Patent ID: 6262238
Filing Date: 2001-07-17
Classification: A61K

Abstract:
A process for improving the stability of an antibody variable domain by modifying an initial antibody variable domain, the process comprising the following steps:(A) providing an initial gene encoding an amino acid sequence of an initial variable domain to be modified;(B) comparing the amino acid sequence encoded by the initial gene with one of consensus tables 1-6, wherein(1) the amino acid sequence of a variable domain of a heavy chain of a human antibody is compared with consensus table 1,(2) the amino acid sequence of a variable domain of a heavy chain of a mouse antibody is compared with consensus table 2,(3) the amino acid sequence of a variable domain of a kappa light chain of a human antibody is compared with consensus table 3,(4) the amino acid sequence of a variable domain of a kappa light chain of a mouse antibody is compared with consensus table 4,(5) the amino acid sequence of a variable domain of a lambda light chain of a human antibody is compared with consensus table 5,(6) the amino acid sequence of a variable domain of a lambda light chain of a mouse antibody is compared with consensus table 6,(C) thereafter producing a modified gene which encodes for a modified antibody variable domain by modifying in the initial gene(1) at least one codon of each pair of codons which encode for cysteine bridges, so that all disulfide bridges present in the initial antibody variable domain are absent from the modified antibody variable domain, and(2) at least one additional codon which codes for an amino acid other than a disulfide bridge-forming cysteine is substituted by another codon using a consensus table selected in part (b) as a guide, wherein(I) the codon coding for the initial amino acid is modified by substituting a codon coding for a substitute amino acid which is listed in the consensus table at a position corresponding to the selected position, when the initial amino acid is unlisted in the selected consensus table at the position corresponding to the selected position, or(II) the codon coding for the initial amino acid is modified by substituting a codon coding for a substitute amino acid having higher frequency in the consensus table at a position corresponding to the selected position, when the initial amino acid is listed a given frequency in the consensus table at the position corresponding to the selected table, the higher frequency being compared to the given frequency;(D) transfecting a eukaryotic cell with the modified gene encoding the modified antibody variable domain; and(E) expressing in the transfected eukaryotic cell the modified antibody variable domain, wherein the modified antibody variable domain expressed by the eukaryotic cell has improved stability when compared to the stability of the initial antibody variable domain.