-hFc proteins were verified by agarose gel electrophoresis experiments to verify the molecular weight...

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Abstract

The invention discloses a fully humanized monoclonal single-domain antibody for resisting human PTH1R and an application of the fully humanized monoclonal single-domain antibody. The amino acid sequence of the antibody is shown as SEQ ID NO: 2. According to the invention, aiming at wild and mutant human PTH1R ECD proteins, antibody screening is carried out by using a phage display technology, and the anti-human PTH1R blocking type single-domain antibody with a blocking effect is screened and obtained through methods such as ELISA, FACS and the like. The antibody can be combined with human PTH1R, can block the interaction between PTH and PTH1R, and has a potential application value in human body dysfunction mediated by PTH-PTH1R, such as chronic kidney disease secondary hyperparathyroidism.

Description

Technical field: [0001] The invention belongs to the field of biomedicine, in particular to an anti-human PTH1R fully human monoclonal single domain antibody with blocking effect and its application. Background technique: [0002] Chronic kidney disease (CKD) refers to chronic kidney structural and functional abnormalities (with a history of kidney damage greater than 3 months) caused by various causes, including pathological damage to glomerular filtration rate (GFR), Abnormal blood or urine composition, abnormal imaging examination, or unexplained decreased GFR (<60mL / min / 1.73m 2 ) for more than 3 months. Recent statistics show that there are 698 million full-stage CKD cases in the world, with a prevalence rate of 9.1%. There are 132 million patients in my country, which is the country with the largest number of CKD patients in the world. [1] . The clinical manifestations of CKD are different in different stages. In the early stage, only mild symptoms such as fatigue...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28C12N15/13C12N15/85C12N5/10A61K39/395A61P13/12A61P5/20
CPCC07K16/2869C12N15/85A61P13/12A61P5/20C07K2317/21C07K2317/569C07K2317/76A61K2039/505
Inventor 高威温嘉奇杨刚赫瑞婷刘晓宇高占辉金潇怡马素娟
Owner NANJING MEDICAL UNIV
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