Conjugate and application of conjugate in degrading beta-catenin protein

A conjugate and protein technology, applied in the field of biomedicine, can solve the problems of unapproved clinical use, difficult to inhibit the function of β-catenin, and compounds that degrade β-catenin protein need to be studied.

Active Publication Date: 2020-06-19
南昌双天使生物科技开发有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In the past, Wnt signaling inhibitors targeting β-catenin protein were almost all chemical small molecules. Since the binding interface between β-catenin and other proteins is large, and the binding proteins are diverse and complex, small molecule inhibitors are difficult to obtain. Completely inhibit the function of β-catenin, so far no small molecule inhibitor of Wnt / β-catenin has been approved for clinical use
[0004] Therefore, compounds suitable for degrading β-catenin protein are still to be studied

Method used

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  • Conjugate and application of conjugate in degrading beta-catenin protein
  • Conjugate and application of conjugate in degrading beta-catenin protein
  • Conjugate and application of conjugate in degrading beta-catenin protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0110] Example 1 Use of conjugates to specifically degrade β-catenin protein and inhibit Wnt / β-catenin signaling pathway

[0111] 1. In HEK293T cells, add 70 μM of the test substance in the control (unstimulated) and Wnt3a stimulated state respectively. Wherein, the DMSO group is a control group; the test product of the control group 1 and the experimental group 1 is the amino acid shown in SEQ ID NO: 11; the test product of the control group 2 and the experimental group 2 is the conjugate 1 (SEQ ID NO: 12); the test substance of the control group 3 and the experimental group 3 is the amino acid shown in SEQ ID NO: 1; the test substance of the control group 4 and the experimental group 4 is the conjugate 2 (SEQ ID NO: 10).

[0112] (SEQ ID NO: 11)

[0113]

[0114] (SEQ ID NO: 12)

[0115]

[0116] Such as figure 1 As shown in A, when Wnt3a is stimulated, the level of β-catenin protein is significantly increased, and only conjugate 2 has a significant down-regulat...

Embodiment 3

[0119] Example 3 The conjugate effectively inhibits the tumorigenesis of colon cancer cell lines in nude mice subcutaneously

[0120] In this experiment, BALB / C immunodeficiency nude mice were selected. First, the colon cancer cell line LoVo cells were pretreated with DMSO and the sample to be tested, and treated with 50 μM concentration for 24 hours, and then the same amount of pretreated cells ( About 3×10 6 ) were injected subcutaneously into nude mice, and after 3 weeks of growth, the subcutaneous tumors of the mice were taken out and weighed.

[0121] The result is as figure 2 As shown, wherein, the DMSO group is a control group; the test substance of the experimental group 2 is conjugate 1 (SEQ ID NO: 12); the test substance of the experimental group 3 is the amino acid shown in SEQ ID NO: 1; the test substance of the experimental group The test article of 4 was conjugate 2 (SEQ ID NO: 10), and there were 10 parallel experiments in each experimental group.

[0122] C...

Embodiment 4

[0123] Example 4 The conjugate effectively reduces the number of tumors in the intestinal tract of mice

[0124] APC was chosen in this experiment min / + Mice, due to the deletion mutation of the APC gene in this mouse, the APC gene is inactivated, resulting in the continuous activation of the Wnt / β-catenin signaling pathway. The most obvious phenotype is the intestinal Massive tumors develop. Select 11-week-old APCs min / +The mice were divided into 5 groups, one group was not treated (Day0) as the control group, and the number of tumors in the intestinal tract was directly dissected and recorded, and the other four groups were respectively treated with 30 mg / kg DMSO, conjugate 1 (experimental group 2), The amino acid shown in SEQ ID NO: 1 (experimental group 3) and the conjugate 2 (experimental group 4) were dissolved in normal saline and injected intraperitoneally to the mice, and the mice were administered 7 times within a 14-day administration cycle. Inject, that is, inje...

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Abstract

The invention provides application of E3 ubiquitin ligase VHL in degradation of beta-catenin protein or inhibition of a Wnt/beta-catenin signal pathway and a conjugate as well as a preparation methodand application of the conjugate. The conjugate comprises a first peptide fragment suitable for binding to beta-catenin protein; a second peptide fragment suitable for binding to E3 ubiquitin ligase VHL; a connecting arm, wherein the first peptide fragment is connected with second peptide fragment through the connecting arm. The E3 ubiquitin ligase VHL can be used for carrying out ubiquitination modification on beta-catenin protein, so that the ubiquitination beta-catenin protein is degraded by protease, a Wnt/beta-catenin signal channel is inhibited, and the effects of inhibiting cancer cellproliferation and tumor-forming ability are achieved; wide scientific research and clinical application prospects are realized.

Description

technical field [0001] The invention relates to the field of biomedicine. Specifically, the present invention relates to conjugates and their application in degrading β-catenin protein. Background technique [0002] The Wnt / β-catenin signaling pathway plays an important role in controlling cell proliferation, differentiation, and survival. Members of this signaling pathway include: Wnt ligands secreted outside the cell, transmembrane receptors on the cell membrane, and intracellular degradation Complex and β-catenin. As a signal transduction molecule of the canonical Wnt signaling pathway, the stability of β-catenin protein is finely regulated. Under pathological conditions, once members of the Wnt / β-catenin signaling pathway undergo gene mutations and lead to regulatory disorder, excess β-catenin will accumulate in the cytoplasm and then enter the nucleus to promote cell proliferation. Many cancers, especially intestinal cancers, have been shown to result from overactiva...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/53A61K47/64A61K47/65A61P1/00A61P35/00G01N33/68
CPCA61K38/53C12Y603/02019A61P35/00A61P1/00A61K47/65A61K47/64G01N33/68
Inventor 陈晔光胡宏岗刘磊廖洪蔚李翔赵联正
Owner 南昌双天使生物科技开发有限公司
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