GENERATION AND CHARACTERIZATION OF A MOUSE HUMAN CHIMERIC ANTIBODY-DIRECTED AGAINST EXTRACELLULAR-MATRIX PROTEIN TENASCIN/

The murine anti-tenascin monoclonal antibody 81C6, following iodinatio n, has been shown to be an efficient localizing and therapeutic agent in both subcutaneous and intracranial human glioma xenograft models in athymic mice and rats. Similarly, effective monoclonal antibody 81C6 localization has been demonstrated in glioma patients, and Phase I tri als with the intact murine IgG(2b) kappa molecule are currently in pro gress. In order to maximize the potential for repeated administration by minimizing murine Fc-mediated immunogenicity and reducing Fc-mediat ed immune effects, we created murine 81C6 variable region/human IgG(2) chimeric monoclonal antibodies by the molecular cloning of the variab le region genes of mouse 81C6 and their genetic linkage to human const ant region exons. The resulting chimeric constructs were introduced in to SP2/0 cells, and stable transfectomas were selected by G418 and myc ophenolic acid resistance. The resistant clones were screened for anti -tenascin activity on tenascin-coated plates by enzyme-linked immunoso rbent assay. The N-terminal amino acid sequence of both heavy and ligh t chains of the purified chimeric 81C6 antibody matched exactly with t hat of the native mouse 81C6 as well as with that deduced from the nuc leotide sequence. The production level of chimeric 81C6 (13.9 mg/ml) f rom ascites in the highest expressing transfectoma was much higher tha n that of native mouse 81C6 (2.5 mg/ml). The chimeric antibody showed the same specificity and equivalent affinity for human intact tenascin or tenascin-expressing cells as the native mouse 81C6 antibody. Direc t comparison of radioiodinated chimeric and radioiodinated mouse 81C6 biodistribution in subcutaneous and intracranial xenograft-bearing mic e showed higher tumor-to-normal tissue ratios for chimeric 81C6 as com pared with native mouse 81C6. The improved localizing and clearance ch aracteristics of chimeric 81C6 in xenograft model systems suggests tha t chimeric 81C6 would be an improved reagent for intracompartmental th erapy of tenascin-expressing tumors in the human central nervous syste m.