Despite a high current standard of care in antiretroviral therapy for HIV, multidrug-resistant strains continue to emerge, underscoring the need for additional novel mechanism inhibitors that will offer expanded therapeutic options in the clinic. We report a new class of small molecule antiretroviral compounds that directly target HIV-1 capsid (CA) via a novel mechanism of action. The compounds exhibit potent antiviral activity against HIV-1 laboratory strains, clinical isolates, and HIV-2, and inhibit both early and late events in the viral replication cycle. We present mechanistic studies indicating that these early and late activities result from the compound affecting viral uncoating and assembly, respectively. We show that amino acid substitutions in the N-terminal domain of HIV-1 CA are sufficient to confer resistance to this class of compounds, identifying CA as the target in infected cells. A high-resolution co-crystal structure of the compound bound to HIV-1 CA reveals a novel binding pocket in the N-terminal domain of the protein. Our data demonstrate that broad-spectrum antiviral activity can be achieved by targeting this new binding site and reveal HIV CA as a tractable drug target for HIV therapy.

Human immunodeficiency virus type 1 (HIV-1

West Nile virus (WNV) is a neurotropic, mosquito-borne flavivirus that can cause lethal meningoencephalitis. Type I interferon (IFN) plays a critical role in controlling WNV replication, spread, and tropism. In this study, we begin to examine the effector mechanisms by which type I IFN inhibits WNV infection. Mice lacking both the interferon-induced, double-stranded-RNA-activated protein kinase (PKR) and the endoribonuclease of the 2,5-oligoadenylate synthetase-RNase L system (PKR ؊/؊ ؋ RL ؊/؊ ) were highly susceptible to subcutaneous WNV infection, with a 90% mortality rate compared to the 30% mortality rate observed in congenic wild-type mice. PKR ؊/؊ ؋ RL ؊/؊ mice had increased viral loads in their draining lymph nodes, sera, and spleens, which led to early viral entry into the central nervous system (CNS) and higher viral burden in neuronal tissues. Although mice lacking RNase L showed a higher CNS viral burden and an increased mortality, they were less susceptible than the PKR ؊/؊ ؋ RL ؊/؊ mice; thus, we also infer an antiviral role for PKR in the control of WNV infection. Notably, a deficiency in both PKR and RNase L resulted in a decreased ability of type I IFN to inhibit WNV in primary macrophages and cortical neurons. In contrast, the peripheral neurons of the superior cervical ganglia of PKR ؊/؊ ؋ RL ؊/؊ mice showed no deficiency in the IFN-mediated inhibition of WNV. Our data suggest that PKR and RNase L contribute to IFN-mediated protection in a cell-restricted manner and control WNV infection in peripheral tissues and some neuronal subtypes.

Previous studies have suggested that ␣-glucosidase inhibitors such as castanospermine and deoxynojirimycin inhibit dengue virus type 1 infection by disrupting the folding of the structural proteins prM and E, a step crucial to viral secretion. We extend these studies by evaluating the inhibitory activity of castanospermine against a panel of clinically important flaviviruses including all four serotypes of dengue virus, yellow fever virus, and West Nile virus. Using in vitro assays we demonstrated that infections by all serotypes of dengue virus were inhibited by castanospermine. In contrast, yellow fever virus and West Nile virus were partially and almost completely resistant to the effects of the drug, respectively. Castanospermine inhibited dengue virus infection at the level of secretion and infectivity of viral particles. Importantly, castanospermine prevented mortality in a mouse model of dengue virus infection, with doses of 10, 50, and 250 mg/kg of body weight per day being highly effective at promoting survival (P < 0.0001). Correspondingly, castanospermine had no adverse or protective effect on West Nile virus mortality in an analogous mouse model. Overall, our data suggest that castanospermine has a strong antiviral effect on dengue virus infection and warrants further development as a possible treatment in humans.Dengue fever, the most prevalent arthropod-borne viral illness in humans, is caused by dengue virus (DEN). DEN is a single-stranded, positive-polarity, enveloped RNA virus that is translated in the cytoplasm as a single polyprotein and cleaved into three structural and seven nonstructural proteins. Four related serotypes of DEN exist in nature and are transmitted to humans primarily by two mosquitoes, Aedes aegypti and Aedes albopictus. DEN is a member of the Flaviviridae family and is genetically related to the viruses that cause yellow fever, hepatitis C, and the Japanese, St. Louis, and West Nile encephalitides. DEN infection results in a spectrum of disease ranging from a debilitating, self-limited illness (dengue fever) to a life-threatening syndrome (dengue hemorrhagic fever [DHF]). DEN causes disease globally with an estimated 25 to 100 million new infections per year (34). At present, no vaccine has been approved for human use and treatment is supportive.Flavivirus assembly takes place at the endoplasmic reticulum (ER) (7). The structural glycoproteins prM and E localize to the luminal side of the ER and form an immature particle with prM and E in a heterodimeric complex (7, 58). Furin-mediated proteolysis of prM in the trans-Golgi network (48) triggers rearrangement, homodimerization of E, and formation of the mature viral particle before release from the infected cell (1, 16). In flavivirus-infected mammalian cells, a 14-residue oligosaccharide, (Glc) 3 (Man) 9 (GlcNAc) 2 , is added in the ER to specific asparagine residues on the prM and E proteins. This high-mannose carbohydrate is sequentially modified in the ER by resident ␣-glucosidases to generate N-linked glycans that lack t...