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Immunotoxins are chimeric molecules, which combine antibody specificity to recognize and bind with high-affinity tumor-associated antigens (TAA) with the potency of the enzymatic activity of a toxin, in order to induce the death of target cells. Current immunotoxins present some limitations for cancer therapy, driving the need to develop new prototypes with optimized properties. Herein we describe the production, purification and characterization of two new immunotoxins based on the gene fusion of the anti-carcinoembryonic antigen (CEA) single-chain variable fragment (scFv) antibody MFE23 to α-sarcin, a potent fungal ribotoxin. One construct corresponds to a conventional monomeric single-chain immunotoxin design (IMTXCEAαS), while the other one takes advantage of the trimerbody technology and exhibits a novel trimeric format (IMTXTRICEAαS) with enhanced properties compared with their monomeric counterparts, including size, functional affinity and biodistribution, which endow them with an improved tumor targeting capacity. Our results show the highly specific cytotoxic activity of both immunotoxins  in vitro , which was enhanced in the trimeric format compared to the monomeric version. Moreover, the trimeric immunotoxin also exhibited superior antitumor activity  in vivo  in mice bearing human colorectal cancer xenografts. Therefore, trimeric immunotoxins represent a further step in the development of next-generation therapeutic immunotoxins.

scientific reports

A novel Carcinoembryonic Antigen (CEA)-Targeted Trimeric Immunotoxin shows significantly enhanced Antitumor Activity in Human Colorectal Cancer Xenografts