Humanisation (also called Reshaping or CDR-grafting) is now a
well-established technique for reducing the immunogenicity of
monoclonal antibodies (mAbs) from xenogeneic sources (commonly rodent) and
for improving their activation of the human immune system; in fact
there are many humanised mAbs in clinical trials and a few have been
given approval to be used as drugs. Although the mechanics
of producing the engineered mAb using the techniques of molecular
biology are relatively straightforward,
simple grafting of the rodent complementarity-determining regions (CDRs) into human frameworks does not
always reconstitute the binding affinity and specificity of the
original mAb. In order to humanize an antibody, the design of the humanized antibody is now the critical
step in reproducing the function of the original molecule.
This design includes various choices: the extents of the
CDRs, the human frameworks to use
and the substitution of residues from the rodent mAb into the
human framework regions (backmutations).
The positions of these backmutations have been
identified principally by sequence/structural analysis or by analysis of an
homology model of the variable regions' 3D structure. Recently, phage
libraries have been used to vary the amino acids at chosen positions.
Similarly, many approaches have been used to choose the most
appropriate human frameworks in which to graft the rodent CDRs. Early
experiments used a limited subset of well-characterised human
mAbs (often where the structure was available), irrespective of the sequence identity to the rodent mAb
(the so-called fixed frameworks approach). Some groups use variable
regions with high amino acid sequence identity to the rodent variable
regions (homology matching or best-fit); others use consensus or germline
sequences while still others select fragments
of the framework sequences within each light or heavy chain variable
region from several different human mAbs. There are also
approaches to humanisation developed which replace the surface rodent
residues with the most common residues found in human mAbs
("resurfacing" or "veneering") and those which use differing definitions of
the extents of the CDRs.
However, some rodent mAbs have proved difficult to humanise using current protocols (Pichla et al (1997) J. Struct. Biol. 119:6-16). The
design and engineering of humanised mAbs are still very much areas
of research, as much for the light they shed on protein structure and function
as well as for the potential therapeutic and diagnostic benefits.
The accompanying web pages supply data and raise
design issues to help the prospective antibody designer through the bewildering variety of choices.
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list of humanised antibodies taken from the
literature. Please email
any queries or problems by clicking on my name at the bottom of the page.