Discovering HIL-6 Antagonist From Synthetic Compound Database By Using Structure-Based Virtual Screening

Human interleukin-6 (hIL-6) is a pleiotropic cytokine, which plays an important role in hematopoietic development, on the immune system and on acute phase reactions. It can exert multi-bioactivities by binding to its receptor on the cell surface. However, the overproduction of hIL-6 and its receptor (hIL-6R) is considered to involve in the development of autoimmune disorders and lymphoid malignancies. So, it is very useful in clinical to discover the antagonist of hIL-6. Now, the researchers are focusing on generating mono-antibody and mutant to block the bioactivity of hIL-6, but few researches about the small molecular antagonists targeted to hIL-6 have been presented. In this research work, based on the 3D (three-dimension) structure of hIL-6 and the interaction information between hIL-6 and hIL-6R, a series of the small molecular antagonists targeted to hIL-6 were screened out, optimized and evaluated using computer-guided virtual screening approach and the bioactivities of the small molecular compounds were analyzed.On the basis of a three-dimensional crystal complex model of (hIL-6hIL-6R·gp 130)₂, molecular docking-based virtual screening was employed to search the chemical databases: ACD (Available Chemicals Directory) and MDDR (MDL Drug Data Report). Compounds were ranked according to their relative binding energy, favorable shape complementarities, potential to form hydrogen bonds with the outer mouth of the hIL-6·hIL-6R model. Then the biological functions of candidate compounds were examined using competition ELISA. XG-7 and 7TD1 cells, two IL-6 dependent cell lines, were chose to evaluate the inhibitory ability on cell proliferation and the effect on cell apoptosis.Three candidate compounds were selected out from the virtual screening. The theoretical studies showed that these compounds offered a practical means of imposing long-term blockade of hIL-6 activity and possessed very high affinity to hIL-6R in vitro. ELISA experiments demonstrated that the three compounds all could block IL-6 to bind with IL-6R specifically. But in cellular level, only compound A could inhibit the proliferation of XG-7 cells, and antagonized the function of hIL-6 in a dose-dependent manner.Considering compound A only could partially antagonize the bioactivity of hIL-6 with the inhibitory rate of 20% at 1μg/ml, we redesigned the antagonist base on the structure of compound A to obtain the novel compound with higher antagonistic activity to hIL-6. Then, compound D with new structure was selected out, which acted as an excellent blocker. Compound D had the ability of interfering with the function of hIL-6, inducing growth arrest and apoptosis on XG-7 and 7TD1 cells. The inhibitory rate of compound D (1μg/ml) reached to 30.4% on XG-7 cell and 28.1% on 7TD1. And it showed no cytotoxicity to SP2/0 and L929 cells in dosage range (from 1 ng/ml to 1μg/ml). All the results demonstrated that the antagonist D, from computer-guided screening and rational design, could inhibit the function of hIL-6 effectively.By structure-based virtual screening and rational drug design, we obtained compound D as the lead compound of small molecular hIL-6 antagonist. The virtual screening approach and evaluating platform may do much help to seek more effective novel small molecular drugs for therapeutic use in hIL-6-associated diseases.