Human monoclonal antibody to human tumor necrosis factor

A technology of tumor necrosis factor and cloning antibody, which is applied in the biological field, can solve the problem of patients losing treatment effect, and achieve the effects of strong neutralizing activity, enhanced anti-aggregation, and good specificity

Active Publication Date: 2021-10-15
武汉班科生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing antibodies against TNFα are prone to anti-antibody phenomenon in clinical use, which leads to the loss of therapeutic effect of the antibody on some patients. Therefore, the development of new human monoclonal antibodies against TNFα will help alleviate this problem

Method used

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  • Human monoclonal antibody to human tumor necrosis factor
  • Human monoclonal antibody to human tumor necrosis factor
  • Human monoclonal antibody to human tumor necrosis factor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0014] Embodiment 1: Obtaining of FCO antibody sequence

[0015] According to the existing literature (Zhu Z, Dimitrov DS, Methods Mol Biol, 2009), a human Fab phage display library was constructed, and human TNFα protein was screened for four rounds based on the library, and by monoclonal phageELISA, the The positive clone was sent for sequencing, and the base sequence of the FCO candidate clone was obtained, and its amino acid sequence was obtained through the amino acid translation software (https: / / web.expasy.org / translate / ), and the light chain amino acid sequence is shown as SEQNo.1 Shown, its heavy chain amino acid sequence is shown in SEQ No.2.

Embodiment 2

[0016] Example 2: Using Tango software to optimize the obtained FCO amino acid sequence

[0017] Use Tango software (http: / / tango.crg.es / ) to predict the aggregation-prone region of the amino acid sequence of the FCO candidate clone, and find that there is a strong aggregation-prone region, such as Picture 1-1 , so point mutation is carried out in this region to obtain FC9 antibody, the amino acid sequence of its light chain is shown in SEQ No.1, the amino acid sequence of its heavy chain is shown in SEQ No.3, the corresponding base sequence, and the light chain is shown in SEQ No. 4, the heavy chain is shown in SEQ No.5. The mutated amino acid sequence was re-predicted by Tango software, and it was found that the aggregation-prone region was significantly improved, as shown in Figure 1-2 , 1-3.

Embodiment 3

[0018] Example 3: 293F cells express FC9 antibody and detect it by SDS-PAGE

[0019] The mutated base sequence was cloned into the dual promoter vector pVitro2-neo-mcs (InvivoGen), and the 293F cells were transfected with PEI to express the protein. After 5-7 days of expression, the culture supernatant of the 293F cells was treated with ProteinA (GE) was purified, and after the purified protein was concentrated and exchanged, the FC9 antibody dissolved in PBS buffer was obtained. After SDS-PAGE electrophoresis detection, it was found that it presented two bands of light chain and heavy chain, such as figure 2 shown.

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Abstract

The invention discloses a human monoclonal antibody against human tumor necrosis factor. After screening and genetic modification, the obtained antibody has good specificity, strong neutralizing activity and increased anti-aggregation, and is suitable for preparing anti-autoimmune Therapeutic drugs for diseases.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a human monoclonal antibody of human tumor necrosis factor. Background technique [0002] Tumor necrosis factor (TNFα) is a human inflammatory response-related protein, and excessive TNFα can lead to the occurrence of autoimmune diseases. The development of antibodies against TNFα will help reduce the content of TNFα in the body and accelerate its clearance in the body, thereby treating related autoimmune diseases. The existing anti-TNFα antibodies are prone to anti-antibody phenomenon in clinical use, which leads to the loss of therapeutic effect of the antibodies on some patients. Therefore, the development of new human monoclonal antibodies against TNFα will help alleviate this problem. Contents of the invention [0003] The purpose of the present invention is to fill the gap in the prior art and provide a human monoclonal antibody against human tumor necrosis factor. [0004]...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/24C12N15/13
CPCC07K16/241
Inventor 龚睿
Owner 武汉班科生物技术有限公司
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