AN AUTOANTIBODY IS MODIFIED FOR USE AS A DELIVERY SYSTEM TO TARGET THE CELL-NUCLEUS - THERAPEUTIC IMPLICATIONS

A murine monoclonal anti-dsDNA antibody was found to penetrate living cells and localize in the nucleus without pathologic effects. A single mutation in V-H markedly enhanced cellular penetration. The mutant an tibody was produced as recombinant Fab and single chain antibody fragm ents to investigate its use as a delivery system to target the cell nu cleus. Complexes were made containing Fab fragments and alkaline phosp hatase conjugated goat antibodies to mouse kappa chains. Fab fragments transported 305 kDa goat antibody-enzyme complexes into the nucleus i n COS-7 and CHO cells. A single chain antibody cDNA was constructed by splice overlap extension PCR and expressed in COS-7 cells. Binding of the single chain antibody to dsDNA was shown by ELISA, and cellular p enetration and nuclear localization were demonstrated in COS-7 and CHO cells. The single chain antibody cDNA was ligated into the expression vector, pEGFP, to produce a fusion protein with green fluorescent pro tein. The fusion protein penetrated COS-7 cells and localized in the c ell nucleus. The single chain antibody produced during sustained expre ssion in CHO cells re-entered antibody-producing cells and localized i n the nucleus without affecting cell viability. Our results demonstrat e the potential use of a modified autoantibody as a delivery system to target the cell nucleus. (C) 1998 Academic Press