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A specific polypeptide targeting and binding to lymphoma cell lines and its application

A cancer cell and lymphatic technology, which can be applied to specific polypeptides that target and bind to lymphoid cancer cell lines and its application fields, which can solve the problems of increasing the economic burden of patients and poor marker specificity.

Active Publication Date: 2021-11-23
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These markers have been widely used clinically, but due to factors such as different subtypes and poor marker specificity, it is often necessary to use multiple markers in combination
These markers have both advantages and limitations, and the combined use of multiple markers will inevitably increase the economic burden of patients

Method used

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  • A specific polypeptide targeting and binding to lymphoma cell lines and its application
  • A specific polypeptide targeting and binding to lymphoma cell lines and its application
  • A specific polypeptide targeting and binding to lymphoma cell lines and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1: Culture of Jeko-1 lymphoma cell line, phage titer determination, phage display panning

[0026] A. Culture of Jeko-1 lymphoma cell line:

[0027]The Jeko-1 lymphoma cell line (ATCC, American Type Culture Collection) was placed at 37°C, 5% CO 2 The cells were cultured in an incubator, and 10% calf serum, 100 U / mL penicillin and 100 U / mL streptomycin were added to the 1640 medium.

[0028] B. Phage titer determination:

[0029] (1) ER2738 glycerol bacteria (NEB Company) were taken at -80°C, streaked on LB-Tet plate, and cultured upside down at 37°C for 12h-16h.

[0030] (2) Use a sterile tip to pick a single colony of ER2738 into 5-10mL LB-Tet medium, culture it on a shaker at 37°C and 220rpm until mid-log phase (OD 600 = 0.5).

[0031] (3) Heat and melt the Top agar in a microwave oven, divide it into 3mL equal portions in sterile centrifuge tubes, and keep warm at 45°C for later use.

[0032] (4) Pre-warm the LB / IPTG / Xgal plate at 37°C.

[0033] (5) The...

Embodiment 2

[0068] Example 2. Acquisition of phage monoclonal and analysis of biological information

[0069] Monoclonal phage amplification and purification:

[0070] (1) Take ER2738 glycerol bacteria at -80°C, streak on LB-Tet plate, and incubate upside down at 37°C for 12h-16h.

[0071] (2) The ER2738 monoclonal plaques obtained in step (1) were cultured in 20 mL LB-Tet at 37° C. and 220 rpm on a shaker for 12h-16h.

[0072] (3) Inoculate ER2738 overnight culture in step (2) at 1:100 dilution in LB medium, divide 1 mL into 15 mL centrifuge tubes.

[0073] (4) Take the LB plate used for the titer determination of the fourth round of panning, pick blue phage plaques with the tip of a pipette in step (3) 1mLER2738 bacterial solution, and culture on a shaker at 37°C and 220rpm for 4h-5h.

[0074] (5) Transfer the culture to a centrifuge tube, centrifuge at 14000rpm for 30s, transfer the supernatant to a new centrifuge tube, repeat the centrifugation for 30s, transfer 80% of the supernata...

Embodiment 3

[0094] Example 3. The cultivation of lymphoma cells, the extraction and primary culture of human lymphocytes, the synthesis of fluorescent polypeptides, and the immunofluorescence experiment of polypeptides

[0095] Culture of Lymphoma Cells:

[0096] Lymphoma cell lines Jeko-1, Romas, Raji, Su-4, Granta-519 (all purchased from ATCC, American Type Culture Collection) were cultured in RPMI-1640 medium containing 10% fetal bovine serum. Mouse mast cell carcinoma cell P815 (purchased from ATCC, American Type Culture Collection) was cultured in DMEM medium containing 10% fetal bovine serum. All cells were stored at 37°C, 5% CO 2 Routine culture in a cell incubator.

[0097] Extraction and primary culture of human lymphocytes:

[0098] (1) Transfer the blood of a normal person into a 15mL centrifuge tube, centrifuge at 500g for 8min, and separate the supernatant serum into a new 15mL centrifuge tube.

[0099] (2) Add an equal volume of PBS to the serum obtained in step (1), and...

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Abstract

The invention relates to a specific polypeptide targeting and binding to lymphoid cancer cell lines and its application, which belongs to the field of biomedicine; the invention is based on in vitro phage display technology, and the specific polypeptide TUZG12 capable of targeting and binding to lymphoid cancer cell lines is obtained by panning in vitro In vitro cell uptake and distribution experiments prove that the polypeptide has targeted binding activity to lymphoma cell lines; the present invention also shows through in vitro cell activity experiments that pro-apoptotic peptide DKK coupled with TUZG12 polypeptide can improve to a certain extent The inhibitory effect of DKK on the proliferation activity of lymphoma cells; the polypeptide screened by the present invention has good targeting binding activity to multiple lymphoma cell lines, and can improve the killing effect of cytotoxic polypeptides on target tumor cells. It has important application value in diagnosis and treatment.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to a specific polypeptide targeting lymphoma cell line and its application. Background technique [0002] Compared with traditional antibody-based targeting molecules, small-molecule polypeptides that specifically bind to tumors have lower immunogenicity, lower production costs, and higher surface density (more peptides per unit surface, so This conjugate has higher affinity) and better tissue penetration effect, and can be used as a marker for tumor diagnosis, and it can also be coupled with traditional chemotherapy drugs to achieve targeted treatment of tumors. Among the peptide drugs approved by the FDA, some peptide drugs have been designated for cancer treatment. Leuprolide, a gonadotropin-releasing hormone analog, has been approved for the treatment of prostate cancer, and the proteosome inhibitor bortezomib is clinically used for the treatment of multiple myeloma. The...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/08C07K19/00A61K47/42A61K38/16A61P35/00
CPCA61K38/00A61K47/42A61P35/00C07K7/08C07K2319/00
Inventor 刘晗青张雅菲卢子文屠志刚梁智全
Owner JIANGSU UNIV
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