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Epidermal growth factor receptor (EGFR) is overexpressed in various tumors and is associated with cancer initiation, progression, and poor prognosis. Despite the achievements made by tyrosine kinase inhibitors and monoclonal antibodies in certain cases, many patients have not benefited from such treatment due to resistance. Immunotoxins (ITs) are antibody‑cytotoxin chimeric molecules with specific cell killing ability, which have achieved different degrees of success in the treatment of a wide range of cancers in clinical trials. The aim of the current study was to examine a novel targeting EGFR recombinant immunotoxin Bs/cucurmosin (CUS) generated by fusing CUS to the EGFR‑specific nanobody 7D12‑9G8. Bs/CUS was successfully expressed in Escherichia coli strain BL21 (DE3) in a soluble form. Furthermore, it retained binding capacity and specificity with EGFR and was superior to rE/CUS, a monospecific IT we reported previously. In vitro results showed that Bs/CUS could be internalized into the cytoplasm and selectively kill cells in the picomolar range. Flow cytometry showed that Bs/CUS killed the cells mediated by the apoptosis pathway. Taken together, results of the current study indicated that Bs/CUS is a promising candidate that should be further evaluated as a cancer therapeutic for the treatment of EGFR‑positive tumors.

Novel EGFR‑bispecific recombinant immunotoxin based on cucurmosin shows potent anti‑tumor efficiency in vitro