THE DIFFICULT PATH TOWARD THE KNOWLEDGE: AN ONGOING STORY ON NEW DENGUE VIRUS INHIBITORS

Eloise Mastrangelo, Delia Tarantino, Gilles Querat, Giuseppe Manfroni and Mario Milani

Biophysics Institute, National Research Council (CNR), Italy

Abstract

From a property library (UNIPG), we recently selected the inhibitor cp1 (with pyridobenzothiazole scaffold) active against Dengue virus polymerase (IC50~1.5 uM). The crystal structure of the protein-inhibitor complex suggested a competitive inhibition mechanism in contrast with enzyme kinetics measurements (Tarantino et al., 2016). Interestingly, the compound was active against different flavivirus in cell culture (EC50~10 uM). In an effort to enhance potency and clarify the mechanism of action we produced a number of variants. Unexpectedly, different experiments with a more potent homologous (cp27, EC50~7 uM) indicated that the polymerase was not the real target of this class of inhibitors in cell. In particular, we demonstrated that cp27 does not interfere with the viral RNA replication (not reducing the amount of RNA nor enhancing the mutation rate) but produced a viral progeny incapable of spread infection efficiently. Despite the failure to select a definite viral target by inhibitor-induced resistance, all our results point to an effect of cp27 in virion maturation and/or assembly, i.e. in the late steps of viral replication. This work shows how, starting from a classical target driven approach, it is sometime necessary to travel an unpredictable path to disclose the true nature of a novel active compound.

Keywords: Dengue virus, RNA dependent RNA polymerase, in silico docking, crystal structure, virus maturation, viral proteins