AT-9010 (2'-methyl-2'-fluoro guanosine triphosphate) is a GTP analog whose prodrug, AT-752 is under consideration in human medicine as a potential antiviral drug against certain flaviviruses. It was previously believed to inhibit viral replication by acting primarily as a chain terminator. However, it was discovered recently that it also binds the GTP binding site of the methyltransferase (MTase) domain of the orthoflavivirus polymerase, thus interfering with RNA capping. Here, we investigated the binding of AT-9010 to Ntaya and Zika virus MTases. Structural analysis using X-ray crystallography revealed similar interactions between the base and sugar moieties of AT-9010 and key residues in both MTases, although differences in hydrogen bonding were observed. Our analysis also suggested that the triphosphate part of AT-9010 is flexible. Despite minor variations, the overall binding mode of AT-9010 was found to be the same for all of the flaviviral MTases examined, suggesting a structural basis for the efficacy of AT-9010 against multiple orthoflavivirus MTases.

Institute of Organic Chemistry and Biochemistry Academy of Sciences of the Czech Republic v v i Flemingovo nám 2 Prague 16610 Czech Republic

Zobrazit více v PubMed

Postler TS, Beer M, Blitvich BJ, Bukh J, de Lamballerie X, Drexler JF, Imrie A, Kapoor A, Karganova GG, Lemey P, Lohmann V, Simmonds P, Smith DB, Stapleton JT, Kuhn JH (2023) Renaming genus Ext binomial species names within family Archives Virol, 168(9). PubMed

Yap TL, Xu T, Chen YL, Malet H, Egloff MP, Canard B, Vasudevan SG, Lescar J (2007) Crystal structure of the dengue virus RNA-dependent RNA polymerase catalytic domain at 1.85-angstrom resolution. J Virol 81(9):4753–4765 PubMed PMC

Dubankova A, Boura E (2019) Structure of the yellow fever NS5 protein reveals conserved drug targets shared among flaviviruses. Antiviral Res 169:104536 PubMed

Upadhyay AK, Cyr M, Longenecker K, Tripathi R, Sun C, Kempf DJ (2017) Crystal structure of full-length Zika virus NS5 protein reveals a conformation similar to Japanese encephalitis virus NS5. Acta Crystallogr F Struct Biol Commun 73(Pt 3):116–122 PubMed PMC

Sebera J, Dubankova A, Sychrovsky V, Ruzek D, Boura E, Nencka R (2018) The structural model of Zika virus RNA-dependent RNA polymerase in complex with RNA for rational design of novel nucleotide inhibitors. Sci Rep 8(1):11132 PubMed PMC

Noble CG, Lim SP, Chen YL, Liew CW, Yap L, Lescar J, Shi PY (2013) Conformational flexibility of the Dengue virus RNA-dependent RNA polymerase revealed by a complex with an inhibitor. J Virol 87(9):5291–5295 PubMed PMC

Arora R, Liew CW, Soh TS, Otoo DA, Seh CC, Yue K, Nilar S, Wang G, Yokokawa F, Noble CG, Chen YL, Shi PY, Lescar J, Smith TM, Benson TE, Lim SP (2020) Two RNA Tunnel Inhibitors Bind in Highly Conserved Sites in Dengue Virus NS5 Polymerase: Structural and Functional Studies. J Virol, 94(24). PubMed PMC

Konkolova E, Krejcova K, Eyer L, Hodek J, Zgarbova M, Fortova A, Jirasek M, Teply F, Reyes-Gutierrez PE, Ruzek D, Weber J, Boura E (2022) A Helquat-like Compound as a Potent Inhibitor of Flaviviral and Coronaviral Polymerases. Molecules, 27(6) PubMed PMC

Good SS, Westover J, Jung KH, Zhou XJ, Moussa A, Colla PL, Collu G, Canard B, Sommadossi JP (2021) AT-527, a Double Prodrug of a Guanosine Nucleotide Analog, Is a Potent Inhibitor of SARS-CoV-2 In Vitro and a Promising Oral Antiviral for Treatment of COVID-19. Antimicrob Agents Chemother, 65(4) PubMed PMC

Konkolova E, Dejmek M, Hrebabecky H, Sala M, Boserle J, Nencka R, Boura E (2020) Remdesivir triphosphate can efficiently inhibit the RNA-dependent RNA polymerase from various flaviviruses. Antiviral Res 182:104899 PubMed PMC

Milisavljevic N, Konkolova E, Kozak J, Hodek J, Veselovska L, Sykorova V, Cizek K, Pohl R, Eyer L, Svoboda P, Ruzek D, Weber J, Nencka R, Boura E, Hocek M (2021) Antiviral Activity of 7-Substituted 7-Deazapurine Ribonucleosides, Monophosphate Prodrugs, and Triphoshates against Emerging RNA Viruses. ACS Infect Dis 7(2):471–478 PubMed

Dong H, Fink K, Zust R, Lim SP, Qin CF, Shi PY (2014) Flavivirus RNA methylation J Gen Virol 95(Pt 4):763–778 PubMed

Song W, Zhang H, Zhang Y, Chen Y, Lin Y, Han Y, Jiang J (2021) Identification and Characterization of Zika Virus NS5 Methyltransferase Inhibitors. Front Cell Infect Microbiol 11:665379 PubMed PMC

Delgado-Maldonado T, Moreno-Herrera A, Pujadas G, Vazquez-Jimenez LK, Gonzalez-Gonzalez A, Rivera G (2023) Recent advances in the development of methyltransferase (MTase) inhibitors against (re)emerging arboviruses diseases dengue and Zika. Eur J Med Chem 252:115290 PubMed

Dong H, Zhang B, Shi PY (2008) Flavivirus methyltransferase: a novel antiviral target. Antiviral Res 80(1):1–10 PubMed PMC

Milani M, Mastrangelo E, Bollati M, Selisko B, Decroly E, Bouvet M, Canard B, Bolognesi M (2009) Flaviviral methyltransferase/RNA interaction: Structural basis for enzyme inhibition. Antiviral Res 83(1):28–34 PubMed PMC

Yap LJ, Luo D, Chung KY, Lim SP, Bodenreider C, Noble C, Shi PY, Lescar J (2010) Crystal structure of the dengue virus methyltransferase bound to a 5'-capped octameric RNA. PLoS ONE, 5(9) PubMed PMC

Kottur J, Rechkoblit O, Quintana-Feliciano R, Sciaky D, Aggarwal AK (2022) High-resolution structures of the SARS-CoV-2 N7-methyltransferase inform therapeutic development. Nat Struct Mol Biol 29(9):850–853 PubMed PMC

Ahmed-Belkacem R, Hausdorff M, Delpal A, Sutto-Ortiz P, Colmant AMG, Touret F, Ogando NS, Snijder EJ, Canard B, Coutard B, Vasseur JJ, Decroly E, Debart F (2022) Potent Inhibition of SARS-CoV-2 nsp14 N7-Methyltransferase by Sulfonamide-Based Bisubstrate Analogues. J Med Chem 65(8):6231–6249 PubMed

Otava T, Sala M, Li F, Fanfrlik J, Devkota K, Perveen S, Chau I, Pakarian P, Hobza P, Vedadi M, Boura E, Nencka R (2021) The Structure-Based Design of SARS-CoV-2 nsp14 Methyltransferase Ligands Yields Nanomolar Inhibitors. ACS Infect Dis 7(8):2214–2220 PubMed

Zilecka E, Klima M, Stefek M, Dejmek M, Nencka R, Boura E (2024) Structure of SARS-CoV-2 MTase nsp14 with the inhibitor STM957 reveals inhibition mechanism that is shared with a poxviral MTase VP39. J Struct Biology: X,: p. 100109 PubMed PMC

Kocek H, Chalupská D, Dejmek M, Dvořáková A, Zgarbová M, Šála M, Chalupský K, Krafčíková P, Otava T, Drexler M (2024) Discovery of highly potent SARS-CoV-2 nsp14 methyltransferase inhibitors based on adenosine 5′-carboxamides. RSC Medicinal Chemistry PubMed PMC

Klima M, Khalili Yazdi A, Li F, Chau I, Hajian T, Bolotokova A, Kaniskan HU, Han Y, Wang K, Li D, Luo M, Jin J, Boura E, Vedadi M (2022) Crystal structure of SARS-CoV-2 nsp10-nsp16 in complex with small molecule inhibitors, SS148 and WZ16. Protein Sci 31(9):e4395 PubMed PMC

Silhan J, Klima M, Chalupska D, Kozic J, Boura E (2022) The structure of monkeypox virus 2′-O-ribose methyltransferase VP39 in complex with sinefungin provides the foundation for inhibitor design. bioRxiv,: p. 2022.09. 27.509668

Skvara P, Chalupska D, Klima M, Kozic J, Silhan J, Boura E (2023) Structural basis for RNA-cap recognition and methylation by the mpox methyltransferase VP39. Antiviral Res 216:105663 PubMed

Zgarbova M, Otava T, Silhan J, Nencka R, Weber J, Boura E (2023) Inhibitors of mpox VP39 2'-O methyltransferase efficiently inhibit the monkeypox virus. Antiviral Res 218:105714 PubMed

Ferrero DS, Albentosa-Gonzalez L, Mas A, Verdaguer N (2022) Structure and function of the NS5 methyltransferase domain from Usutu virus. Antiviral Res 208:105460 PubMed

Li R, Niu Z, Liu Y, Bai X, Wang D, Chen C (2022) Crystal structure and cap binding analysis of the methyltransferase of langat virus. Antiviral Res 208:105459 PubMed

Krejcova K, Krafcikova P, Klima M, Chalupska D, Chalupsky K, Zilecka E, Boura E (2024) Structural and functional insights in flavivirus NS5 proteins gained by the structure of Ntaya virus polymerase and methyltransferase. Structure PubMed

Nencka R, Silhan J, Klima M, Otava T, Kocek H, Krafcikova P, Boura E (2022) Coronaviral RNA-methyltransferases: function, structure and inhibition. Nucleic Acids Res 50(2):635–650 PubMed PMC

Krafcikova P, Silhan J, Nencka R, Boura E (2020) Structural analysis of the SARS-CoV-2 methyltransferase complex involved in RNA cap creation bound to sinefungin. Nat Commun 11(1):3717 PubMed PMC

Posthuma CC, Te Velthuis AJW, Snijder EJ (2017) Nidovirus RNA polymerases: Complex enzymes handling exceptional RNA genomes. Virus Res 234:58–73 PubMed PMC

Good SS, Shannon A, Lin K, Moussa A, Julander JG, La Colla P, Collu G, Canard B, Sommadossi JP (2021) Evaluation of AT-752, a Double Prodrug of a Guanosine Nucleotide Analog with In Vitro and In Vivo Activity against Dengue and Other Flaviviruses. Antimicrob Agents Chemother, 65(11): p. e0098821 PubMed PMC

Feracci M, Eydoux C, Fattorini V, Lo Bello L, Gauffre P, Selisko B, Sutto-Ortiz P, Shannon A, Xia HJ, Shi PY, Noel M, Debart F, Vasseur JJ, Good S, Lin K, Moussa A, Sommadossi JP, Chazot A, Alvarez K, Guilemot JC, Decroly E, Ferron F, Canard B (2023) AT-752 targets multiple sites and activities on the Dengue virus replication enzyme NS5. Antiviral Res, 212 PubMed

Lin K, Good SS, Julander JG, Weight AE, Moussa A, Sommadossi JP (2022) AT-752, a double prodrug of a guanosine nucleotide analog, inhibits yellow fever virus in a hamster model. PLoS Negl Trop Dis 16(1):e0009937 PubMed PMC

Zhou XJ, Lickliter J, Montrond M, Ishak L, Pietropaolo K, James D, Belanger B, Horga A, Hammond J (2024) -human trial evaluating safety and pharmacokinetics of AT-752, a novel nucleotide prodrug with pan-serotype activity against dengue virus. Antimicrob Agents Chemother 68(5):e0161523 PubMed PMC

Petersen E, Wilson ME, Touch S, McCloskey B, Mwaba P, Bates M, Dar O, Mattes F, Kidd M, Ippolito G, Azhar EI, Zumla A (2016) Rapid Spread of Zika Virus in The Americas - Implications for Public Health Preparedness for Mass Gatherings at the 2016 Brazil Olympic Games. Int J Infect Dis 44:11–15 PubMed

Weaver SC, Costa F, Garcia-Blanco MA, Ko AI, Ribeiro GS, Saade G, Shi PY, Vasilakis N (2016) Zika virus: History, emergence, biology, and prospects for control. Antiviral Res 130:69–80 PubMed PMC

Smithburn KC, Haddow AJ (1951) Ntaya virus; a hitherto unknown agent isolated from mosquitoes collected in Uganda. Proc Soc Exp Biol Med 77(1):130–133 PubMed

Braack L, Gouveia de Almeida AP, Cornel AJ, Swanepoel R, de Jager C (2018) Mosquito-borne arboviruses of African origin: review of key viruses and vectors. Parasit Vectors 11(1):29 PubMed PMC

Dubankova A, Horova V, Klima M, Boura E (2019) Structures of kobuviral and siciniviral polymerases reveal conserved mechanism of picornaviral polymerase activation. J Struct Biol 208(2):92–98 PubMed

Hercik K, Brynda J, Nencka R, Boura E (2017) Structural basis of Zika virus methyltransferase inhibition by sinefungin. Arch Virol 162(7):2091–2096 PubMed

Kabsch W (2010) Xds. Acta Crystallogr D Biol Crystallogr 66(Pt 2):125–132 PubMed PMC

Liebschner D, Afonine PV, Baker ML, Bunkoczi G, Chen VB, Croll TI, Hintze B, Hung LW, Jain S, McCoy AJ, Moriarty NW, Oeffner RD, Poon BK, Prisant MG, Read RJ, Richardson JS, Richardson DC, Sammito MD, Sobolev OV, Stockwell DH, Terwilliger TC, Urzhumtsev AG, Videau LL, Williams CJ, Adams PD (2019) Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix, vol 75. Acta Crystallographica Section D-Structural Biology, pp 861–877 PubMed PMC

Emsley P, Lohkamp B, Scott WG, Cowtan K (2010) Features and development of Coot. Acta Crystallogr D Biol Crystallogr 66(Pt 4):486–501 PubMed PMC

Noble CG, Li SH, Dong HP, Chew SH, Shi PY (2014) Crystal structure of dengue virus methyltransferase without -adenosyl–methionine. Antiviral Res 111:78–81 PubMed PMC

Zhang C, Feng T, Cheng J, Li Y, Yin X, Zeng W, Jin X, Li Y, Guo F, Jin T (2016) Structure of the NS5 methyltransferase from Zika virus and implications in inhibitor design. Biochem Biophys Res Commun PubMed