A Translation Inhibitor That Suppresses Dengue Virus In Vitro and In Vivo

Abstract

We describe a novel translation inhibitor that has anti-dengue virus (DENV) activity in vitro and in vivo. The inhibitor was identified through a high-throughput screening using a DENV infection assay. The compound contains a benzomorphan core structure. Mode-of-action analysis indicated that the compound inhibits protein translation in a viral RNA sequence-independent manner. Analysis of the stereochemistry demonstrated that only one enantiomer of the racemic compound inhibits viral RNA translation. Medicinal chemistry was performed to eliminate a metabolically labile glucuronidation site of the compound to improve its in vivo stability. Pharmacokinetic analysis showed that upon a single subcutaneous dosing of 25 mg/kg of body weight in mice, plasma levels of the compound reached a C max (maximum plasma drug concentration) above the protein-binding-adjusted 90% effective concentration (EC 90 ) value of 0.96 M. In agreement with the in vivo pharmacokinetic results, treatment of DENV-infected mice with 25 mg/kg of compound once per day reduced peak viremia by about 40-fold. However, mice treated with 75 mg/kg of compound per day exhibited adverse effects. Collectively, our results demonstrate that the benzomorphan compounds inhibit DENV through suppression of RNA translation. The therapeutic window of the current compounds needs to be improved for further development.Dengue virus (DENV), a member of the Flavivirus genus from the Flaviviridae family, causes approximately 50 million to 100 million human infections annually (12). Besides DENV, many other flaviviruses are important human pathogens, including West Nile virus (WNV), yellow fever virus (YFV), Japanese encephalitis virus (JEV), and tick-borne encephalitis virus (TBEV). There is no clinically approved antiviral treatment for any flavivirus infection. The current treatment for flavivirus infection is only supportive. Human vaccines for flaviviruses are available only for YFV, JEV, and TBEV (12). Development of a dengue vaccine is challenging because a successful vaccine requires a balanced immune response to all four serotypes of DENV; an unbalanced vaccine could lead to enhanced virus replication mediated by serotype-cross-reactive antibodies (21). Therefore, antiviral therapy is urgently needed for treatment of flavivirus infections.The flaviviral genome is a plus-sense RNA about 11 kb in length. The genomic RNA is composed of a 5Ј untranslated region (UTR), a single open reading frame, and a 3Ј UTR. The single open reading frame encodes a long polyprotein that is cleaved by a combination of viral and host proteases into 10 viral proteins, three structural proteins (capsid [C], premembrane [prM], and envelope [E]) and seven nonstructural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5). The structural proteins form virus particles. The nonstructural proteins, together with host factors, form a replication complex to replicate viral RNA (17). Only two of the flavivirus proteins have enzymatic activities. NS3 functions as a viral serine protease, with ...

A Novel Inhibitor of Dengue Virus Replication That Targets the Capsid Protein

“…Virus-specific inhibitors have been identified that target the viral envelope (13), methyl transferase (14), protease (15), NS4B protein (16), polymerase (17,18), and virus-specific RNA translation (19). In addition, compounds that target host enzymes, such as ER glucosidases (20)(21)(22)(23), dihydroorotate dehydrogenase (19), and an intracellular cholesterol transporter (24), have been shown to have antiviral activity. Although these compounds appear to be effective at inhibiting DENV replication, there is still no approved antiviral therapeutic for the treatment of DENV infection in humans.…”

Novel Indole-2-Carboxamide Compounds Are Potent Broad-Spectrum Antivirals Active against Western Equine Encephalitis Virus In Vivo

“…Detailed mechanism-of-action studies, breadth of antiviral activity outside the alphavirus genus, and in vivo activity were not explored with this natural product-derived compound, so a more detailed comparison with CCG205432 and related compounds is not yet possible. However, Wang et al recently described a novel benzomorphan compound (NITD-451) with in vitro and in vivo antiviral activity (45) that shares intriguing contrasts with our indole-2-carboxamide compounds. The benzomorphan series of compounds were first developed as antivirals against DENV, and NITD-451 shows potent activity against several flaviviruses but not WEEV, whereas CCG205432 is active against WEEV and related alphaviruses but not DENV.…”

“…The benzomorphan series of compounds were first developed as antivirals against DENV, and NITD-451 shows potent activity against several flaviviruses but not WEEV, whereas CCG205432 is active against WEEV and related alphaviruses but not DENV. Furthermore, although both series of compounds have enantiospecific activity and modulate cellular gene expression, NITD-451 suppresses translation in cells and in in vitro assays (45), whereas CCG205432 was active only in cells and did not inhibit expression from mRNA templates in vitro, suggesting different mechanisms of action. However, Wang et al were also unable to generate NITD-451-resistant DENV, similar to our failed attempts to generate CCG205432-resistant WEEV and FMV, consistent with both compounds having host factors as targets.…”

“…Thus, the number of cellular processes that serve as potential targets for antiviral agents likely exceeds the number of viral targets with pathogens such as alphaviruses, whose genome encodes a limited number of proteins with known enzymatic activities (4). Several recently published cell-based antiviral screens have identified a number of compounds that target host factors, from pyrimidine biosynthesis (17,51,57) to protein translation (45,58). Targeting a cellular process also has the potential advantage of presenting a higher barrier for the development of resistance, a concept with substantial experimental and clinical support.…”

Ethyl 4-(4-methylphenyl)-4-pentenoate from Vetiveria zizanioides Inhibits Dengue NS2B–NS3 Protease and Prevents Viral Assembly: A Computational Molecular Dynamics and Docking Study

“…Proteolysis of the dengue virus genome yields three structural and seven nonstructural proteins [6]. During viral maturation, the polyprotein is cleaved by the NS3 serine protease at the N-terminal domain, interceded by the cofactor NS2B [7]. Processing of this polyprotein is a fundamental process that must occur before viral RNA replication process.…”