Design and Characterization of Ricin Based Immunotoxins Against EPHA2 Receptor for Breast Cancer Therapy: An In-Silico Study

Abstract

Background: One of the most promising strategies to combat cancer is the use of immunotoxins. Objectives: This study aimed to design two immunotoxins composed of antibody fragments against the EphA2 receptor, which is highly expressed in breast cancer. Methods: EphA2-N-ricin and EphA2-C-ricin were designed by fusing scFv against the EphA2 receptor with the A chain of ricin in varying orders. mFold was used to analyze the mRNA stability of the constructs. The 2D and 3D protein structures of the constructs were predicted using prediction tools and verified by quality assessment tools. The physicochemical properties were calculated using ProtParam. Docking between the constructs and the EphA2 receptor was performed using HADDOCK software, and the 2D interaction plots of the complexes were generated using LigPlus. A 100 ns molecular dynamics (MD) simulation was conducted for docked complexes using Gromacs. Ultimately, the allergenicity and antigenicity of the constructs were determined. Results: The designed immunotoxins had stable mRNAs, reliable 2D and 3D protein structures, and demonstrated high affinity and stable interactions with the receptor protein, as revealed by docking and MD analyses. Higher binding affinity and stability were observed for construct 2. Moreover, the designed immunotoxins lacked allergenicity and were identified as antigens. Conclusions: Based on these observations, it is reasonable to conclude that both designed immunotoxins could serve as suitable immunotoxins; however, construct 2 exhibits more promising properties. Given these results, these immunotoxins could be used in empirical studies to treat breast cancer in vitro or in vivo.