Figure 1.

Receptor mediated delivery concept. (A) Conjugation scheme. Human holo-transferrin was activated by incubation with SPDP, and conjugates were isolated and incubated with purified ZFN as described in ‘Materials and Methods’ section. A scissile disulfide bond joins the transferrin and the ZFN. (B) Cellular delivery. 1. The transferrin-nuclease complex binds to its cognate receptor, which induces uptake of the cargo to the cell interior in the early recycling endosome. 2. Under the reducing conditions of the endosomal milieu, the ZFN protein is released from the ligand-receptor complex by ‘self-immolation’ of the disulfide bond. 3. ZFN protein escapes the endosome. 4. ZFN protein translocates to the cell nucleus where the two ZFN subunits bind to opposing DNA strands and cleave the target sequence. Pm, plasma membrane; End, endosome; Nuc, nucleus.