Immunologic Function and Molecular Insight of Recombinant Interleukin-18

Area of Science:

• Immunology
• Biotechnology
• Molecular Biology

Background:

• Cytokine-mediated therapy is an advancing alternative in cancer treatment.
• Interleukin-18 (IL-18) demonstrates anti-cancer potential, particularly when combined with IL-12.
• Engineering IL-18 aims to improve its therapeutic efficacy.

Purpose of the Study:

• To enhance the anti-cancer activity of Interleukin-18 (IL-18) through single point mutagenesis.
• To investigate the impact of specific mutations (M33Q, M60Q, E6K, T63A) on IL-18's structure and function.
• To evaluate the synergistic effects of combined mutations on IL-18's ability to stimulate interferon-gamma (IFN-γ) production.

Main Methods:

• Single point mutations were introduced into the IL-18 gene based on binding residues and polarity.
• Mutated IL-18 proteins were expressed in Pichia pastoris and purified.
• The activity of IL-18 variants was assessed by measuring IFN-γ stimulation in the NK-92MI cell line.
• Molecular dynamics simulations were employed to analyze the structural effects of mutations.

Main Results:

• The E6K and T63A IL-18 mutants exhibited significantly higher IFN-γ induction activity compared to native IL-18.
• The combined E6K and T63A mutations demonstrated a synergistic effect, increasing IFN-γ induction by 16-fold.
• Mutations M33Q and M60Q did not show significant IFN-γ inducing activity.
• Molecular dynamics revealed that E6K enhances structural stability, T63A increases flexibility, and M33Q/M60Q cause substantial conformational changes.

Conclusions:

• Engineered IL-18 variants, particularly E6K and T63A, show enhanced potential for cancer therapy.
• The synergistic combination of E6K and T63A mutations offers a novel strategy for potent cytokine-mediated therapy.
• These findings introduce a new version of IL-18 as a promising candidate for advanced cancer treatment.