Identification Of a Novel Small-Molecule TNFα Inhibitor With Activity Against Inflammation In a Hepatitis Mouse Model

Over-expression of tumor necrosis factor α (TNFα) is a hallmark of many inflammatory diseases including rheumatoid arthritis, inflammatory bowel disease and septic shock and hepatitis, making it a potential therapeutic target for clinical interventions.

To date, significant advances have been made in the development of biological agents targeting TNFα and its signaling components. There are several well known commercial TNFα inhibitors, such as infliximab, adalimumab and etanercept, all of which are TNFα antibodies or TNFR1-Fc chimeras and function to prevent TNFα from binding to its receptor. Those biomacromolecular agents have been proved to be effective in the treatment of inflammatory bowel disease and rheumatoid arthritis due to their unique superiorities such as high specificity. However, several severe limitations such as poor stability, cost-ineffective commercial-scale production and exclusion from blood/brain barrier have also emerged. Instead, small-molecule chemical compounds have been appreciated as appropriate alternatives for overcoming most disadvantages associated with macromolecular inhibitors. Furthermore, they offer additional clinical benefits such as simpler preparation for oral medicine.

Now by the use of computer-aided drug design (CADD) and cell-based assays in vitro, several selective small-molecule antagonists of TNFα activity have been identified. They include broad-spectrum inhibitors targeting the key molecules of the intracellular TNFα pathway, functionally uncharacterized inhibitors of TNFα expression, inhibitors of the processing enzyme TNFα converting enzyme (TACE), and molecules that directly bind to TNFR or prevent TNFα-TNFR interactions. Although the small-molecule inhibitors are capable of blocking the biological activity of TNFα in vitro, few have been shown to abrogate or reduce TNFα-induced inflammatory responses in vivo and exhibit high IC50 and severe side effects. Also, none of the small-molecular inhibitors have been reported to successfully block TNFα’s interaction with TNFR through direct binding to TNFα. Thus, development of small molecules for TNFα therapy remains a major challenge.

In this study, to explore chemical inhibitors against TNFα activity, we applied CADD combined with in vitro and cell-based assays and identified a lead chemical compound (named as C87 thereafter) from a compound library including about 90,000 small molecular compounds, which directly binds to TNFα indicated by SPR assay, and it potently inhibits TNFα-induced cytotoxicity (IC50=8.73μM) and effectively blocks TNFα–triggered signaling activities. More importantly, by using a murine acute hepatitis model, we showed that C87 attenuates TNFα-induced inflammation, thereby markedly reducing injuries to the liver and improving animal survival. Thus, our results lead to a novel and highly specific small-molecule TNFα inhibitor, which can be potentially used to treat TNFα mediated inflammatory diseases.