Small molecule inhibitors of influenza virus nucleoprotein

Influenza is a continuing worldwide public health threat, with seasonal activity that is not adequately controlled
by the yearly vaccine program. The severity of this problem is likely to increase with emergence of new influenza
virus strains in the human population, and this situation is made even more complex by the COVID-19 pandemic.
Both COVID-19 and the 2009 H1N1 influenza pandemics are reminders of the challenge posed by emergent
viruses, and they also highlight weaknesses in global preparedness should additional threats arise. Despite
existing seasonal vaccines and two classes of chemical pharmaceuticals in current use, there is an urgent need
for new anti-influenza therapeutic agents to provide broader coverage, ensure against emergence of drug
resistance and prepare for future inevitable pandemics. This Phase I STTR application is to develop potent
inhibitors to a novel viral target, the nucleoprotein NP. We have identified and characterized two inhibitor classes,
with promising affinity and potency, lack of cytotoxicity and good PK properties. The goals of Phase I are to
demonstrate in vivo efficacy in the mouse model, select and characterize escape mutants in cell culture, and
further explore one of our series with excellent opportunities for additional medicinal chemistry. Included in this
effort is the important goal of creating broad-spectrum antivirals that are active against H1N1 and H3N2 seasonal
strains – which is achievable given the highly conserved nature of the NP target. Our approach is divided among
three, integrated Specific Aims. In Aim 1 the BALB/c mouse infection model will be used for efficacy studies of
leads JJNP9-4 and MC-2, based on successful MTD and PK studies in the mouse. Several parameters will be
measured as indicators of efficacy, including mortality, body weight, viral load in the lung and other standard
behavioral and physical parameters. In Aim 2, JJNP9-4 and MC-2 will be used for selection of virus escape
mutants, followed by sequencing of the mutants, reverse genetic construction of drug-resistant variants, and
characterization of viral fitness of the escape mutants. The results will inform future structural studies to identify
the binding sites of the inhibitors. In Aim 3 the potency of one inhibitor series will be addressed by developing
an SAR with the synthesis of individual analogs and combinatorial libraries Two top-prioritized analogs will be
used for MTD, PK and efficacy studies in the mouse model of infection.Project Narrative
Yearly influenza epidemics affect about 10% of the worldandapos;s population and estimates of annual mortality range
from 250,000 – 500,000, including approximately 30,000 deaths and 200,000 hospitalizations in the United
States. The proposed research is directed at developing new drugs that can combat influenza virus, the virus
that causes influenza.