Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Antibodies and methods for making and using them

a technology of antigen binding fragments and cell culture, which is applied in the field of cell culture production of can solve the problems of not providing an antibody that can be produced in high yield in cell culture, and reducing the time to prepare humanized antibodies or antigen binding fragments, so as to and improve the yield of antibodies

Inactive Publication Date: 2008-08-07
GENENTECH INC
View PDF48 Cites 80 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

The present invention is about improving the process of humanizing antibodies or antigen binding fragments and increasing the yield of antibodies or antigen binding fragments in cell culture. The invention involves modifying the primary sequence of antibody variable domains to improve their folding, assembly, and yield in cell culture. The invention provides methods for identifying which residues to modify and which consensus sequences to use for the modified residues, resulting in improved yield of antibodies or antigen binding fragments. The invention also includes expressing a modified framework region consensus sequence in a host cell and recovering the modified antibody or antigen binding fragment from the host cell. Overall, the invention provides methods for efficiently producing humanized antibodies or antigen binding fragments in high yield in cell culture.

Problems solved by technology

It has been discovered that selecting the most commonly occurring heavy and light chain consensus sequences may not provide an antibody that can be produced in high yield in cell culture.
The method may decrease the time to prepare a humanized antibody or antigen binding fragment that can be produced in high yield in cell culture.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Antibodies and methods for making and using them
  • Antibodies and methods for making and using them
  • Antibodies and methods for making and using them

Examples

Experimental program
Comparison scheme
Effect test

example 1

Heavy Chain Aggregation During Expression and Folding of Antibodies

[0210]Antibodies produced in cell culture can accumulate intracellularly, in the periplasm or in the extracellular medium. Antibody production typically involves expression of the light and heavy chains in the cytoplasm, secretion into the periplasmic space, folding of the light and heavy chains, and assembly of the folded light and heavy chains to form an antibody molecule. Multiple covalent and non-covalent interactions occur between and within the heavy and light chains during these folding and assembly processes. Antibody yield can be greatly affected by the efficiency and fidelity of these processes. Following synthesis of the heavy and light chains, protein aggregation or proteolysis can occur thereby reducing the yield of the antibody. Solubility experiments were carried out in order to assess the efficiency of steps in the antibody production pathway in bacterial cells for the purpose of improving antibody yi...

example 2

Preparation of Anti-VEGF Antibodies with Improved Yield

[0226]The yield of antibodies or antigen binding fragments from cells can be influenced by the in vivo folding and / or assembly of the antibody. In order to increase antibody or antigen binding fragment yield, the sequence of the antibody or fragment was modified and the effect on folding and yield on the antibody was examined. As described in Example 1, aggregation of heavy chains may contribute to a decrease in antibody folding, assembly and yield. Targeted modification of the heavy chain variable region FR1 of anti-VEGF antibodies was performed. Antibody variants with different human variable domain subgroup consensus sequences in the FR1 were prepared and the yield of completely assembled antibody products was examined.

Materials and Methods

A. Preparation of Expression Vectors Encoding Modified Anti-VEGF Antibodies

[0227]To evaluate the effect of different heavy chain FR1 human subgroup consensus sequences on the expression, fo...

example 3

Selection of a Heavy Chain FR1 Sequence for Antibodies Based on Comparison of HVR1 Antibody Sequences to HVR1 Consensus Subgroup I-III Sequences

[0278]Substitution of heavy chain FR1 consensus sequence subgroup III with consensus sequence subgroup I in anti-VEGF antibodies increased assembled antibody yield, presumably due to improved folding efficiency. Framework region sequences that provide for improved yield for other antibodies or antigen binding fragments can be identified based upon identifying the consensus sequence subgroup that has the most sequence identity in the HVR1 region of the variable domain when the HVR1 region of the antibody or fragment is compared to the corresponding HVR1 sequences of each of the subgroup consensus sequences. The method of identifying a heavy chain FR1 sequence for increasing the yield of antibodies based on HVR1 consensus subgroup I-III comparisons was tested for an anti-IgE antibody, E25.

A. Identification of a Heavy Chain FR1 Sequence for Hum...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
surface areaaaaaaaaaaa
temperatureaaaaaaaaaa
temperatureaaaaaaaaaa
Login to View More

Abstract

The present invention provides methods for producing humanized antibodies and increasing the yield of antibodies and / or antigen binding fragments when produced in cell culture. In one aspect of the invention, at least one framework region amino acid residue of the variable domain is substituted by a corresponding amino acid from a variable domain consensus sequence subgroup that has the most sequence identity with the HVR1 and / or HVR2 amino acid sequence of the variable domain. In another aspect, an amino acid is placed at a position proximal to a cys residue that participates in an intrachain variable domain disulfide bond that corresponds to an amino acid found at that position in a variable domain consensus sequence subgroup that has the most sequence identity with the HVR1 and / or HVR2 amino acid sequence of the variable domain.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]This application claims priority to U.S. Ser. No. 60 / 442,484, filed Jan. 23, 2003, under 35 U.S.C. 119(e), which application is hereby incorporated by reference.FIELD OF THE INVENTION[0002]The present invention generally concerns the production of antibodies or antigen binding fragments in cell culture. More specifically, the invention provides methods for improving the yield of recombinant antibodies or antigen binding fragments in cell culture.BACKGROUND[0003]Antibodies, particularly humanized antibodies, have become very useful for diagnostic and therapeutic purposes. Humanized antibodies are antibodies in which CDRs or hypervariable regions (HVRs) from a non-human antibody are combined with human framework regions to form an antigen binding molecule. This exchange is sometimes known as a “CDR swap”. There are different ways of selecting human framework sequences for humanized antibodies. One method involves selecting a human variable ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(United States)
IPC IPC(8): C12N15/09C07K16/00C07K16/22C07K16/42
CPCC07K16/00C07K16/22C07K2317/567C07K16/4291C07K16/42
Inventor SIMMONS, LAURA
Owner GENENTECH INC
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com