Super-humanized antibody

A chimeric antibody, non-human technology, applied in the direction of hybrid immunoglobulin, etc., can solve problems such as no explanation

Inactive Publication Date: 2011-02-02
杰斐逊 富特
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there is still a need for humanized framework sequences, and these references do not describe methods for selecting a sufficient number of human framework sequences for use with a given set of murine CDRs

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0098] Example 1. Humanized anti-chicken lysozyme antibody

[0099] Mouse antibody D1.3 binds to chicken lysozyme antigen. The peptide sequence of D1.3 variable region comes from the protein database, accession number 1VFA. The light chains are numbered according to Kabat, and the canonical structure type is assigned to the mouse CDR as follows.

[0100] The light chain CDR1 labeled according to Kabat consists of the following sequence:

[0101] 24 25 26 27 28 29 30 31 32 33 34

[0102] R A S G N I H N Y L A

[0103] Because there is no insertion or deletion between residues 27 and 31, CDR1 has a canonical structure type 2.

[0104] The light chain CDR2 labeled according to Kabat consists of the following sequence:

[0105] 50 51 52 53 54 55 56

[0106] Y T T T L A D

[0107] This is not a special sequence, its structure type is Type 1.

[0108] The light chain CDR3 labeled according to Kabat consists of the following sequence:

[0109] 89 90 91 92 93 94 95 96 97

[0110] Q H F W S T P R T

...

Embodiment 2

[0128] Example 2. Humanized anti-human CD28 antibody

[0129] The mouse anti-human CD28 antibody called 9.3 was used as the non-human antibody under study. The murine 9.3 hybridoma line was isolated, and Hansen et al. (1980) described the hybridoma line.

[0130] The heavy and light chain variable region genes of 9.3 were cloned by reverse transcription and polymerase chain reaction, starting from messenger RNA separated by the guanidinium isothiocyanate method (Chomczynski and Sacchi, 1987) and oligo dT column chromatography. The amplification is guided by oligonucleotides complementary to the constant region and oligonucleotides corresponding to the single peptide region or the N-terminal framework sequence.

[0131] Number the light chain according to Kabat and refer to Picture 8 , Assign the canonical structure type to the CDR as follows.

[0132] The light chain CDR1 numbered according to Kabat consists of the following sequence

[0133] 24 25 26 27 a b c d 28 29 30 31 32 33 34 ...

Embodiment 3

[0161] Example 3. Humanized anti-scorpion toxin antibody

[0162] The mouse anti-scorpion toxin antibody called BCF2 was used as a non-human sequence of humanized anti-scorpion toxin. Murine BCF2 hybridomas have been described, and Licea et al. (1996) demonstrated the effectiveness of BCF2 antibodies in murine models. Selisko et al. (1999) clarified the variable region sequence of BCF2, in Picture 12 These sequences are listed in.

[0163] As described above, the canonical structure type of the light chain is determined as CDR1 is type 5, CDR2 is type 1, and CDR3 is type 1. The canonical structure types of heavy chain CDRs are type 1 for CDR1 and type 2 for CDR2. Using the considerations discussed above for the selection of human germline V and J gene sequences, a humanized version of BCF2 was designed.

[0164] The composition of the light chain variable region is: the Kabat CDR sequence of the BCF2 light chain; and the framework sequence, which is consistent with human gene A2 / ...

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Abstract

The invention discloses a method for humanizing an antibody. In the invention, through comparing a standard CDR (Complementarity-Determining Region) structure type of a CDR sequence in a variable region of an inhuman antibody with a standard CDR structure type of a corresponding CDR in a human antibody sequence library, a frame sequence of the variable region is selected from human antibody genes, preferably the gene segments of an embryonic system antibody. The variable region of the human antibody which has the standard CDR structure type similar to an inhuman CDR forms a subgroup of a member antibody sequence and a human frame sequence can be selected from the subgroup. The subgroup members can be further rated through the amino acid similarity between the human and inhuman CDR sequences. The ahead human sequence in the rating is selected to provide the frame sequence and construct a chimeric antibody by utilizing the selected the subgroup member human frame. The chimeric antibody replaces the human CDR sequence by the counterpart functionality of the inhuman CDR, therefore, the humanized antibody which has high affinity and low immunogenicity is provided without comparing the frame sequences between the inhuman antibody and the human antibody. The invention also discloses the chimeric antibody prepared according to the method of the invention.

Description

[0001] This application is a divisional application of the Chinese invention patent application No. 02829598.6 filed on July 12, 2002 with the title of "superhumanized antibody". [0002] Cross references to related applications [0003] This application claims the priority of U.S. Provisional Application 60 / 305,111 filed on July 12, 2001. [0004] Government funding statement [0005] The research and development of the present invention was funded by the National Institutes of Health, the fund number is CA-18029. Technical field [0006] The present invention relates to a method for humanizing antibodies, in particular to a method for humanizing antibodies by generating chimeric antibodies containing CDR sequences derived from non-human antibodies and framework sequences of human antibodies, and more particularly to methods for selection Methods for humanization of appropriate human antibody framework sequences, and still more particularly involving the comparison of the canonical CD...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/46
Inventor 杰斐逊·富特
Owner 杰斐逊 富特
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