IL-6 Antagonist Designed Rationally On The Conformational Information Of Ligand And Receptor

Since the three-dimensional structure of IL-6 has been determined by X-ray crystallographic analysis at 1997, researchers have made much progress in the study of human IL-6 antagonist. However, the effect of their clinical applications is limited because of it's non-specificity and poor effect to inhibit the activity of human IL-6 in vivo. Based on the three-dimensional complex structure of IL-6/sIL-6R/gp130 and the interaction pattern of human IL-6 and its receptor (i.e. EL-6R. gp130) derived from distance geometry, inter-molecular hydrogen bond forming theory, the interleukin-6 antagonist peptide was designed rationally by the means of reasonable screening and computer-guided molecular design. The reasonable plan how to design ligand antagonist based on the complex structural information of the ligand and receptor was present. Furthermore, aimed on designed IL-6 antagonist peptide, the biological activity was evaluated through experiment in vitro. The study highlights to provide more effective leading compound for clinical therapy of IL-6 related disease. The outline of the study was following:1. The IL-6 antagonist peptide designed rationallyBased on the three-dimensional complex structure of IL-6/sIL-6R/gp130,dealing with the technology of Computer Graphics, distance geometry and the intermolecular hydrogen bond forming theory, the binding domain between the ligand (i.e. hIL-6) and it's receptor (i.e. hIL-6R and gpl3O) was determined. The structural information of the functional domain and the interaction mode were analyzed theoretically. The IL-6 antagonist peptide was designed using site-linking method. The spatial conformation of the antagonist peptide was modeled using ab initio method. The complex structure of the antagonist peptide and ML-6R was constructed using molecular docking and optimized with molecular mechanism and dynamics. The interaction mode between the antagonist and hIL-6R was analyzed and the bioactivity of the antagonist peptide was evaluated theoretically.2. Experimental evaluation on the biological function of the antagonist peptide.(D Dealing with MTT and 3H-Tdr methods, the activity of IL-6 antagonist peptide was tested. XG-7 cell, the interleukin-6 dependent cell line, was chosen to examine the peptide function on proliferation. And then the influence on hIL-6 was studied. The results showed that the proliferation of XG-7 cell was stimulated by IL-6 could be effective inhibited by the antagonist peptide. It revealed that the antagonist peptide could inhibit EL-6 biological function at cellular level.(2) The activity of the IL-6 antagonist peptide on how to inhibit IL-6/IL-6R binding with binding assay was studied. The results showed that the peptide could compete IL-6 to bind with IL-6R specifically.(3) Based on the immunohistochemical staining method (Alkaline phosphatase-anti-alkaline phosphatase technique, APPAP), the effect whether the antagonist peptide influenced the cellular signal transduction of EL-6 was studied. Chosen human hepatoma cell line HepG2 induced with hIL-6, different concentrations of IL-6 antagonist peptide were added. With immunohistochemical staining on HepG2 cell, the results showed that the quantity of stained cells and the intensity of stained cells were decreased with the increasing concentrations of...