Production and functional characteristics of the recombinant targeted toxin based on the HER2-specific non-immunoglobulin scaffold

Designing of new high-affinity protein compounds that can selectively destroy human cancer cells is a priority research area in biomedicine. In this work we report on the production procedure and describe functional characteristics of genetically encoded targeted toxin DARPin-PE40 for selective destruction of HER2-positive human cancer cells. HER2-specific non-immunoglobulin scaffold protein with ankyrin repeat was used as a targeting moiety. As a cytotoxic module, we used a fragment of the Pseudomonas exotoxin A. Using cytoplasmic expression in a bacterial system, we produced a monomelic protein with the yield of 180 mg per 1 L of culture medium. It was shown that targeted recombinant toxin possesses a considerable cytotoxic activity in vitro against HER2-positive cells (IC₅₀ 2 pM).