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Full-humanized anti-PD-1 monoclonal antibody and preparation method and application thereof

A monoclonal antibody, PD-1 technology, applied in the direction of antibody, anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, antiviral agent, etc., to achieve the effect of promoting secretion, high affinity and wide application prospect

Inactive Publication Date: 2013-08-14
ZHENGZHOU UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

These results confirm that blocking PD-1 signaling may be applied to the clinical treatment of viral infections such as HIV

Method used

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  • Full-humanized anti-PD-1 monoclonal antibody and preparation method and application thereof
  • Full-humanized anti-PD-1 monoclonal antibody and preparation method and application thereof
  • Full-humanized anti-PD-1 monoclonal antibody and preparation method and application thereof

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Embodiment Construction

[0027] The present invention will be further described below in conjunction with specific examples and test examples, but it should not be understood as a limitation of the present invention.

[0028] 1. Preparation of cDNA

[0029] Collect 10 ml of peripheral blood from 50 people with liver cancer, kidney cancer, melanoma, and healthy people, anticoagulate with heparin sodium, separate mononuclear cells (PBMC) by density gradient centrifugation, and use PD-1 antigen for in vitro sensitization. After PBMC were sensitized in vitro, press 4×10 6 Add 1ml of EBV culture supernatant to PBMC, incubate at 37°C for 3 hours, discard the supernatant, add RPMI 1640 complete medium containing 100ml / L FCS, culture for 2 weeks, replace with RPMI 1640 complete medium containing 200ml / L NBS . The culture supernatant of transformed cells was screened by indirect ELISA method, using PD-1 as the tumor cell antigen solid phase plate, the collected B cell culture supernatant as the primary antib...

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Abstract

The invention discloses a full-humanized anti-PD-1 monoclonal antibody having a heavy-chain amino acid sequence shown in a sequence 7 in a sequence table and a light-chain amino acid sequence shown in a sequence 8 in the sequence table. The full-humanized anti-PD-1 monoclonal antibody has the advantages that the antibody has high appetency and low immunogenicity on PD-1, is efficiently expressed in animal cells, and can be used for industrial production. An experiment proves that the full-humanized anti-PD-1 monoclonal antibody specifically blocks a PD-1 / PD-L inhibiting signal, so that a disabled effector cell in an organism restores a biological function; activation proliferation of a tumor and a virus specificity CD8+T cell and secretion of a cell factor are facilitated; the killability of lymphocyte on tumor antigen, an exotic invasive virus and the like is enhanced; the immunity of the organism is improved; and the tumor cell and the virus are timely cleared. Therefore, the antibody disclosed by the invention has a wide application prospect on treatment of tumors, infectious diseases and autoimmune diseases.

Description

technical field [0001] The invention relates to the field of development and production of antibodies and antibody drugs, in particular to the acquisition of a fully humanized monoclonal antibody against human PD-1, and also relates to a preparation method and application of the monoclonal antibody. Background technique [0002] PD-1 (CD279) is an immunoglobulin superfamily type I transmembrane glycoprotein composed of 288 amino acids, which was originally cloned from the apoptotic mouse T cell hybridoma 2B4. Named programmed death-1 (programmed death-1, PD-1). When initially discovered, PD-1 was recognized as a member of the CD28 superfamily because of its structural resemblance to the CD28 molecule. However, PD-1 protein lacks the MYPPPY sequence that mediates the combination of CD28 / CTLA-4 and B7.1 / B7.2, and the FDPPPF sequence that mediates the combination of ICOS and ICOS-L, so it is structurally related to CD28, CTLA -4 and ICOS are significantly different. Therefo...

Claims

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Application Information

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IPC IPC(8): C07K16/28C12N15/85A61K39/395A61P31/12A61P35/00A61P37/02
Inventor 方惠敏张守涛张一折
Owner ZHENGZHOU UNIV
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