EFFECT OF VK FRAMEWORK-1 GLYCOSYLATION ON THE BINDING-AFFINITY OF LYMPHOMA-SPECIFIC MURINE AND CHIMERIC LL2 ANTIBODIES AND ITS POTENTIAL USE AS A NOVEL CONJUGATION SITE

A potential asparagine (Asn)-linked glycosylation site was identified in the VK FRI sequence of an anti-B lymphoma monoclonal antibody (MAb) , LU. SDS-PAGE analysis and endo-F treatment of both murine and chimer ic LL2 antibodies indicated that this site was glycosylated; however, no differences in the binding affinity to Raji cells were observed bet ween the native murine LU and the endo-F-deglycosylated murine LL2 ant ibodies. Elimination of the glycosylation site from the chimeric LU an tibody was accomplished by an Asn to Gin mutation in the tri-acceptor site found in the light chain. The resultant aglycosylated chimeric LL 2 exhibited a similar Raji cell binding affinity to that of the glycos ylated form. The results are in agreement with computer modeling studi es which suggested the lack of interactions between the oligosaccharid e moiety and the CDRs. The finding is interesting because it enables a wider choice of human framework sequences, which in most cases do not have a corresponding glycosylation site, for the humanization of the LU VK domain, as well as a greater latitude of host expression systems . Most importantly, the LL2 VK carbohydrate moiety might be used as a novel conjugation site for drugs and radionuclides without compromisin g the immunoreactivity of the antibody. (C) 1995 Wiley-Liss, Inc.