Screening Of HDHODH Inhibitors And Crystal Structures Of HDHODH-Inhibitor Complex

Autoimmune diseases have greatly affected people’s lives, such as rheumatoid arthritis, systemic lupus erythematosus, organ transplant rejection and so on. At present, glucocorticoid, microbial metabolites, antimetabolites, alkylating agents have been widely used as immunosuppressants in clinical applications. But most of them have side effects at different degrees, which limited their clinical applications, so nowadays the development of efficient, low toxic, targeted small molecule inhibitors to apply for the treatment of autoimmune diseases has become a research hotspot.Dihydroorotate dehydrogenase (DHODH) is a key enzyme of the de novo pyrimidine synthesis. Inhibitors of DHODH could inhibit pyrimidine synthesis, thereby blocking DNA and RNA synthesis, so DHODH has been an appealing drug target to treat some autoimmune diseases. Currently, leflunomide is a well-known inhibitor of human dihydroorotate dehydrogenase (hDHODH) and has been widely used in the treatment of rheumatoid arthritis, but long-term use of the leflunomide will cause headaches, nausea, rashes and other side effects, therefore, we hope to discover new inhibitors of hDHODH with more efficiency and less side effects as drug candidates.In this study, we obtained lead compound b 1 as hDHODH inhibitor by virtual screening, with the half maximal inhibitory concentration (IC50) equal to 0.365μM; we solved the complex crystal structure of hDHODH with inhibitor b1, the PDB ID was 4LS0; we designed and synthesized 25 derivatives of compound b1 based on the complex structure of compound bl-hDHODH; and we evaluated all of these compounds, among which compounds b13 and b24 showed highest inhibition to hDHODH with double-digit nanomolar, the values of IC50 were 32 nM and 43 nM, respectively; we determined another two complex crystal structures of b13-hDHODH and b21-hDHODH, the PDB ID were 4LS1 and 4LS2, respectively. From these three crystal structures solved in our study, we found water molecules played an important role in facilitating inhibitors binding to the pocket of hDHODH by generating a strong water-mediated hydrogen bond network between inhibitors and hDHODH.In summary, we designed two inhibitors of hDHODH with double-digit nanomolar inhibition activity based on the X-ray crystal structure of hDHODH in complex with lead compound b1; moreover, the three complex crystal structures solved in this study demonstrated that hydrogen bond network interactions played a pivotal role in drug design and optimization.