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JP-2026075092-A - Vaccinia virus polymerase-mediated viral replication

JP2026075092AJP 2026075092 AJP2026075092 AJP 2026075092AJP-2026075092-A

Abstract

[Problem] To provide a method for regulating the activity of poxvirus viral polymerase in cells infected with poxvirus. [Solution] Methods and compositions for modulating the activity of poxvirus viral polymerase by modulating the assembly and/or interaction of one or more subunits of viral polymerase are described. [Selection Diagram] Figure 3A

Inventors

  • サーレイ，アラダー・エイ

Assignees

  • イミュノルクス インターナショナル コーポレーション

Dates

Publication Date
20260507
Application Date
20251205
Priority Date
20191211

Claims (20)

  1. A method for regulating the activity of poxvirus viral polymerase in cells infected with a poxvirus, comprising the step of contacting the cells with a compound that reduces or inhibits the interaction between viral polymerase and glutamine tRNA (tRNA Glu ).
  2. The method according to claim 1, wherein tRNA Glu is uncharged tRNA Glu .
  3. The method according to claim 1 or 2, wherein the poxvirus is smallpox virus or a variant thereof.
  4. The method according to any one of claims 1 to 3, wherein the compound comprises a small molecule, antisense RNA, an antibody, an aptamer, or a polypeptide.
  5. The method according to any one of claims 1 to 4, wherein the viral polymerase is a virus-encoded RNA polymerase.
  6. The method according to claim 5, wherein the viral polymerase is a multi-subunit RNA polymerase (vRNAP) encoded by the virus.
  7. The method according to any one of claims 1 to 6, wherein the cells are stem cells, immune cells, or cancer cells.
  8. The method according to claim 7, wherein the stem cells are selected from adult stem cells, embryonic stem cells, fetal stem cells, mesenchymal stem cells, neural stem cells, totipotent stem cells, pluripotent stem cells, multipotent stem cells, oligopotent stem cells, unipotent stem cells, adipocytes, endothelial stem cells, induced pluripotent stem cells, bone marrow stem cells, umbilical cord blood stem cells, adult peripheral blood stem cells, myoblast stem cells, young stem cells, dermal fibroblast stem cells, and combinations thereof.
  9. A method for treating or preventing poxvirus infection in a subject requiring treatment or prevention of poxvirus infection, wherein the poxvirus comprises a viral polymerase, and the method comprises the step of administering to the subject a compound that reduces or inhibits the interaction between the viral polymerase and glutamine tRNA (tRNA Glu ).
  10. The method according to claim 9, wherein tRNA Glu is uncharged tRNA Glu .
  11. The method according to claim 9 or 10, wherein the poxvirus is smallpox virus or a variant thereof.
  12. The method according to any one of claims 9 to 11, wherein the compound comprises a small molecule, antisense RNA, an antibody, an aptamer, or a polypeptide.
  13. The method according to any one of claims 9 to 12, wherein the viral polymerase is a virus-encoded RNA polymerase.
  14. The method according to claim 13, wherein the viral polymerase is a multi-subunit RNA polymerase (vRNAP) encoded by the virus.
  15. A method for modulating the activity of poxvirus viral polymerase in cells infected with a poxvirus, comprising the step of contacting the cells with glutamine, wherein glutamine modulates the interaction between viral polymerase and glutamine-tRNA (tRNA Glu ).
  16. The method according to claim 15, wherein the poxvirus is smallpox virus or a variant thereof.
  17. The method according to claim 15, wherein the poxvirus is a vaccinia virus or a variant thereof.
  18. The method according to any one of claims 15 to 17, wherein glutamine reduces or inhibits the interaction between viral polymerase and tRNA Glu .
  19. The method according to any one of claims 15 to 17, wherein glutamine increases or promotes the interaction between viral polymerase and tRNA Glu .
  20. The method according to any one of claims 15 to 19, wherein the viral polymerase is a virus-encoded RNA polymerase.

Description

Cross-reference of related applications [0001] This application claims the benefit of U.S. Provisional Application No. 62/946,828, filed December 11, 2019, which is incorporated in whole for all purposes. References to "sequence listings," tables, or computer program listings appendices submitted as ASCII files. [0002] The sequence listing written in the file 055523-504001WO_SequenceListing_ST25.txt, 4,096 bytes, machine format IBM-PC, MS Windows operating system, created on 11 December 2020, is incorporated herein by reference. [0003] The nucleus of eukaryotes contains the mechanisms of DNA replication and gene transcription. Numerous viruses depend on host cell factors for their replication and transcription, and therefore require at least a transient intranuclear phase to ensure viral proliferation. A notable exception among eukaryotic DNA viruses is the poxviridae family, where replication and transcription are confined to the cytoplasm (Moss, 2013). These processes require virally encoded factors to produce mature mRNA from the viral genome. [0004] The Poxviridae family includes the smallpox virus (varicella) and the vaccinia virus (varicella vaccine). While natural varicella was declared eradicated worldwide in 1980, the risk remains that the varicella virus, or its variants, could be used as agents of bioterrorism. Furthermore, the vaccinia virus is being studied as a potential cancer treatment (e.g., a tumor-disintegrating virus). [0021] This figure shows the measured total integrated intensity of CV-1 cells over time during the glutamine experiment. The x-axis represents the time since infection in time, and the y-axis represents the total integral. Error bars represent the calculated standard error. "+" and "-" represent the presence or absence of glutamine during the first medium change, respectively.[0022] This figure shows the measured total integrated intensity of CV-1 cells over time during the glutamine experiment. The x-axis represents the time since infection in time, and the y-axis represents the total integral. Error bars represent the calculated standard error. "+" and "-" represent the presence or absence of glutamine between the second medium changes, respectively.[0023] This figure shows the measured total integrated intensity of CV-1 cells over time during the glutamine experiment. The x-axis represents the time since infection in time, and the y-axis represents the total integral. Error bars represent the calculated standard error. "+" and "-" represent the presence or absence of glutamine during the third medium exchange, respectively.[0024] This figure shows the percentage viral titer of each sample compared to sample +/+/+. Error bars represent the standard deviation. Statistically significant differences based on triplicates against the positive control +/+/+ (Student's t-test, p < 0.05) are marked with an asterisk.[0025] Figure 3A shows a schematic representation of vRNAP EC. The subunits are colored as shown, as in Grimm et al., 2019. The helix is shown as a cylinder. Nucleic acids are shown in blue (template DNA), cyan (non-template DNA), and red (RNA). Metal ions are shown as spheres. Figure 3B shows a magnified view of the active site of vRNAP. Proteins and nucleic acids are shown as sticks and are colored as in Figure 3A. Cryo-EM density is shown as a gray mesh. The vRNAP EC is in the post-translocation state, and the +1 template base is ready to base pair with the incoming nucleotide. Residues specific to vRNAP discussed in the study are highlighted in green. Figure 3C shows a schematic diagram of the nucleic acid scaffold used in this study. Individual bases are shown as circles, and bases are abbreviated as single-letter codes. Bases that can exist in the EC structure are shown as solid circles, and invisible bases are shown as hollow circles. The active site metal A is shown as a pink sphere. vRNAP residues within a distance of 4 Å from the nucleic acid are shown, and S. cerevisiae Pol The data is colored according to its storage method in II. Residues specific to the vRNAP discussed herein are highlighted in green. See also Figures 10, 11, and 12.Figure 3A shows a schematic representation of vRNAP EC. The subunits are colored as shown, as in Grimm et al., 2019. The helix is shown as a cylinder. Nucleic acids are shown in blue (template DNA), cyan (non-template DNA), and red (RNA). Metal ions are shown as spheres. Figure 3B shows a magnified view of the active site of vRNAP. Proteins and nucleic acids are shown as sticks and are colored as in Figure 3A. Cryo-EM density is shown as a gray mesh. The vRNAP EC is in the post-translocation state, and the +1 template base is ready to base pair with the incoming nucleotide. Residues specific to vRNAP discussed in the study are highlighted in green. Figure 3C shows a schematic diagram of the nucleic acid scaffold used in this study. Individual bases are shown as circles, and bases are abbreviated as single-letter codes. Bases