Synthesis of Novel <i>N</i>⁴-Hydrocytidine Analogs as Potential Anti-SARS-CoV-2 Agents

Coronavirus disease 2019 (COVID-19) is an emerging global pandemic with severe morbidity and mortality caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Molnupiravir, an ester prodrug form of N4-hydroxycytidine (NHC), was recently emergency-use approved for the treatment of early SARS-CoV-2 infections. Herein, we report the synthesis and evaluation of a series of novel NHC analogs

Nucleoside Analogs with Selective Antiviral Activity against Dengue Fever and Japanese Encephalitis Viruses

Dengue virus (DENV) and Japanese encephalitis virus (JEV) are important arthropod-borne viruses from the Flaviviridae family. DENV is a global public health problem with significant social and economic impacts, especially in tropical and subtropical areas. JEV is a neurotropic arbovirus endemic to east and southeast Asia. There are no U.S. FDA-approved antiviral drugs available to treat or to prevent DENV and JEV infections, leaving nearly one-third of the world’s population at risk for infection. Therefore, it is crucial to discover potent antiviral agents against these viruses. Nucleoside analogs, as a class, are widely used for the treatment of viral infections. In this study, we discovered nucleoside analogs that possess potent and selective anti-JEV and anti-DENV activities across all serotypes in cell-based assay systems. Both viruses were susceptible to sugar-substituted 2=-C-methyl analogs with either cytosine or 7-deaza-7-fluoro-adenine nucleobases. Mouse studies confirmed the anti-DENV activity of these nucleoside analogs. Molecular models were assembled for DENV serotype 2 (DENV-2) and JEV RNA-dependent RNA polymerase replication complexes bound to nucleotide inhibitors. These models show similarities between JEV and DENV-2, which recognize the same nucleotide inhibitors. Collectively, our findings provide promising compounds and a structural rationale for the development of direct-acting antiviral agents with dual activity against JEV and DENV infections

Substrates and Inhibitors of SAMHD1

<div><p>SAMHD1 hydrolyzes 2'-deoxynucleoside-5'-triphosphates (dNTPs) into 2'-deoxynucleosides and inorganic triphosphate products. In this paper, we evaluated the impact of 2' sugar moiety substitution for different nucleotides on being substrates for SAMHD1 and mechanisms of actions for the results. We found that dNTPs ((2'<i>R</i>)-2'-H) are only permissive in the catalytic site of SAMHD1 due to L150 exclusion of (2'<i>R</i>)-2'-F and (2'<i>R</i>)-2'-OH nucleotides. However, arabinose ((2'<i>S</i>)-2'-OH) nucleoside-5'-triphosphates analogs are permissive to bind in the catalytic site and be hydrolyzed by SAMHD1. Moreover, when the (2'<i>S</i>)-2' sugar moiety is increased to a (2'<i>S</i>)-2'-methyl as with the SMDU-TP analog, we detect inhibition of SAMHD1’s dNTPase activity. Our computational modeling suggests that (2'<i>S</i>)-2'-methyl sugar moiety clashing with the Y374 of SAMHD1. We speculate that SMDU-TP mechanism of action requires that the analog first docks in the catalytic pocket of SAMHD1 but prevents the A351-V378 helix conformational change from being completed, which is needed before hydrolysis can occur. Collectively we have identified stereoselective 2' substitutions that reveal nucleotide substrate specificity for SAMHD1, and a novel inhibitory mechanism for the dNTPase activity of SAMHD1. Importantly, our data is beneficial for understanding if FDA-approved antiviral and anticancer nucleosides are hydrolyzed by SAMHD1 <i>in vivo</i>.</p></div

Cartoon for SAMHD1 homotetramerization.

<p>SAMHD1 monomers bind GTP (or dGTP) at allosteric 1 (A1) site leading to the formation of homodimers. Next, dNTPs bind to allosteric 2 (A2) sites allowing for the formation of homotetramers. The catalytic (Cat) site then can accompany dNTPs for hydrolysis, leading to the generation of deoxynucleosides (dNs) and inorganic triphosphates (iPPP).</p