Project 4 - Inhibitors of Flavivirus Replication

Emerging and endemic members of the genus Flavivirus pose significant worldwide public health threats. The global ranges of dengue viruses, Japanese encephalitis virus, West Nile virus (WNV), tick-borne encephalitis virus and ZIKV have recently expanded. Dengue virus is estimated to infect ~400 million, cause illness in ~100 million, and kill ~21,000 people a year. Yellow fever virus remains a public health concern in 34 African countries and 13 South American countries. Newly emerging flaviviruses include Powassan virus (POWV) (northeast US and Canada) and Usutu virus (Europe). Vaccines are available for humans for only some flaviviruses, but they are not always used. A few individuals develop multiorgan disease after vaccination with the live 17D yellow fever vaccine which is typically fatal. There are currently no effective flavivirus-specific small molecule therapeutics for treatment of humans with flavivirus infections. The goal of the research proposed in Project 4 of the AC/DC is to develop clinical candidate compounds that can be delivered orally, have broad anti-flavivirus activity and directly act on a viral component. Under Aim 1, we will develop four ribonucleoside analogs synthesized by Core B, for which preliminary data indicating anti-flavivirus activity has already been obtained, by a reiterative analog synthesis-biological testing strategy with Cores B and C. Compounds will be tested for antiviral activity against a panel of flaviviruses and cytotoxicity in primary mouse and human macrophages and neural cells. Mode of action and development of drug resistance will also be analyzed. Under Aim 2, reporter POWV and WNV viruses will be generated and used in high throughput screening of ~300,000 unique compounds under BSL3 containment by Core F. Hits validated by counter-screening will be further developed under Aim 1. Compounds meeting the advancement criteria will be tested for in vivo efficacy in flavivirus mouse and hamster infection models and in human cortical organoids. The effect of the gut microbiome on the activity of lead flavivirus antiviral compounds will be assessed in collaboration with Project 7. The impact of antiviral treatment on acute and chronic virus-induced neurological signatures in POWV and WNV infected animals and organoids will be evaluated in collaboration with Project 2. Core D will provide whole genome sequencing of viral mutants generated during viral resistance profiling of lead compounds.