Design and Characterization of Ricin Based Immunotoxins Against EPHA2 Receptor for Breast Cancer Therapy: An
Atefeh Faraz1, Jafar Amani2, Sedigheh Arbabian1
1Department of Cellular and Molecular Biology, Faculty of Biological Science, North Tehran Branch, Islamic Azad University, Tehran, Iran.
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.
Insights
Two novel immunotoxins targeting breast cancer
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Immunotoxins represent a promising strategy for cancer therapy.
- The EphA2 receptor is overexpressed in breast cancer, making it a potential therapeutic target.
Purpose of the Study:
- To design and computationally evaluate two novel immunotoxins targeting the EphA2 receptor for breast cancer treatment.
- To assess the stability, binding affinity, and safety profiles of the designed immunotoxins.
Main Methods:
- Constructing immunotoxins by fusing anti-EphA2 single-chain variable fragments (scFv) with ricin A chain.
- Utilizing bioinformatics tools (mFold, HADDOCK, Gromacs) for mRNA stability, protein structure prediction, molecular docking, and molecular dynamics simulations.
- Evaluating physicochemical properties, allergenicity, and antigenicity of the designed constructs.
Main Results:
- The designed immunotoxins exhibited stable mRNA and reliable protein structures.
- Both constructs demonstrated high affinity and stable interactions with the EphA2 receptor, with construct 2 showing superior binding.
- The immunotoxins were found to be non-allergenic and antigenic.
Conclusions:
- The designed immunotoxins show potential as therapeutic agents for breast cancer.
- Construct 2 displays enhanced properties, suggesting it as a more promising candidate for further empirical investigation.
- These findings support the use of these immunotoxins in preclinical studies for breast cancer treatment.
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