Transient Expression of an IL-1Ra−Fc Fusion Cytokine in Tobacco Species for Improved Anti-inflammatory Therapeutic Potential

Abstract The interleukin-1 receptor antagonist (IL-1Ra) is a key anti-inflammatory cytokine that inhibits interleukin-1 signaling by competitively binding to IL-1 receptors. However, recombinant IL-1Ra, currently produced in Escherichia coli, exhibits limited pharmacokinetics and requires frequent administration due to its short plasma half-life and absence of glycosylation. To overcome these limitations, we designed a novel IL-1Ra-Fc fusion protein consisting of human IL-1Ra fused to a human IgG1 Fc fragment via a flexible linker and transiently expressed in Nicotiana benthamiana to enhance protein yield, stability, and enable glycosylation. The recombinant protein accumulated to approximately 340 mg/kg fresh leaf biomass and achieved an overall DSP recovery of 45% protein yield, with a characteristic high α-helical secondary structure, as confirmed by CD spectroscopy. Receptor-binding analysis shows that rhIL-1Ra-Fc exhibited significantly enhanced IL-1 receptor binding activity, showing approximately 17% higher activity than that of commercial IL-1Ra. Overall, this is the first study to demonstrate the feasibility of producing functionally active IL-1Ra-Fc in plants for potential therapeutic applications in inflammatory diseases.