Immunotoxins: the role of the toxin

Abstract

Immunotoxins are antibody-toxin bifunctional molecules that rely on intracellular toxin action to kill target cells. Target specificity is determined via the binding attributes of the chosen antibody. Mostly, but not exclusively, immunotoxins are purpose-built to kill cancer cells as part of novel treatment approaches. Other applications for immunotoxins include immune regulation and the treatment of viral or parasitic diseases. Here we discuss the utility of protein toxins, of both bacterial and plant origin, joined to antibodies for targeting cancer cells. Finally, while clinical goals are focused on the development of novel cancer treatments, much has been learned about toxin action and intracellular pathways. Thus toxins are considered both medicines for treating human disease and probes of cellular function.

Figure 1

Graphic representations of three toxins, diphtheria toxin (DT), Pseudomonas exotoxin (PE) and the plant toxin, ricin. Above each “domain” is a functional label. Below each domain is a common name that was used in early publications. DT has an N-terminus catalytic domain (C-domain) also known and the A fragment followed by a protease processing site, then a nine helix domain (commonly known as the “T” or translocation-domain) followed by a receptor binding domain (R-domain). The B-fragment includes both the T-domain and the R-domain. PE has an N-terminal receptor-binding domain followed by a processing domain. Then at the C-terminus there is a catalytic domain followed by a KDEL-like sequence. Ricin has a catalytic domain at the N-terminus, followed by a processing site and then a duplicated receptor-binding domain with a preference for binding galactose residues. Each toxin has a helical domain where several helices follow in close sequence. For DT there are nine helices while PE has 6; and these helices are arranged in what appears to be a separate domain between C and R-domains. Ricin also has a cluster of helices but these are located in the middle of its catalytic domain. A simple view of these helical domains is that they function in the translocation of each toxin’s C-domain. However, this has only been established for the T-domain of DT. The site of proteolytic processing is shown for each toxin.

Figure 2

Immunotoxin construction-from oldest to newest. First generation immunotoxins were constructed by using chemical crosslinking agents to attach intact toxins to intact antibodies. Second generation immunotoxins used modified toxins lacking receptor-binding domains. Third generation molecules used cloned antibody fragments fused to modified toxin genes; allowing for the recombinant production of homogeneous protein. Further improvements of the third generation molecule might include the removal of immunogenic amino acids including (as shown) much of the multi-helical domain of PE.