Epidermal growth factor receptor as a target of antitumor activity of binase mutants

BACKGROUND: The epidermal growth factor receptor (EGFR) is one of the key proteins in cell signaling that regulates mitogen-activated protein kinase (MAPK) pathways. EGFR dysregulation is associated with various neoplasms, highlighting the importance of developing targeted EGFR inhibitors. Binase, a ribonuclease (RNase) from Bacillus pumilus, is a promising antitumor agent capable of interacting with EGFR. Its cytotoxic potential is primarily determined by its activity against intracellular RNA. Binase mutants with reduced catalytic activity, K26A and H101E, also demonstrate antitumor properties.

AIM: This study aimed to assess the effect of interactions between binase mutants and EGFR on their cytotoxic potential.

METHODS: The antiproliferative activity of binase and its mutants K26A and H101E, with residual catalytic activity of 11.0% and 0.02%, respectively, was evaluated using the MTT assay in A431 epidermoid carcinoma cells, which overexpress wild-type EGFR. Immunofluorescence analysis was used to examine the interactions of RNases with EGFR and their ability to modulate the MAPK/ERK pathway. Hypothetical protein–protein interaction models were generated using computational modeling. The antimigratory activity of RNases was assessed using the standard scratch assay.

RESULTS: Binase and its mutants reduced the proliferative activity of A431 tumor cells by 40%. Pretreatment of cells with the monoclonal anti-EGFR antibody cetuximab attenuated the cytotoxic potential of binase mutants. Computational modeling indicated that the tested RNases may interact with EGFR, with binase having a higher affinity for the ATP-binding site of the tyrosine kinase domain and the mutant derivatives binding preferentially to regions involved in receptor endocytosis. This may underlie the observed differences in EGFR internalization rates. Unlike binase and the K26A mutant, the catalytically inactive H101E mutant lacks antimigratory activity, indicating the importance of maintaining a certain level of enzymatic activity.

CONCLUSION: EGFR is a target of the cytotoxic activity of binase mutants, and their interaction inhibits the MAPK/ERK signaling pathway and causes tumor cell death.