Humanisation of monoclonal antibodies Flashcards
monoclonal antibodies made in mice and bacteria ave to be
disguised as human
why do monoclonal antibodies have to be disguised as human
otherwise immune system will recognise them as foreign and destroy them
if you put a mouse antibody into a human
around 5 days after it will be destroyed and excited from the system
two main reasons monoclonal antibodies have to be humanised
1) immunogenicity
2) Origin
immunogenicity issues
most mAbs used in vivo are of mice origin and therefore Xenogenic
due to mAbs being xenogenic (of mouse origin)
human anti-mouse antibody (HAMA) reaction occurs
HAMA
human antibody recognised the mouse antibody as foreign and destroys it
origin of mAbs issue
mouse origin diminishes mAbs ability to elicit effector mechanisms such as complement-mediated lysis or antibody dependent cell-mediated cytotoxicity
if antibody is not humanised
will not elicit complement mediated lysis or antibody dependent cell-mediated cytotoxicity
use of xenoantibodies in human has proven
severely limiting due to them being seen as foreign proteins by the immune system
name 4 strategies used to increase the human content of the mAbs to prevent rejection
- production of human hybridomas
- replacement of the consent regions of mouse mAbs with those of human antibodies
- replacement of CDRs of human antibody with those of a mouse antibody via CDR grafting
4) production of fully humanised mAbs in transgenic mice
Production of human hybridomas
ethical issues and low efficiency
Replacement of the constant regions of mouse mAbs (powerful) with those of human antibodies
chimeric antsy
- leaving important VH/VL parts of the Fab fragment from the the mouse–> however will eventually eb recognised by humans
Replacement of complementarity- determining regions (CDRs) of human antibody with those of a mouse antibody via CDR grafting
makes humanised antibody
CDR grafting
takes a human antibody dn swaps out the CDRs with the CDrs of a mice so that the hyper variable regions are specific for antigen of interest
a set of CDRs constitute a
paratope
antibody expression in chinese hamster ovary cells
1) take mouse monoclonal cell line and extract mRNA for variable domains
clone into mammalian expression vector and produces a full length human antibody
2) take that vector and transfect into a Chinese Hamster Ovary cell (CHO)
3) these CHO will grow in vitro and will produce human antibodies with CDRs of the mouse..
mAbs with Xi in name
chimeric
mAbs with Zu in name
humaniZed
mAbs with Mu in name
fully human
fully human are
phage display
