The incidence of dengue fever epidemics has increased dramatically over the last few decades. However, no vaccine or antiviral therapies are available. Therefore, the need for safe and effective antiviral drugs has become imperative. The entry of dengue virus into a host cell is mediated by its major envelope (E) protein. The crystal structure of the E protein reveals a hydrophobic pocket that is presumably important for low-pHmediated membrane fusion. High-throughput docking with this hydrophobic pocket was performed, and hits were evaluated in cell-based assays. Compound 6 was identified as one of the inhibitors and had an average 50% effective concentration of 119 nM against dengue virus serotype 2 in a human cell line. Mechanism-ofaction studies demonstrated that compound 6 acts at an early stage during dengue virus infection. It arrests dengue virus in vesicles that colocalize with endocytosed dextran and inhibits NS3 expression. The inhibitors described in this report can serve as molecular probes for the study of the entry of flavivirus into host cells.

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Growing evidence suggests that the presence of a subpopulation of hypoxic non-replicating, phenotypically drug-tolerant mycobacteria is responsible for the prolonged duration of tuberculosis treatment. The discovery of new antitubercular agents active against this subpopulation may help in developing new strategies to shorten the time of tuberculosis therapy. Recently, the maintenance of a low level of bacterial respiration was shown to be a point of metabolic vulnerability in Mycobacterium tuberculosis. Here, we describe the development of a hypoxic model to identify compounds targeting mycobacterial respiratory functions and ATP homeostasis in whole mycobacteria. The model was adapted to 1,536-well plate format and successfully used to screen over 600,000 compounds. Approximately 800 compounds were confirmed to reduce intracellular ATP levels in a dose-dependent manner in Mycobacterium bovis BCG. One hundred and forty non-cytotoxic compounds with activity against hypoxic non-replicating M. tuberculosis were further validated. The resulting collection of compounds that disrupt ATP homeostasis in M. tuberculosis represents a valuable resource to decipher the biology of persistent mycobacteria.

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