Method to screen phage display libraries with different ligands

Abstract

Abstract of the Disclosure The present invention relates to methods for selecting repertoires of polypeptides using generic and target ligands. In particular, the invention relates to a library comprising a repertoire of polypeptides of the immunoglobulin superfamily, wherein the members of the repertoire have a known main chain conformation.

Description

Detailed Description of the InventionIntroduction[0001] The present invention relates to methods for selecting repertoires of polypeptides using generic and target ligands. In particular, the invention describes a method for selecting repertoires of antibody polypeptides with generic ligand to isolate functional subsets thereof.[0002] [0002]The antigen binding domain of an antibody comprises two separate regions: a heavy chain variable domain (VH) and a light chain variable domain (VL: which can be either Vκ or Vλ). The antigen binding site itself is formed by six polypeptide loops: three from VH domain (H1, H2 and H3) and three from VL domain (L1, L2 and L3). A diverse primary repertoire of V genes that encode the VH and VL domains is produced by the combinatorial rearrangement of gene segments. The VH gene is produced by the recombination of three gene segments, VH, D and JH. In humans, there are approximately 51 functional VH segments (Cook and Tomlinson (1995) Immunol Today, 16:...

AI Technical Summary

Benefits of technology

[0015]In a third aspect, the invention provides a method for detecting, immobilising, purifying or immunoprecipitating one or more membe

Problems solved by technology

Although synthetic libraries help to overcome the inherent biases of the natural repertoire which can limit the effective size of phage libraries constructed from rearranged V genes, they require the use of long degenerate PCR primers which frequently introduce base-pair deletions into the assembled V genes.

This high degree of randomisation may also lead to the creation of antibodies which are unable to fold correctly and are also therefore non-functional.

Furthermore, antibodies selected from these libraries may be poorly expressed and, in many cases, will contain framework mutations that may effect the antibodies immunogenicity when used in human therapy.

Since it is desirable to produce artificial human antibodies which will not be recognised as foreign by the human immune system, the use of consensus frameworks which, in most cases, do not correspond to any natural framework is a disadvantage of this approach.

A further problem with the libraries of the prior art is that, because the main-chain conformation is heterogeneous, three-dimensional structural modelling is difficult because suitable high resolution crystallographic data may not be available.

This is a particular problem for the H3 region, where the vast majority of antibodies derived from natural or synthetic antibody libraries have medium length or long loops and therefore cannot be modelled.

Images