We have employed a structure-based drug design approach to systematically evaluate the structure-activity relationship (SAR) of zafirlukast, a known inhibitor that targets the pre-ligand assembly domain (PLAD) of tumor necrosis factor receptor 1 (TNFR1) and inhibits its function. Receptor-specific inhibition of TNFR1 is a highly sought-after strategy for the treatment of autoimmune diseases such as rheumatoid arthritis. Through performing an innovative FRET-based high-throughput screening, we have recently identified zafirlukast as a small-molecule inhibitor of TNFR1 activation which acts through a novel mechanism of disrupting TNFR1 PLAD-PLAD interaction without ablating native ligand binding. In the current study, we focus on establishing a SAR of zafirlukast by adopting lipophilic ligand efficiency (LLE) and topological polar surface area (TPSA) as metrics to optimize the molecule for improved potency, as well as to investigate the portion of the molecule important for this inhibition mechanism. We have synthesized over 30 analogues of zafirlukast and have managed to improve its activity significantly by 10-fold to a half maximal inhibitory concentration (IC<sub>50</sub>) of 6 μM, while improving on the drug disposition properties. We are testing two more series of analogues, potentially achieving an IC<sub>50</sub> in the nanomolar range. The chemical structures and molecular weights of the analogues were confirmed by NMR and mass spectrometry. In addition, through both experimental observations and computational simulations, we have found that both the sulfonamide and carbamate groups of the molecule are responsible in receptor binding, and the aromatic ring on the sulfonamide group plays a key role in the disruption of PLAD-PLAD interactions. This is a significant discovery not only because our optimized molecule is a novel and potent inhibitor of TNFR1, but more importantly we are the first to provide information on the chemical structures crucial in disrupting TNF receptor-receptor interaction.

Structure-activity relationship analysis of TNF receptor inhibitors for elucidation of inhibition mechanisms and therapeutic developments

256th National Meeting (2018)

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