Dengue virus NS1 protein as a mediator of pathology (2016–2019)
Severe complications of dengue virus infection include potentially fatal hemorrhage and shock, which have been linked with a cytokine storm resulting from excessive monocytes/macrophage activation. Understanding the triggers that lead to these events is critical for developing appropriate therapeutic strategies. Dengue virus non-structural protein NS1 has a secreted hexameric form that can circulate at high levels. We have found that the secreted dengue NS1 directly activates mouse macrophages and human blood cells via toll-like receptor 4 (TLR4), inducing the release of a range of cytokines and chemokines. LPS contamination was excluded as a source of this activity. Thus NS1 is a dengue virus-encoded pathogen-associated molecular pattern (PAMP). In an in vitro model of vascular leak we further showed that treatment with NS1 resulted in the disruption of endothelial cell monolayer integrity. We obtained complete inhibition of NS1-mediated cytokine release and monolayer disruption using the TLR4 antagonist LPS-RS, suggesting a route for therapeutic intervention in severe disease. Indeed, our preliminary work suggests that the TLR4 antagonist prevents vascular leak in a mouse model of dengue infection. Similar to the role of LPS in septic shock, NS1 may contribute to vascular leak and shock in dengue patients. In this project we will investigate (i) the molecular characteristics of NS1 required for TLR4 activation; (ii) the molecular interaction between NS1 and the TLR4 receptor complex, and its inhibition by established TLR4 antagonists; (iii) the effect of therapeutic blockade of TLR4 signalling in a mouse model of infection using eritoran, which has been used already in clinical trials; (iv) the possibility that NS1 from other flaviviruses such as West Nile virus and Japanese encephalitis virus (JEV) plays a role in pathology. The successful outcome will be to establish existing TLR4 antagonist drugs as candidates for dengue virus therapy.