Design Of TNF-α Antagonists Based On The Interaction Between TNF-α And Neutralizing Monoclonal Antibody (Z12)

Tumor necrosis factor-α (TNF-α) is a multifunctional cytokine that plays a central role in infection, inflammation and immune regulation. While normal level of TNF-a is very important to immune homeostasis, overproduction of TNF-a can result in significant pathological changes in diseases including sepsis, inflammatory bowel disease, rheumatoid arthritis, and Crohn's disease, etc. Neutralization of excessive TNF-a has become an effective therapeutic strategy. In this study, three novel strategies were used to design TNF-a inhibitors under the computer-aided guidance.Firstly, one peptide (PT4) was chosen from previously designed peptides and linked with FC fragment of human IgG1. The interaction mode of PT4-linker-FC (abbr. PLF) with TNF-a was analyzed with computer-guided molecular modeling method. After expression in E.coli and purification, recombinant PT4-linker-FC could bind directly with TNF-a coated on the ELISA plates. Furthermore, PLF could competitively inhibit the binding of Z12 to TNF-a and also inhibit the TNF-a-induced cytotoxicity on L929 cells. The TNF-a antagonizing activity of PLF was significantly higher than the free peptide.Secondly, Frameworks of heavy chain variable region (VH) of human antibody were used as scaffolds to display three peptides (PT2, PT3 and PT4) simultaneously. Complementarity-determining regions (CDRs) in VH were substituted with PT2, PT3 and PT4 derived from previously designed peptides, respectively. New engineered antibody-like molecule was called PTVH. Seven PTVHs were constructed when seven...