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PD-L1 IgV affinity peptide with antineoplastic activity, and preparation method and application of thereof

A technology with anti-tumor activity and affinity, applied in anti-tumor drugs, medical preparations containing active ingredients, peptides, etc.

Active Publication Date: 2014-10-15
ZHENGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, the use of PD-L1 blockers as tumor immunotherapy drugs or adjuvants has good application prospects and safety, but in the prior art, there is still a lack of better PD-L1 blocker products

Method used

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  • PD-L1 IgV affinity peptide with antineoplastic activity, and preparation method and application of thereof
  • PD-L1 IgV affinity peptide with antineoplastic activity, and preparation method and application of thereof
  • PD-L1 IgV affinity peptide with antineoplastic activity, and preparation method and application of thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] The targeting PD-L1 IgV affinity peptide with anti-tumor activity provided by the present invention, which specifically binds to the PD-L1 IgV region, is screened by phage display peptide library technology, and its amino acid sequence is: Ala-Asn-Gly -Ser-Arg-Leu-Val, namely A-N-G-S-R-L-V, the molecular weight is 715.4.

[0021] Due to the high cost of monoclonal antibody drugs and the inability to produce them on a large scale, we selected the PD-L1 IgV region protein expressed and purified from prokaryotic cells as the target, and screened the protein that can specifically bind to PD-L1 IgV through phage display technology. Peptides to block PD-1 / PD-L1 signaling pathway. For the convenience of those skilled in the art to implement the present invention, its screening process is briefly described as follows:

[0022] The preparation instructions for the media and master solutions used in the screening process are as follows:

[0023] LB liquid medium: Weigh 10 g o...

Embodiment 2

[0075] The affinity peptide of PD-L1 IgV with anti-tumor activity is synthesized by Fomc solid-phase peptide synthesis method, and the synthesis steps are briefly described as follows:

[0076] The main reagents used in the synthesis process are:

[0077] Heading liquid: acetic anhydride / pyridine solution (1:1, v / v);

[0078] Indene detection reagent: A. Ninhydrin / ethanol solution (5%, w / v)

[0079] B. Phenol / Ethanol (4:1, w / v)

[0080] C. Potassium cyanide / pyridine (2%, v / v)

[0081] Deprotection solution: piperidine / DMF solution (20%, v / v);

[0082] Cleavage reagent: by volume, TFA (82.5%), H 2 O (5%), phenol (5%), thioanisole (5%), ethanedithiol (2.5%).

[0083] The synthetic steps are briefly described as follows:

[0084] (1) Swell the resin, add the first amino acid

[0085] A. Swelling resin: Take 0.3~0.5 g Rink resin (the C-terminal amino acid of the peptide connected to the resin is an amide) and place it in a cleaned and dried peptide synthesizer, add an ap...

Embodiment 3

[0127] Taking the PD-L1 IgV affinity peptide with anti-tumor activity prepared in Example 2 as an example, the inventors conducted further in vivo experiments on tumor-bearing mice, and the specific experimental process was as follows:

[0128] Select 24 experimental Balb / c mice, adjust the cell concentration to 5×10 mouse colon cancer (CT26) cells with normal saline (NS) 6 cells / mL, 0.1 mL cell suspension (containing 5×10 5 cells) were inoculated subcutaneously in the armpit of the right forelimb of each Balb / c mouse, and the growth of the subcutaneous tumor was continuously observed.

[0129] The affinity peptide prepared in Example 2 was dissolved in physiological saline to prepare a polypeptide drug, which was subpackaged and stored at -20°C for future use.

[0130] Nine days after inoculation with mouse-derived colon cancer (CT26) cells, the mice were divided into groups according to tumor volume, 6 mice in each group, and divided into negative control group (NS), positi...

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Abstract

The invention belongs to the technical field of biopharmacy, particularly relates to a PD-L1 IgV targeted affinity peptide product with antineoplastic activity, and a preparation method and application of the affinity peptide product. According to the invention, the affinity peptide is specifically bonded in the PD-L1 IgV area, the amino acid sequence of the affinity peptide is ANGSRLV (Ala-Asn-Gly-Ser-Arg-Leu-Val), and the molecular weight of the affinity peptide is 715.4; the affinity peptide can be artificially synthesized by adopting the Fomc solid-phase polypeptide synthesis, and can be adopted as the mainly active ingredient in preparing colon cancer (CT26) resistance medicines. Bacteriophages are adopted to display the peptide bank screening technique, the PD-L1 IgV is adopted as the target for high-flux screening, and the affinity peptide with the antineoplastic activity is artificially synthesized; the inventor further proves that the affinity peptide has the better antineoplastic activity, can remarkably inhibit growth of tumors in rats, and provides the novel thinking and theoretical basis for the research and development of PD-L1-based medicines.

Description

technical field [0001] The invention belongs to the technical field of biopharmaceuticals, and specifically relates to the screening, preparation and application of a polypeptide product with anti-tumor activity, more specifically, the invention relates to an affinity peptide product targeting PD-L1 IgV with anti-tumor activity and its preparation and application. Background technique [0002] In recent years, the situation of tumor prevention and control is very severe. With the improvement of clinical diagnosis, surgical treatment, chemotherapy and radiotherapy, some patients can get early detection, early treatment, and better prognosis. However, finding new treatment methods and drugs has always been a global challenge. Research hotspots. Compared with traditional treatment methods, tumor immunotherapy can activate or induce tumor patients to establish a specific immune response to tumor antigens, eliminate primary tumor cells, and establish immune memory to prevent t...

Claims

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Application Information

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IPC IPC(8): C07K7/06C07K1/04A61K38/08A61P35/00
Inventor 高艳锋刘蓓媛祁元明李国栋李雯雯陈鲤翔赵园园周秀曼
Owner ZHENGZHOU UNIV
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