Polyamine depletion inhibits norovirus infection by blocking virus-induced apoptosis

Abstract Noroviruses are non-enveloped, positive-sense RNA viruses in the family Caliciviridae. Human noroviruses (HNoVs) are the leading cause of viral gastrointestinal disease. Despite their clinical relevance, therapeutic strategies for norovirus infections remain limited, partly due to challenges in cultivating HNoV and limited understanding of virus-host interactions. Polyamines (PAs) are small polycationic metabolites derived from amino acid metabolisms and are essential for diverse cellular functions and important host factors for many RNA viruses. However, their role in norovirus infection has remained elusive. Here, we demonstrate that PAs are critical for productive infection for both HNoV and murine norovirus (MNV), underscoring a conserved and functionally significant dependency across species. Functional analysis in MNV-infected murine macrophages showed that PA depletion did not significantly impair virus attachment and viral genome replication. However, it resulted in a marked reduction in infectious virus titers following the first replication cycle, coinciding with a loss of virus-induced apoptosis and impaired virion release. Exogenous supplementation with two metabolites of the PAs biosynthetic pathway (spermine or spermidine), or induction of apoptosis using a PI3-kinase inhibitor, restored both viral titers and apoptosis in PA-depleted macrophages. In human intestinal enteroids (HIEs), depletion of PAs completely ablated HNoV infection, which was partially rescued adding exogenous PAs. Collectively, our findings establish PAs as a host dependency factor that facilitates norovirus infection, promoting virus-induced apoptosis and egress in the murine system. Furthermore, these results point to PAs metabolism as a potential therapeutic target for antiviral intervention against HNoV infections. Importance HNoV are a major cause of acute viral gastroenteritis worldwide, yet the development of effective antivirals and vaccines has been hampered by the lack of small-animal models and robust in vitro systems to propagate the virus. MNV shares key features with HNoV and serves as a valuable surrogate for studying norovirus biology. In this study, we identify host-derived PAs as essential and conserved metabolites required for productive infection for both HNoV and MNV. PAs exert multifactorial effects on norovirus infection, particularly by facilitating virus-induced apoptosis and promoting virion release in MNV-infected cells. Notably, PA depletion using difluoromethylornithine (DFMO), a licensed and well-tolerated inhibitor of PA synthesis, effectively suppressed norovirus infection. Our findings highlight the potential of repurposing DFMO as a host-targeting antiviral, especially for the treatment of chronic norovirus infections in immunocompromised individuals. This work provides new insight into norovirus-host interactions and proposes a readily translatable therapeutic avenue. Competing Interest Statement The authors have declared no competing interest.