Antibody Name: BR96 M1
References: Rosok, M.J., Yelton, D.E., Harris, L.J., Bajorath, J., Hellstrom, K-E., Hellstron, I., Cruz, G.A., Kristensson, K., Lin, H., Huse, W.D. and Glaser, S.M. "A combinatorial library strategy for the rapid humanization of anticarcinoma BR96 Fab" J. Biol. Chem. (1996) 271:22611-22618.
Donor Antibody: mouse
Acceptor Antibody: Human Fabs
Antigen: Tumour-associated antigen expressing Lewis Y related carbohydrate.
Laboratory: Bristol-Myers Squibb, WA; Ixsys, CA
Design: M13 phage Fab expression library containing combinations of human and murine residues at several positions chosen by consideration of canonical residues, a model of the Fv region and buried residues as defined by Padlan (1991). Library incorporated mouse/human alternatives at the following positions:
VH 24 49
VL 11 36 37 43 104
Frameworks: germline FRs 1,2,3 VH HSIGDP51 SGIII/JH4 | VL HSIGVA23 SGII/Jk2
CDRs: All 6 VH and VL, Kabat Definition except P60A and T62S in CDR-H2.
Backmutations: I2V:L (canonical but not necessary) L36Y:L P43S:L
Binding: Humanised shows 2-fold less affinity compared to mouse mAb by surface plasmon resonance (SPR).
Expression: E.coli strains XL-1 and MK30-3. Better expression of humanised than mouse Fabs.
Comment: Taq polymerase introduced PCR error. Now experimenting with Pfu and Vent. Compared their humanisation with that of B3 and found none of the latter's backmutations had any effect on BR96.
Clinical Indication: human carcinomas

explanation of symbols and abbreviations

José Saldanha © 1997-8. Birkbeck College, London WC1E 7HX.