Antineoplastic effect of anti-erbB-2 intrabody is not correlated with scFvaffinity for its target

Intracellular single-chain antibodies (scFvs) have emerged as a powerful me thod to knock out expression of oncoproteins. We have demonstrated previous ly that scfvs directed against a variety of molecular targets induce specif ic toxicity in tumor cells. Recently, the utility of an anti-erbB-2 scFv ha s predicated its evaluation in a phase I gene therapy clinical trial. The u tility of scFv as an intrabody is closely linked to its interaction with a target, Limiting the contribution of the latter to the neoplastic phenotype . In this study, we sought to determine whether improvement in the affinity of the scFv far its cognate target could improve the efficiency of intrabo dy-mediated oncoprotein knockout. We compared in erbB-2-positive and -negat ive tumor cells the function of plasmids encoding a newly developed C6.5 an ti-erbB-2 scFv, which has a 1000-fold higher affinity, with our original e2 3 anti-erbB-2 scFv. Intracellular scFv expression, target binding, and tumo r cell cytotoxicity were found to be similar in all conditions tested, incl uding dose-response studies with limiting dilutions of the scFv. On this ba sis, we have concluded that the antineoplastic effect of anti-erbB-2 intrab ody is not correlated with scFv affinity for its target.