Mechanistic Study and in vivo Efficacy Test of BPR3P---A Novel Enterovirus 71 Agent that May Inhibit Viral Replication Complex Formation (I)

Recurring outbreaks of enterovirus 71 (EV71) cause substantial financial and social burdens annually in Taiwan. The current lack of vaccination and approved therapeutic agents has prompted us to develop new antiviral compounds. This proposal is aimed at identifying the antiviral mechanisms of a novel compound, BPR3P0128 (patent pending), and testing its in vivo actions in an animal model. In collaboration with the National Health Research Institute, we have used a systematic compound library to screen for potential new inhibitors against influenza viruses. Intriguingly, some antiviral candidates also possess broad potent activity against other RNA viruses. These candidates include the BPR3P series of compounds having anti-EV71 activity of submicromolar EC50 in cell culture; the pharmacokinetic data show at least 80% oral bioavailability in the rat. This grant proposal to discover new antiviral agents and to study and optimize their in vitro and in vivo actions is in partnership with Drs. Tsuan Hsu (徐祖安) and Hsing-Pang Hsieh (謝興邦) of the National Health Research Institute, Taiwan. We propose to study the underlying anti-EV71 mechanism of BPR3P and its efficacy in vivo. The in vivo efficacy in mice of BPR3P and rosmarinic acid (RA) will be tested at the Research Center for Emerging Viral Infections by Professor Shin-Ru Shih (component project #1). If the agent show effectiveness in mice, it will become more attractive for development as a drug candidate. Our specific aim is to conduct mechanism-of-action studies, laboratory animal testing, and target identification studies to understand further the antiviral mechanisms of BPR3P. The mechanistic studies will include assessing the inhibition spectrum and using in vitro polymerase assays, time-of-addition assays, resistant virus selection, and combinational assays with inhibitors identified by the other two component projects. We will also develop the pharmacokinetic profile by measuring the half-life of the drug; evaluating its bioavailability, area under the curve, and clearance; and making interspecies comparisons. The BPR3P targets will be identified using in silico and affinity chromatography. Alternatively, we propose to study the inhibitory mechanisms and in vivo activity of a pure natural compound, RA, a natural phenol antioxidant carboxylic acid found in many herbs such as lemon balm, rosemary, oregano, sage, thyme, and peppermint. RA has potent activity against EV71 and inhibits the viral titer by at least 100-fold at nontoxic concentrations in RD cells. Viral proteins 2C and 3A are involved in the formation of the viral replication complex. Using a yeast two-hybrid assay, we have identified several 2C- and 3A-interacting cellular proteins that are involved in viral replication. We will test whether the inhibitory effect of BPR3P results from disruption to the interaction between these cellular proteins and viral proteins. These cellular proteins may serve as drug targets.

Project ID:PC10106-0041
External Project ID:NSC101-2325-B182-016