Selection of tumor-specific internalizing human antibodies from phage libraries

Antibody internalization into the cell is required for many targeted therap eutics, such as immunotoxins, immunoliposomes, antibody-drug conjugates and fur targeted delivery of genes or viral DNA into cells. To generate direct ly tumor-specific internalizing antibodies, a non-immune single chain Fv (s cFv) phage antibody library was selected on the breast tumor cell line SKBR 3. Internalized phage were recovered from within the cell and used for the next round of selection. After three rounds of selection, 40% of clones ana lyzed bound SKBR3 and other tumor cells but did not bind normal human cells . Of the internalizing scFv identified, two (F5 and C1) were identified as binding to ErbB2, and one (H7) to the transferrin receptor. Both F5 and H7 scFv were efficiently endocytosed into SKBR3 cells, both as phage antibodie s and as native monomeric scFv. Both antibodies were able to induce additio nal functional effects besides triggering endocytosis: F5 scFv induces down stream signaling through the ErbB2 receptor and H7 prevents transferrin bin ding to the transferrin receptor and inhibits cell growth. The results demo nstrate the feasibility of selecting internalizing receptor-specific antibo dies directly from phage libraries by panning on cells. Such antibodies can be used to target a variety of molecules into the cell to achieve a therap eutic effect. Furthermore, in some instances endocytosis serves as a surrog ate marker for other therapeutic biologic effects, such as growth inhibitio n. Thus, a subset of selected antibodies will have a direct therapeutic eff ect. (C) 2000 Academic Press.