The receptor for an epidermal growth factor (EGFR) is an attractive target for cancer immunotherapy, as it is a tyrosine kinase receptor overexpressed on tumor cells with a key role in the development of malignancy. Cetuximab (Cet) is an EGFR inhibitor medication used for metastatic colorectal cancer (CRC); however, its therapeutic efficacy is limited due to resistance mechanisms in some patients. The current study aimed to provide a novel immune-RNase based on the conjugation of Cet with RNase enzyme as an anti-EGFR biosystem against CRC.

A novel immuno-RNase, called anti-EGFR antibody-RNase A (Cet-RNaseA), is produced from the conjugation of anti-EGFR antibody Cet and bovine pancreatic RNase (RNase A). After synthesizing Cet-RNaseA, the physicochemical properties of Cet-RNaseA were characterized by SDS-PAGE, BCA kit, and UV-vis spectrum. Furthermore, its biological impacts, including cell viability, apoptosis, and ERK1, ERK2, Bax, and Bcl2 gene expression, were evaluated in the KRAS mutant SW-480 cells.

Cet-RNaseA retained the enzymatic activity of RNase A and was specifically bound to EGFR-positive cells with an affinity comparable with the free RNase A and Cet. In addition, the maximum anti-apoptosis effect was obtained after treating cells with an IC50 dose of Cet-RNase A compared to free RNase A and Cet. Based on real-time PCR data, in the treated KRAS mutant SW-480 cells with the Cet-RNase A, the expression level of Bax, ERK1, and ERK2 were significantly increased, while the expression of Bcl2 was significantly decreased compared to the untreated control cells.

The novel immune-RNase showed an effective and targeted antiproliferative activity against EGFR-positive CRC cells, which was more potent than the free RNase A and Cet alone. In conclusion, Cet-RNase A could be considered a promising candidate for the immunotherapy of EGFR-positive tumors.