CONSTRUCTION AND CHARACTERIZATION OF A HUMANIZED, INTERNALIZING, B-CELL (CD22)-SPECIFIC, LEUKEMIA LYMPHOMA ANTIBODY, LL2/

The murine monoclonal antibody, LL2, is a B-cell (CD22)-specific IgG2a which has been demonstrated to be clinically significant in the radio immunodetection of non-Hodgkin's B-cell lymphoma. The antibody carries a variable region-appended glycosylation site in the light chain, and is rapidly internalized upon binding to Raji target cells. Humanizati on of LL2 was carried out in order to develop LL2 as a diagnostic and immunotherapeutic suitable for repeated administration. Based on the e xtent of sequence homology, and with the aid of computer modeling, we selected the EU framework regions (FR) 1, 2 and 3, and the NEWM FR4 as the scaffold for grafting the heavy chain complementarity determining regions (CDRs), and the REI FRs for that of light chains. The light c hain glycosylation site, however, was not included. Construction of th e CDR-grafted variable regions was accomplished by a rapid and simplif ied method that involved long DNA oligonucleotide synthesis and the po lymerase chain reaction (PCR). The humanized LL2 (hLL2), lacking light chain variable region glycosylation, exhibited immunoreactivities tha t were comparable to that of chimeric LL2 (cLL2), which was shown prev iously to have antigen-binding properties similar to its murine counte rpart, suggesting that the VK-appended oligosaccharides found in mLL2 are not necessary for antigen binding. Moreover, the hLL2 retained its ability to be internalized into Raji cells at a rate similar to its m urine and chimeric counterparts.