INTRACELLULAR EXPRESSION OF SINGLE-CHAIN ANTIBODIES REVERTS ERBB-2 TRANSFORMATION

We report a novel approach for specific in vivo inactivation of the Er bB-2 receptor tyrosine kinase and suppression of ErbB-2-induced transf ormation. Genes encoding single chain antibodies that specifically bin d to the extracellular domain of human ErbB-2 were constructed and exp ressed intracellularly in NIH/3T3 fibroblasts transformed by activated ErbB-2. The single chain antibodies are derived from monoclonal antib odies FRP5 and FWP51 (Harwerth, I. M., Wels, W., Marte, B. M., and Hyn es, N. E. (1992) J. Biol. Chem. 267, 15160-15167) and are composed of heavy and light chain variable domains connected by a flexible peptide linker. The antibodies were provided with: 1) an N-terminal hydrophob ic leader sequence to target their synthesis to the lumen of the endop lasmic reticulum, and 2) a C-terminal retention signal to prevent secr etion. When expressed in ErbB-2-transformed cells, the single chain an tibodies bound to the receptor and prevented its transit through the e ndoplasmic reticulum. This resulted in the functional inactivation of the receptor and reversion of the transformed phenotype. This is the f irst demonstration of a targeted and stable inactivation of a cellular onco-protein via intracellular antibody expression. The use of such a strategy represents a simple and powerful approach to study the in vi vo function of receptors and other cellular proteins.